Sickle Cell Disease by the HbS haplotype

HbS of African derivation...some parties almost always associate its presence in non-African locations, or else what is perceived to be "non-Sub-Saharan" Africa locations, with historic slavery. How accurate is this observation?

Before one gets into the details about the issue of legitimacy of the above mentioned perception, it is perhaps necessary to know some basic things about HbS:

It is essentially an altered beta-globin (b-globin) cluster of the Hemoglobin A, a cluster which is located at position 15.5 away from the centromere of chromosome 11 on its short arm; the specific alteration that gives rise to HbS instead of the "normal" HbA, is a "point mutation" [*1] in GAC codon at position 6 of the b-globin cluster, which converts GAC to GTG, resulting in the replacement of glutamic acid amino acid by valine. Several haplotypes are associated with this mutation; there are supposed to be some 475 identified allele variants of the b-globin gene, of which the HbS haplotypes are a subset. This includes the African derivatives, namely; the Senegalese, Benin and Bantu haplotypes. Outside of African derivatives, there is the Asian haplotype, mainly found in Arabia and the Indian sub-continent.

Now, since it is difficult to age HbS variants with respect to one another, due to selective pressure acting on the extent of its availability, the distribution pattern becomes worthy of noting, as well as geographical origin. Apparently, the African haplotypes have the highest prevalence among African subjects and recent African descendants. Of the African variants, the Benin haplotype appears to have wider geographical reach, perhaps because it originated in the vicinity of the Niger River Valley, and spread through ancient contact in the Sahara. So, treating African HbS haplotypes unanimously as the product of slave trade is not only simplistic, but also erroneous. Consider the fact for example, that there are locations where African slaves had historically been brought as "pick up points" for trade between the trading parties, after having been picked up from their home region in some other geographical location, but are nonetheless virtually free from the sickle cell disease; the African Horn comes to mind as an example, and yet HbS has hardly penetrated that region. Some may be tempted to attribute this phenomenon to the environment of that region, but that assessment would be oblivous to the fact that Bantu-speaking populations neighbouring the populations of these regions, have high prevalence cases of HbS variants [i.e. Bantu haplotype]. This point is especially relevant, when its presence in Egypt is merely explained off as the product of that region being a historic corridor for slave trade. Additionally, it is necessary to note documented cases of HbS outside of sub-Saharan Africa, in order to realize that the antiquity of its heritage in those regions precedes historic slave trade involving slaves brought from sub-Saharan Africa. For examples, we have the following:

Haplotypes of the beta-globin gene as prognostic factors in sickle-cell disease.

el-Hazmi MA, Warsy AS, Bashir N, Beshlawi A, Hussain IR, Temtamy S, Qubaili F.Medical

Biochemistry Department, World Health Organization Collaborating Centre for Haemoglobinopathies, Thalassaemias and Enzymopathies, College of Medicine, King Khalid University Hospital, Riyadh, Saudi Arabia.

Objective
This study was conducted with two objectives: to determine the b-globin gene haplotypes associated with Hb S in Arabic-speaking populations from different countries (Egypt, Syrian Arab Republic and Jordan), and to compare the results with those of Saudi SCD patients from different regions of Saudi Arabia, where both mild and severe forms of the disease exist.

Patients and methods
The study group of 126 SCD patients comprised 14 Egyptians, 9 Syrians, 10 Jordanians and 93 Saudis. Some of the Egyptians, Syrians and Jordanians were living in Riyadh, while others were living in their own countries, from where buffy coats were received. The Saudis were from three areas of Saudi Arabia where the sickle-cell gene has been reported at a high frequency, these being eastern (22 patients), south-western (67) and north-western (4) regions...

Buffy coat was used to extract DNA [18]. Portions of DNA were first subjected to polymerase chain reaction (PCR) amplification, and the PCR product was restricted using Ava II, HindIII, HincII, Hpa I and Xmn I, following the procedure published earlier [19,20]. The presence or absence of each site (shown in Figure 1) was identified, based on the size of the fragments produced (Table 1).

Results
"We collaborated with researchers from Egypt, Syrian Arab Republic and Jordan in a study of patients with sickle-cell disease from those countries, and from various parts of Saudi Arabia, in order to investigate the influence of genetics on the clinical presentation of the disease, and to attempt to determine the **origin** of the sickle-cell gene in Arabs. Our results suggest that beta-globin gene haplotypes influence the clinical presentation of sickle-cell disease, and that there are at least two major foci for the origin of the sickle-cell gene, one in the eastern part of Saudi Arabia, and the other in the populations of North Africa and the north-western part of the Arabian peninsula…

The Benin haplotype was found in patients with severe disease, either as homozygous or in combination with another haplotype. The majority of Syrians and Jordanians had the Benin haplotype, and severe disease. However, one in three Syrians and one in five Jordanians had a milder disease, and the Saudi-Indian haplotype was identified.

All Saudi patients from south-western and north-western areas, where the disease is generally severe, had the Benin haplotype in the homozygous or heterozygous state. Of the Saudi patients from the eastern area, where a mild form of SCD exists, only 9% had the Benin haplotype. The remainder had the Saudi-Indian haplotype, either in its homozygous or heterozygous state…

Restriction endonuclease restriction sites have provided a useful insight into the normal polymorphic variations in the DNA surrounding various gene loci, where a combination of two or more polymorphic sites has led to the identification of specific haplotype patterns [13,14]. This has been of significance in the study of the regions surrounding the b-globin gene (i.e. the b-globin gene cluster), where several polymorphic sites have been identified, and population differences have been found on analysis of the haplotype pattern [9]. An interesting observation is that the sickle-cell mutation has occurred on chromosomes carrying different polymorphic sites and different b-globin gene haplotypes, and this seems to play a role in the clinical expression of SCD [9].

We compared the haplotype pattern of SCD patients from different Arabic-speaking countries. Benin haplotype was the major haplotype in all countries with a severe presentation of SCD and it was present in both the homozygous and heterozygous state. This was true for those SCD patients from south-western and north-western areas of Saudi Arabia, and for those from Egypt, Jordan and Syrian Arab Republic. On the other hand, patients from the eastern part of Saudi Arabia, who present with a significantly milder clinical picture, carried the Saudi-Indian b-globin gene haplotype either in its homozygous or heterozygous state."

Source: East Mediterr Health J. 1999 Nov;5(6):1154-8 http://www.emro.who.int/Publications/EMHJ/0506/10.htm

From the above, one gets the impression that the Benin haplotype is dominant over the other haplotypes wherein it appears in a heterozygous case. Not sure how that specifically goes on a case by case basis, wherein the Benin haplotype would appear with the other African variants, but at the least, this study demonstrates that it is the more likely dominant haplotype over that of the Asian haplotypes; the present author takes this to be the case [with the understanding that from the African side, only the Benin haplotype has notably penetrated the regions under discussion, while the others mentioned are of the Asian type], because it has always been identified with the severe cases of SCD in not only the homozygous cases, but also in the heterozygous cases...that is to say, what are the chances of heterozygous cases, especially in areas like eastern Saudi where the Asian haplotype is the prevalent HbS haplotype, being inclusive of Benin-Asian haplotype pairings? Quite likely—albeit to varying degrees, notwithstanding the discernable distribution patterns of these haplotypes as mentioned in the study. On the other hand, as noted, and to reiterate:

one in three Syrians and one in five Jordanians had a milder disease, and the Saudi-Indian haplotype was identified.

...this even when it occurred in either homozygous or heterozygous cases, not involving the Benin haplotype [which consistently appeared in severe cases of SCD, regardless of whether it was a heterozygous or homozygous case].

Another noteworthy case, is that SCD appears in the remains of predynastic subjects, and the only HbS haplotype noted in Egypt today is the Benin haplotype. As we will shortly see, the Benin haploype also accounts for all the known HbS cases outside of Africa, save for the Asian haplotype, largely confined to eastern Saudi Arabia and the Indian sub-continent.

Use of the amplification refractory mutation system (ARMS) in the study of HbS in predynastic Egyptian remains.

Marin A, Cerutti N, Massa ER.

1999 May-Jun

Dipartimento di Biologia Animale e dell'Uomo, Università degli Studi di Torino.

We conducted a molecular investigation of the presence of sicklemia in six predynastic Egyptian mummies (about 3200 BC) from the Anthropological and Ethnographic Museum of Turin. Previous studies of these remains showed the presence of severe anemia, while histological preparations of mummified tissues revealed hemolytic disorders. DNA was extracted from dental samples with a silica-gel method specific for ancient DNA. A modification of the polymerase chain reaction (PCR), called amplification refractory mutation system (ARMS) was then applied. ARMS is based on specific priming of the PCR and it permits diagnosis of single nucleotide mutations. In this method, amplification can occur only in the presence of the specific mutation being studied. The amplified DNA was analyzed by electrophoresis. In samples of three individuals, there was a band at the level of the HbS mutated fragment, indicating that they were affected by sicklemia. On the basis of our results, we discuss the possible uses of new molecular investigation systems in paleopathological diagnoses of genetic diseases and viral, bacterial and fungal infections.

PMID: 11148985 [PubMed - indexed for MEDLINE]

The interesting thing about the study, is that it too talks of severe cases of SCD, which as we've just seen, is the case with the Benin haplotype as well, and it takes us to a period that precedes any known documentation of slave trade involving sub-Saharan Africans.

Let's take a look at the distribution pattern and type of HbS haplotypes present in global samples:

The Benin haplotype accounts for HbS associated chromosomes in Sicily,4 Northern Greece,10 Southern Turkey,11 and South West Saudi Arabia,6,7 suggesting that these genes had their origin in West Africa. The Asian haplotype is rarely encountered outside its geographic origin because there have been few large population movements and Indian emigrants have been predominantly from non HbS containing populations. However, it is of interest that the Asian haplotype was first described among descendants of Indian indentured laborers in Jamaica.12 — Graham R. Serjeant, MD, FRCP, MRC Laboratories (Jamaica), University of the West Indies, Kingston.

The African type in the above regions appear to be exclusively the Benin haplotype, in contrast to...

From these original foci of the HbS mutation, the gene spread along trading routes to North Africa and the Mediterranean, was transported in large populations to North and South America and the Caribbean during the slave trade, and latterly has spread to Northern Europe by immigration from the Caribbean, directly from Africa to the United Kingdom, France, Belgium, and Holland, and from Turkey to Germany. The relative prevalence of these haplotypes in the Americas reflects the different origins of their African peoples, approximately 70% of HbS associated chromosomes having the Benin haplotype, 10% Senegal and 10% Bantu. Haplotype frequencies in Jamaica are similar to the USA but the Bantu haplotype accounts for the majority of HbS associated chromosomes in Brazil.9 — Graham R. Serjeant, MD, FRCP, MRC Laboratories (Jamaica), University of the West Indies, Kingstom.

It is of no coincidence that the other African haplotypes outside of the Benin haplotype, have found their way into the Americas, regions that have been affected by the historic slave trade, and have large size settler populations of recent common ancestry from sub-Saharan Africa.

What all these show, when taken together, is that blanket attribution of African HbS variants solely to historic slave trade events is a blatant distortion of reality.

Ps - A good map showing the geographical range of HbS haplotype variants:

Lucotte et al.'s haplotype IV

So exactly what UEP marker is Lucotte et al.'s haplotype IV associated with?

Lucotte et al. don't exactly tell us which specific NRY monophyletic unit haplotype IV belongs to, but S.O.Y Keita clues us in on it:

Haplotype IV, designating the M2/PN1 subclade, as noted, is found in high frequency in west, central, and sub-equatorial Africa in speakers of Niger-Congo—which may have a special relationship with Nilosaharan—spoken by Nubians; together they might form a superphylum called Kongo-Saharan or Niger-Saharan (see Gregersen 1972, Blench 1995), but this is not fully supported. The spatial distribution of p49a,f TaqI haplotypes in the geographically-widespread speakers of Nilosaharan languages has not been fully characterized, but the notable presence of haplotype IV in Nubians speaking the Eastern Sudanic branch is interesting in that this subgroup is in the Sahelian branch of speakers, whose ancestors may have participated in the domestication of cattle in the eastern Sahara (Ehret 2000, Wendorf and Schild 2001). Sometimes haplotype IV (and the M2 lineage) is seen as being associated with the “Bantu expansion” (2000-3000 bp), but this does not mean that it is not much older, since expansion and origin times cannot be conflated. Haplotype IV has substantial frequencies in upper Egypt and Nubia, greater than VII and VIII, and even V. Bantu languages were never spoken in these regions or Senegal, where M2 is greater than 90 percent in some studies. — Keita, 2005.

Aside from referencing various Lucotte et al. publications from 1996 through to 2003, and relating their findings on RFLP markers to those of Al Zahery et al. 2003, the frequency and distribution pattern of haplotype IV no doubt reinforced Keita's perception of its link with E3a bearing chromosomes. Keita tabulated the frequencies of various RFLP haplotypes across the samples studied in the aforementioned works, and it can be viewed here:

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Haplotypes and percentages

                                   IV    V      XI     VII   VIII  XII   XV

Falasha (38)                       0.0   60.5   26.3   0.0   0.0   0.0   0.0

Ethiopians [non Falasha](104)      0.0   40.4   25.9   0.0   23.1  0.0   0.0

Berbers (74)                       1.4   68.9   2.8    1.4   6.8   4.1   0.0
(Morocco)

“Sephardic” Jews (381)             8.4   18.6   6.8    19.9  34.1  4.2   2.1

“Oriental” Jews(56)                1.8   8.9    0.0    7.1   78.6  0.0   1.8

“Near Eastern” (27)                0.0   7.4    0.0    7.4   85.1  0.0   0.0

Askenazic Jews(256)                0.0   3.1    15.2*  22.7  24.6  9.0   10.9

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1 - Lucotte and Mercier (2003a)
2 - Lucotte and Mercier (2003b)
3 - Al-Zahery et al. (2003); *haplotype XI here is documented from two biallelic lineages
4 - Lucotte et al. (2000)
5 - Lucotte and Smets (1999)

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country (n)                     Haplotypes and percentages

                                IV    V      XI    VII   VIII   XII   XV

Egypt(274)                      13.9  39.4   18.9  6.6   7.3    2.2   5.5

Lebanon(54)                     3.7   16.7   7.4   20.4  31.5   5.6   1.9

Palestine(69)                   1.4   15.9   5.8   13.0  46.4   0.0   4.3

Iraq(139)                       1.4   7.2    6.4*  20.1  36.0   1.4   0.7

Egypt(52)                       7.7   40.4   21.2  9.6   7.7    3.8   1.9

Libya (38)                      7.9   44.7   10.5  0.0   5.3    13.2  0.0

Algeria (141)                   8.5   56.7   5.0   1.4   7.1    4.2   5.0

Tunisia (73)                    0.0   53.4   5.5   4.1   2.7    26.0  2.7

Morocco (102)                   0.98  57.8   8.8   4.9   7.8    0.98  10.8

Mauretania (25)                 8.0   44.0   8.0   0.0   4.0    0.0   0.0

Suprasah(composite)(505)        4.4   55.0   7.7   3.2   6.3    7.1   4.2

Ethiopia(composite)(142)        0.0   45.8  26.1   0.0   16.9   0.0   0.0

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6 - Lucotte et al. (1996)

— References 2, 4, 5, as in Table 2A.

— *Haplotype XI in groups admixed with northern Europeans is usually affiliated with haplogroup R1; in Africa it is usually associated with haplogroup E (al-Zahery 2003).

The distributions of haplotype 4 in those tables make perfect sense, if they were strongly associated with Hg E3a chromosomes.

As the present author has noted elsewhere, an apparent limitation of just relying on RFLP markers lies with the high probability for re-occurrence of the same RFLP sequences in two, if not more, distinct haplogroups. However, lest one thinks that Keita is alone in his observation, about linking haplotype IV to E3a bearing chromosomes, then consider this:

A total of 21 different 49a,f haplotypes were found and are illustrated in Fig. 4 as a sub-classification of the Iraqi Y-chromosome haplogroups. The most represented haplotype of haplogroup E is haplotype 5 (A 2 C 0 D 0 F 1 I 1 ). This is followed by haplotype 11 (A 3 C 0 D 0 F 1 I 1 ) at a much lower frequency. Haplotypes 5 and 11 were observed both in Africa (Lucotte et al., 2001; Passarino et al., 1998; Persichetti et al., 1992; Santachiara-Benerecetti and Semino, 1996; Spurdle and Jenkins, 1992; Torroni et al., 1990) and Eurasia (Passarino et al., 2001; Semino et al., 2000b) but in Africa they belong to haplogroup E, whereas in Eurasia, particularly in *Northeastern Eurasia*, they belong mainly to the haplogroup R-M17. Interestingly, the proportion of haplotypes 5 and 11 in haplogroups E and R-M17 is reversed, with haplotype 5 prevalent in haplogroup E and haplotype 11 in haplogroup R-M17. By considering that the two haplotypes differ by a single band change and their different proportion in the two lineages, it is likely that haplotype 11 is a derivative of haplotype 5 in haplogroup E and just the opposite in haplogroup R-M17.

And...

It is worth mentioning that in haplogroup E two subjects belong to the African specific E-M2 clade, which is very frequent in the Western and Southern part of the continent (Cruciani et al., 2002; Passarino et al., 1998; Scozzari et al., 1999; Seielstad et al., 1994; Semino et al., 2002; Underhill et al., 2000) and has been related to the Bantu expansion. These two Y chromosomes **harbor haplotype 4** (A 1 C 0 D 0 F 1 I 1 ) which is also African- specific and **shows the same geographic distribution** (Excoffier et al., 1987; Passarino et al., 1998; Spurdle and Jenkins, 1992; Torroni et al., 1990). Within haplogroup J, haplotypes 7 (A 2 C 0 D 1 F 1 I 0 ) and 8 (A 2 C 0 D 1 F 1 I 1 ) are the most represented, but haplotype 7 is observed only in the J-M172 sub-set. This suggests that the 49a,f haplotype 7 arose on a 12f2-8Kb/M172 Y chromosome. — N. Al Zahery et al. 2003, Y-chromosome and mtDNA polymorphisms in Iraq, a crossroad of the early human dispersal and of post-Neolithic migrations.

To understand how Al Zahery et al. were able to link RFLP haplotypes to SNP markers on the chromosomes under study, it is worth noting that they identified the former by using TaqI restriction enzyme digests and the latter with the likes of PCR and DHPLC analysis; essentially this sort of approach to testing for both RFLP markers and binary markers has been exemplified in the following link—we discussed the methods utilized in relative detail, in the sorting out of RFLP markers into respective sub-clades: RFLPs: Lucotte et al., A case study — Pt. 1 [clickable]

Therefore, E3a in the Nile Valley?

Getting back to Keita's linking of haplotype IV to Hg E3a chromosomes, and as verified above in the Al Zahery et al. study; some parties are perplexed by the detection of the rather considerable frequencies of Hg E3a in the Nile Valley, as denoted by the distribution and frequency patterns of haplotype IV in Lucotte et al.'s study, i.e. if haplotype IV is to be unequivocally accepted as a marker characteristic of Hg E3a-bearing chromosomes - again, as maintained by Keita and Al Zahery above, for instance. Even Lucotte et al. themselves had placed haplotype IV in a context not inconsistent with that maintained by Al Zahery et al. and Keita, for they say:

Haplotype IV, characteristic of sub-Saharan populations, shows a southern geographic distribution in Egypt. — G. Lucotte and G. Mercier

...and they also refer to it as "African haplotype" elsewhere, in contrast to:

Haplotype V is a characteristic Arab haplotype, with a northern geographic distribution in Egypt in the Nile River Valley. — G. Lucotte and G. Mercier, Y-chromosome haplotypes in Egypt, 2002.

If one recalls, it has already been noted that "Arab" here, as used by Lucotte and Mercier, is in reference to "Arabized" north Africans; it is necessary to realize this, for it has the potential of misleading those in the little know.

It is necessary to note, that while haplotype IV has considerable presence in the Upper Nile Valley, which Lucotte and Mercier refer to as "Upper Egypt" and "Lower Nubia" respectively, its average frequency still follows that of haplotype V and haplotype XI, both of which have been linked to Hg E3b-bearing chromosomes:

We analyzed Y-chromosome haplotypes in the Nile River Valley in Egypt in 274 unrelated males, using the p49a,f TaqI polymorphism. These individuals were born in three regions along the river: in Alexandria (the Delta and Lower Egypt), in Upper Egypt, and in Lower Nubia. Fifteen different p49a,f TaqI haplotypes are present in Egypt, the three most common being haplotype V (39.4%), haplotype XI (18.9%), and haplotype IV (13.9%).

Haplotype IV has a gradient that decreases as one moves from the so-named Upper Nile Valley regions to Lower Egypt, with higher frequencies in "Upper Egypt" and "Lower Nubia" and lower frequencies in "Lower Egypt". Nonetheless, does haplotype IV's presence in Lower Egypt mean that Hg E3a has been detected therein? Well, if Luis et al.'s work of "the Nile Valley corridor vs. the African Horn" is anything to go by, this should come as no surprise; they show that low frequencies of Hg E3a occur in "northern Egypt", which in itself should be instructive, considering that this is the region of the Nile Valley relatively further away from sub-Saharan Africa [where this haplotype predominates]. So, if lower Egypt can show low frequencies of E3a, as verified by Luis et al., then does it not follow that Upper Egypt could and would have this marker, in relatively higher frequencies?

The aforementioned perplexion that the present author has come across, concerning this E3a presence, was presumably justified by the idea that few other studies that have been done on samples taken from Upper Egypt and Sudan, little to no Hg E3a had been documented. Now of course, the results of samplings can be affected by choice of sampling, in terms of its ethnic diversity and geographical range, the type and number of markers typed and methods used to amplify or detect them. The present author recalls a discussion wherein he was told that Arredi et al.'s study, "A Predominantly Neolithic Origin for Y-Chromosomal DNA Variation in North Africa", was one such study which studied samples from the Upper Nile Valley of Egypt, but found no E3a markers therein. It goes back to the sample issues just mentioned, and this is what the present author had to say about that issue:

"Arredi et al.'s study primarily focuses on E3b and J lineages, not E3a. Their samples, as far as I know consist of candidates from two locations: 44 candidates from Mansoura and 29 from Luxor. Of these samples, 14 binary markers were identified in the 44 candidates from Mansoura, while the 37 identifications were done by tandem repeats, and likewise, of the 29 candidates, 9 binary markers was identified amongst them while 27 markers were defined only by tandem repeats. It is also of interest that this study also shows E3a detection in the Ethiopian sample. Whereas in the Luis et al. sample, more diverse speaking groups in northern Egypt were sampled, namely Arabic and Tamazight speakers. So it is the question of sampling selection, sampling size, and ability to detect as much binary markers along with STR markers as possible."

Yet, what's interesting in all this, is that the individual who raised the issue of lack findings of Hg E3a in the aforementioned Arredi et al. study, didn't and perhaps was not capable of demonstrating that RFLP haplotype IV corresponds to any other but Hg E3a chromosomes. To top that, the frequency and distribution pattern of haplotype IV—which would make sense if it is associated with E3a, in terms of sporting the aforementioned decreasing south-to-northward gradient, was not taken into consideration; nor was the correspondence between the frequency of haplotype IV in Lower Egypt, as noted by Lucotte et al., and that of Hg E3a in Lower Egypt, as noted by Luis et al. (2004), taken into consideration.

As it stands, pending new information to the contrary, haplotype IV is linked to E3a bearing chromosomes, not to mention the other peculiarities specific to and shared by haplotype IV and Hg E3a, and as such, one would have to concede that Hg E3a is present in the Nile Valley [and there is no doubt about that], with a south-to-north gradient showing higher frequencies in the Upper Nile Valley, that progressively decreases as one goes further down the Nile, and finally reaching the lowest frequencies in northern Egypt.

Last but not least, of note:

One Y-specific DNA polymorphism (p49/Taql) was studied in a sample of 469 African males coming from twelve populations of sub-Saharan Africa. An high frequency (62.5%) of the Y-haplotype IV was observed in these populations, the most elevated percentage of this haplotype being observed in Mossis (from Burkina-Fasso). The “Arabic” haplotype V is present in these populations at a mean frequency of 8.7%. The “oriental” haplotype XI is present at a mean frequency of 11.3%, the most elevated percentage of this haplotype being observed in Songhaiis (from Niger).—Lucotte G. and Gérard N. 2000, Haplotypes of the Y chromosome in some populations of west Africa
___________________________________________________________
Brief references:

*As noted by Keita 2005, Explanation of the Pattern of P49a,f TaqI RFLP Y-Chromosome Variation in Egypt:

1 - Lucotte and Mercier (2003a)
2 - Lucotte and Mercier (2003b)
3 - Al-Zahery et al. (2003); *haplotype XI here is documented from two biallelic lineages
4 - Lucotte et al. (2000)
5 - Lucotte and Smets (1999)
6 - Lucotte et al. (1996)

*Lucotte et al.; North African genes in Iberia studied by Y-chromosome DNA haplotype 5; 2001.

*G. Lucotte and G. Mercier, Y-chromosome haplotypes in Egypt, 2002.

*N. Al Zahery et al. 2003, Y-chromosome and mtDNA polymorphisms in Iraq, a crossroad of the early human dispersal and of post-Neolithic migrations.

*Luis et al. 2004, The Levant versus the Horn of Africa: Evidence for Bidirectional Corridors of Human Migrations.

*Arredi et al. 2004, A predominantly neolithic origin for Y-chromosomal DNA variation in North Africa.

RFLPs: Lucotte et al., A case study — Pt. 2

Continuing with Lucotte et al. 2006, North African Berber and Arab Influences in the Western Mediterranean Revealed by Y-Chromosome DNA Haplotypes.

Link to the first part of this topic:
RFLPs: Lucotte et al., A case study — Pt. 1[clickable]

There is the question of haplotype V corresponding to more than one binary marker—which is why it is important for one to understand the *context* of the specific study in question, rather than assume that one group of researchers' context per designator is the same as the other. Many a times, researchers use a standardized designation like the Y Chromosome Consortium nomenclature, applying the same context to facilitate cross-reference between researchers, and hence, compare the results at hand, but not every researcher adheres to the most popular standardized nomenclature system—some researchers use one or another system of nomenclature, dependent on the methods applied to discern haplotypic differentiation.

From what the present author can tell, Lucotte et al. didn't run their haplotypes for binary markers for the earlier studies, as exemplified in their 2001 study in "A case study — Pt. 1", but given that Lucotte et al. this time around [i.e. in the present study] proclaimed to have not only used the same sequencing methods used by them in the earlier studies [namely the 2001 and 2003 studies for example], but also the DNA polymerase chain reaction used by Underhill et al. to test for binary markers M81 and M78, presumably using samples already tested by Lucotte et al. in previous studies, they were able to verify that Lucotte et al.'s 2001 and 2003 haplotype V [which they deemed to be 'a characteristic Berber haplotype' and ' of predominantly Berber origin'] were mainly M81 chromosomes, after having further discerned haplotype V into at least two discernable groups: Vb and Va, using Gonçalves et al.'s PCR method. All the Moroccan bearers of the sub-group Vb, turned out to be M81 carriers, including *all* the 'Berber' designated speakers of this bunch and 21 of the 59 'Arab' designated North Africans; the remaining 38 'Arab' designated north African bearers of the sub-group Va either tested positive for M78 or they didn't. To be specific, 31 of the 38 Arabic speaking North African bearers of haplotype V [sub-group Va] tested positive for M78, while the rest didn't. This results confirm the sub-group Vb to be M81, and Va to be partly M78. Going by this, the majority of what Lucotte et al. refer to as a 'characteristic Berber haplotype', are M81 chromosomes.

In that Lucotte et al.'s haplotype V appears to be largely of M81 [all Vb subgroups in the intro study] and partly M78 [sub-group Va], with the remaining [sub-group Va] yet to be identified by other markers, it is obviously associated with E3b macrohaplogroup. Additionally, according to Keita, haplotype XI also turns out to be Hg E3b affiliated. Thus, alerting oneself to the context at hand, is warranted!

As to the question of whether the authors in the present study were able to genetically differentiate Moroccan "Berbers" and "Arabs" into distinct genealogical camps, the answer is that they were probably going by self-ethnic identifications of the contributors of the samples in question. Apparently much of north African populations were really 'Arabized' populations, not original ethnic Arabs. However, as the present author noted before, there is something to be discerned here:

• All* subgroup Vb individuals are either "Berber" identified Tamazight individuals [47 individuals] or what appears to be "Arabized" Tamazight individuals [21 individuals], testing positive for E-M81, whereas...

• ...out of the 38 remaining "Arabic" identified individuals who tested positive for the subgroup Va, 31 turned out to be positive for E-M78. The remaining 7 individuals of this 38 "Arabic" identified individuals didn't test positive for either E-M81 or E-M78. Those individuals *could* [but not necessarily] possibly turn out positive for a non-M78 and non-M81 marker like "R1a"; however, **if** J happens to have the same combination of RFLPs , then it could well be a possible candidate here, as it is the next frequent paternal line which isn't M35 derived, but still quite less frequent than the aboriginal E-M81 marker. Other non-M81 and non-M78 E-M35 [E3b] derived lineages could just as well still be a candidate.

• Four Moroccan "Berber" individuals out of the 51 tested, appear to be unaccounted for here. They probably fell into the Va subgroup, and likely didn't test positive for either E-M81 or E-M78. These individuals could just as well test positive for the aforementioned non-M78 and non-M81 possible candidates.

In any case, non-M81 and non-M78 lineages appear to be in the minority. The majority of Moroccan "Berbers" are M81, while the majority of the Moroccan "Arabs" are largely of subgroup Va E-M78 lineage [31 individuals], but carry a significant amount of subgroup Vb E-M81 [21 individuals]. So clearly, the "Berber" group, while they share lineages [largely E-M81] with their Moroccan "Arab" counterparts, can clearly be discerned from the said "Arab" counterparts, in that the later seems to have relatively more M78 lineages, as well as relatively more non-M81 subgroup Va lineage [7 individuals] than their "Berber" identified counterparts [possibly the unaccounted for 4 "Berber" individuals].

RFLPs: Lucotte et al., A case study — Pt. 1

From Lucotte et al., we have:

North African Berber and Arab Influences in the Western Mediterranean Revealed by Y-Chromosome DNA Haplotypes

Nathalie Gérard, Sala Berriche, Annie Aouizérate, Florent Diéterlen, Gérard Lucotte. Human Biology. Detroit: Jun 2006. Vol.78, Iss. 3; pg. 307, 10 pgs

During the 7th century A.D., Muslim people coming from the Arabian peninsula and the Middle East invaded North Africa. The most important population movement relating both sides of the Mediterranean Sea was the conquest of the Iberian peninsula by North African populations (with recruited Berbers), soon after the first Muslim invasion. More than eight centuries (8th to 15th centuries) of Muslim domination in the southern part of Iberia imparted an important cultural legacy (Conrad 1998) and probable gene exchanges between North African and Iberian populations.

Variations in DNA sequences specific to the nonrecombinant part of the Y chromosome, relating to paternal ancestry, are particularly interesting from a human population genetics point of view. The first published and most informative probe used in Southern blots for this objective is p49 (locus DYSl), which is able to identify at last five TaqI male-specific fragments (A, C, D, F, and I) that are polymorphic between individuals (Lucotte and Ngo 1985). Sixteen main corresponding haplotypes (numbered I-XVI) were identified using the p49 probe on DNA samples of unrelated males living in France (Ngo et al. 1986). Only recently has the molecular basis of the p49 TaqI polymorphisms been established (Jovelin et al. 2003); the polymorphisms correspond to variable TaqI sites located in the four DAZ genes located in the AZF-c region of the Y chromosome.

In fact, the conventional p49 TaqI polymorphisms were the most popular markers used in various populations because of their ability to detect more than 100 different haplotypes [for a compilation on the subject until the end of 1995, see Poloni et al. (1997)]. Haplotype XV (A3,C1,D2,F1,I1) was the most widespread haplotype in our initial study (Ngo et al. 1986). Haplotype XV was also predominant in the first European study we published (Lucotte and Hazout 1996), with elevated frequencies in French Basques. The geographic distribution of haplotype XV in Europe reveals a gradient of decreasing frequencies from this Basque focus toward eastern peripheral countries (Lucotte and Loirat 1999) but also toward southwestern countries. According to the Y Chromosome Consortium (2002) nomenclature, haplotype XV corresponds to the M173 lineage (Diéterlen and Lucotte 2005).

Haplotype V (A2,C0,D0,F1,I1) is the most frequent haplotype in North Africa (Lucotte et al. 2000), with a particularly high frequency (55%) in the populations with a relative predominance of Berber origin. Our previous study on the subject examined the relative frequencies of haplotype V in four Iberian populations compared with a Berber population living in North Africa (Lucotte et al. 2001). The highest frequency of haplotype V (68.9%) was observed in Berbers from Morocco, and the geographic distribution of haplotype V revealed a gradient of decreasing frequencies with latitude in Iberia (40.8% in Andalusia, 36.2% in Portugal, 12.1% in Catalonia, and 11.3% in the Basque Country) (Lucotte et al. 2001); such a cline of decreasing haplotype V frequencies from the south to the north in Iberia clearly established a gene flow from North Africa toward Iberia.

According to the Y Chromosome Consortium (2002) nomenclature, haplogroup E is characterized by the mutations SRY4064, M96, and P29 on a background defined by the insertion of an Alu element (YAP + ). The third clade, E3 (defined by the mutation P2), of haplogroup E is further subdivided into two monophyletic forms, the second one (E3b) being characterized by mutations M35 and M125. All of the 110 p49 TaqI haplotype V subjects from Morocco (51 Berbers and 59 Arabs) that we had previously tested correspond to haplogroup E3b.

In the present study we have subdivided haplotype V into its Berber (Vb) and Arab (Va) components in order to distinguish the relative contributions of these two ethnicity-specific markers in the gene pools of the populations living in Iberia and in other populations in the northern part of the western Mediterranean area.

Materials and Methods
DNA Samples. This study concerns 2,196 unrelated male DNA samples (Table 1). We collected 904 new unrelated males subjects, from three different countries (Portugal, France, and Italy): 79 from North Portugal and 59 from South Portugal; 243 from the Marseilles region of France; 192 from Genoa, 64 from Rome, and 128 from Naples in continental Italy; 39 from Sicily; and 100 from Sardinia. All these new samples correspond to adult males, whose origin is based on the local birthplace of their fathers and (at least) grandfathers. We have obtained informed consent from each of the French subjects studied.

We add for comparison the following subjects, already tested as bearing haplotype V in previous studies: 11 subjects from Mauritania, 51 Berbers from Morocco, 59 Arabs from Rabat, 80 subjects from Algeria, 39 subjects from Tunisia, and 17 subjects from Libya (Lucotte et al. 2000); 29 Spaniards from Sevilla (Lucotte et al. 2001); 4 Spaniards from Barcelona and 9 French Catalans from Perpignan (Lucotte and Loirat 1999); 11 French Basques, 1 subject from Montpellier, and 7 subjects from Grasse in France and 6 subjects from Milan in Italy (Lucotte and Hazout 1996); and 44 subjects from Corsica (Lucotte et al. 2002).
Figure 1 indicates the representative geographic points in the western Mediterranean area for each of the 22 populations studied.

Genetic Analysis. Blood samples were collected by venipuncture using EDTA as an anticoagulant. Genomic DNA was extracted as described by Gautreau et al. (1983), using proteinase K and phenol-chloroform extractions.

Y-chromosome haplotypes were obtained using Southern blot analysis by hybridizing TaqI-restricted DNA successively with the p49f,a-specific probes, oligolabeled by random priming, according to the method described by Lucotte et al. (1994). Subdivision of haplotype V detected by Southern blot analysis into subhaplotypes Va and Vb was realized by polymerase chain reaction (PCR), using the assay published by Gonçalves and Lavinha (1994); the presence of the "low" XY275 allele (275G) defines subhaplotype Vb, and the other allele defines subhaplotype Va.

To compare subhaplotypes Va and Vb with the E3b1 and E3b2 subhaplo-groups [according to the Y Chromosome Consortium (2002) nomenclature], we further analyzed our Berber and Arabic DNA samples from Morocco for biallelic markers M78 and M81 using PCR (Underbill et al. 2000).

Realization of the Isofrequency Haplotype Maps. The maps of subhaplotype Vb and Va isofrequencies have been drawn with the Spatial Analyst program (Arcview software) using the Kringing procedure. We have used the inverse distance weighting method (with a power of 2), which is well adapted to scarce data in coastal areas and on islands in the western Mediterranean area. Five neighbors are calculated for each quadrant.

Results
Table 1 summarizes the frequencies we obtained for haplotype V and sub-haplotypes Vb and Va in the 22 study populations. For the 2,196 males typed, 491 (22.3%) bear haplotype V. The frequency of haplotype V is 35.5% in Portugal, with a more elevated proportion in the south (49.2%) than in the north (25.3%). The frequency of haplotype V in the Marseilles region (11.1%) has a value similar to the mean value in continental France (9%). In Italy the highest frequency is attained in Sicily (28.2%), followed by Naples at 17.2%. As previously shown (Lucotte et al. 2000), haplotype V is found at the highest frequency (68.9%) in Berbers from Marrakech in Morocco; an apparently increasing east-west cline in haplotype V frequencies is shown in North Africa from Libya (44.7%) to Rabat (57.7%), with intermediate values for Tunisia (53.4%) and Algeria (56.7%). In Spain haplotype V is much more frequent (40.9%) in the south of the country [in Andalusia (Sevilla)] than in the north (12.9%) [in Catalonia (Barcelona)].

Subhaplotype Vb is the Berber subhaplotype because its most elevated relative value (63.5%) is obtained for the Berber population of Marrakech. In the non-Berber population of Rabat in Morocco, the frequency of subhaplotype Vb is only 20.6%, whereas the frequency of subhaplotype Va (Arab) is 37.3%. In order of decreasing values, the subhaplotype Vb frequencies are 40% in Mauritania, 35.9% in South Portugal, 25.4% in Andalusia, and 15.8% in Libya. Low frequencies of subhaplotype Vb are found in Sicily (5.1%), Algeria (2.8%), Tunisia (2.7%), and North Portugal (2.5%); frequencies less than 2% are found in French Basques (1.9%), in Naples (0.8%), and in Corsica (0.6%), Subhaplotype Vb is absent in Catalonia (Barcelona and Perpignan), in the south of France (Montpellier, Grasse, and the region of Marseilles), in continental Italy (Milan, Genoa, and Rome), and in Sardinia.

Table 2 summarizes the frequencies of subhaplotype Vb in North Africa, Iberia, the south of France, and Italy. The maximum value (63.5%) concerns the Berber population, but this frequency is notably lower (9.3%) for other populations from North Africa. In southern Iberia an elevated value (30%) is observed, but the frequency of subhaplotype Vb is only 1.8% in northern Iberia. These frequencies are less than 1% in France and Italy.

Figure 2 shows the isofrequency map of subhaplotype Vb in the western Mediterranean area (coordinates on the map: x = longitude, y = latitude). From the Berber focus in Berbers from southern Morocco, the frequencies of subhaplotype Vb decrease in North Africa to the north of Morocco and to the east in Algeria and Tunisia. For Iberia the most elevated value of subhaplotype Vb frequencies is in southern Portugal; relatively elevated values are observed in Andalusia, moderate values are observed in the southern part of Spain, and low values are seen in Catalonia.

In the present study all haplotype V non-subhaplotype Vb subjects are termed subhaplotype Va (Arab) subjects. Their maximum relative frequencies are 53.9% (Algeria), 50% (Tunisia), and 37.3% (Rabat) in North Africa. Table 3 summarizes the frequencies of subhaplotype Va in North Africa, Iberia, southern France, and Italy. The maximum value (45.8%) is found in North Africa. In northern Iberia a slightly more elevated value is observed (20%) compared to southern Iberia (14.6%). A frequency of 10.3% is seen in France, and in Italy the 14.6% value observed in the south is relatively more elevated than in the north (3.4%)

Figure 3 gives the isofrequency map of subhaplotype Vb. In North Africa frequencies decrease from east to west and southward. For southern Europe the map shows the relatively higher percentages observed in the south of Italy versus the north and (to a lesser degree) in the north of Iberia versus the south.

In our PCR assay the 68 Moroccan subjects with subhaplotype Vb (47 Berbers and 21 Arabs) were tested for the M81 marker: All subjects were positive for the M81 marker, so subhaplotype Vb is homologous with subhaplogroup E3b2. The 38 Moroccan non-Berber subjects were further tested for the M78 marker: Only 31 of them (80.8%) were positive for the M78 marker; we conclude that, in Morocco at least, subhaplotype Va corresponds only partly to subhaplogroup E3b1

Discussion
P49a,f TaqI haplotype V, which is homologous with haplogroup E3b according to the Y Chromosome Consortium (2002) nomenclature, is the predominant Y-chromosome haplotype in North Africa (Lucotte et al. 2000), where its geographic distribution shows an east to west cline. In the present study we have extended the research of haplotype V frequencies (Lucotte et al. 2001) in various European populations located in the western Mediterranean basin to include France, Portugal, and Italy. The frequency of haplotype V in the Marseilles region is 11.1%, a value similar to the main value we obtained previously for continental France (Lucotte and Hazout 1996). In continental Italy we observed the highest haplotype V frequency in Naples (17.2%); Sicily, with a frequency of 28.2%, corresponds to the most elevated value we observed for Italy. In South Portugal the frequency of haplotype V is very high (49.2%); we had previously obtained a similar value for Libya and for Mauritania. The frequency of haplotype V for North Portugal (25.3%) is similar to the value we obtained for Sicily in the present study.

To better divide haplotype V into its ethnic components, we have subdivided it into subhaplotypes Vb (Berber) and Va (Arab). We have established that subhaplotype Vb is the Berber haplotype, because it is present at very elevated frequencies (63.5%) in our Berber population from Morocco but at relatively low frequencies (20.6%) in our non-Berber population of Rabat. Such a distinction of a Berber component was also realized by Scozzari et al. (2001), because they observed that the haplogroup they named 25.2 was also more frequent in the Berber population from Morocco than in Arabs. Our present results show that subhaplotype Vb frequencies in North Africa decrease from west to east, starting from the Berber focus in Morocco; in the western Mediterranean area subhaplotype Vb is at low frequencies along the south coast of Europe but occurs at relatively elevated frequencies in southern Iberia (peaking at 35.9% in South Portugal). Flores et al. (2004), in their important study of various locations in Iberia, observed that subhaplogroup E3b2 is more frequent in southern Iberia, attaining a maximum value of 11.5% in the region of Málaga.

In the present study all the non-subhaplotype Vb subjects bearing haplo-type V are classified as subhaplotype Va (Arab); they probably correspond to a heterogeneous group representing various ethnicities (our results concerning the incomplete correspondence between subhaplotypes Va and E3b1 in Morocco suggest that). We have shown here that in North Africa the focus of subhaplotype Va frequencies is in Algeria (53.9%) and Tunisia (50.6%); from this focus frequencies of subhaplotype Va decrease in the south and the west of the region.
Subhaplotype Va attains substantial frequencies along the southern coast of Europe; these frequencies reached relatively elevated frequencies in France (Perpignan, 11.8%) and in southern Italy (Naples, 16.4%; Sicily, 23.1%). For Iberia, relatively more elevated values are attained for Andalusia (15.5%) and for North Portugal (22.8%). Brion et al. (2004) also showed relatively higher frequencies of haplogroup E* (xE3a) (up to 18.3%) in their study concerning northern Iberia.

We had previously established (Lucotte et al. 2001) that haplotype V showed a gradient of decreasing frequencies with latitude in Iberia, and we interpreted this pattern as a consequence of the historical Islamic occupation of the peninsula (Conrad 1998). The results reported in the present study concerning subhaplotypes Vb and Va (subhaplotype isofrequencies maps given in Figures 2 and 3) have again shown both of these gradients. From this perspective, the opposite pattern of gradient frequencies observed in Iberia for the western European haplotype XV (Diéterlen and Lucotte 2005) is reconciled with the slow reconquest of the Iberian peninsula from the north by the Christians, which lasted seven centuries and ended in Granada in 1492

Examination of the above, with the assistance of references to older Lucotte et al. studies:

With regards to Lucotte et al.'s earlier reference to RFLP haplotype V as "Arab", Keita is right about the "Arabic" label being misleading, but in fact, if one reads Lucotte et al. later work, it is clear that they associate this haplotype with North Africans. Lucotte et al. refers to North Egyptians, as Egyptian "Arabs", and makes reference to groups in other parts of North Africa as "Arabs" as well. So, in actuality Lucotte et al. were associating haplotype V with what they perceived as "Arabized" north Africans. And so, as one can see, they refered to haplotype V as "Arab" and "Berberian", and made note of the fact that the Falasha had a high frequency "haplotype V and XI", which attests to their African provenance.

Keita associates haplotype V and XI with African origin, but so does Lucotte et al. Keita associates V and XI [barring his reference to other contexts used by other researchers] with M35/215, but if Lucotte et al. associate these with "North Africans" and Ethiopian Jews, and proclaim that is of African provenance, they too must be associating it with M35/215. M81 is the predominant "Berber" variant of M35. So the question is, if haplotype V is predominantly "Berber" and associated with "Berber origin", and haplotype XI is noted to have high frequencies in Eastern Africa, and decreases as one moves west of the African continent, then what is haplotype V and haplotype XI, as presented by Lucotte et al.?...In the meantime from Keita's publication:

Some TaqI 49 a,f variants have multiple associations; for example VIII is affiliated with several lineages (Al-Zahery, 2003). So far research indicates that haplotype V in Africa is associated with the M35/215 (or 215/M35) subclade, **as is XI**, and IV with the M2/PN1/M180 lineage, both of the YAP/M145/M213 cluster. These lineages that in Africa that affiliate with haplotypes V, XI, and IV (called “sub-Saharan”), are joined by a transition mutation: “(M)ost notably the PN2 transition…unites two high frequency subclades, defined by M2/PN1/M180 mutations in sub-Saharan Africa, and M35/215 in north and east Africa…” (Underhill et al., 2001, p.50; see also Cruciani et al., 2002).

Hence, Lucotte et al.'s 'Arab' and 'Berber' appellations to V haplotypes in north Africa, appear to be what they deem 'Arabic' speaking North Africans and 'Berber' speaking North Africans. On another note, it also appears that haplotype XI is also affiliated with E-M35 in the Lucotte et al. data Keita used. To be certain about any of this, one might as well examine primary texts from Lucotte et al. themselves:

Y-chromosome DNA haplotypes in North African populations

Abstract:
The frequency distribution of Y-chromosome haplotypes at DNA polymorphism p49/TaqI was studied in a sample of 505 North Africans from Mauritania, Morocco, Algeria, Tunisia, Libya, and Egypt. A particulary high frequency (55.0%) of Y-haplotype 5 (A2,CO,DO,F1,11 ) was observed in these populations, with a relative predominance in those of Berber origin. Examination of the relative frequencies of other haplotypes in these populations, mainly haplotype 4 (the "African" haplotype), haplotype 15 (the "European" haplotype), and haplotypes 7 and 8 (the "Near-East" haplotypes), permit useful comparisons with neighboring peoples living in sub-Saharan Africa, Europe, and the Near East.

The highest frequency of haplotype 5 (68.9%) was previously observed in Berbers from Morocco, and it has been established that this haplotype is a characteristic Berber haplotype in North Africa....

Haplotype 5 (A2, C0, D0, F1, I1) has a particularly high frequency (55%) in North Africa (Lucotte et al. 2000), and is of predominantly Berber origin. — Lucotte et al.; North African genes in Iberia studied by Y-chromosome DNA haplotype 5; 2001.

At least based on this, with caution, it is strongly suggestive of M81 derivatives. The authors note that it is supposed to be a characteristic Berber haplotype in North Africa, and is of predominantly Berber origin. However, we also know that the Lucotte et al. data cited by Keita also shows V haplotypes in Egypt, along with XI haplotypes. V haplotype in Egypt has a gradient that increases as one moves from south to north, while that of XI is the opposite, with a gradient increasing as one moves from north to south. So in the Egyptian context, does this mean that V is still suggestive of E-M81 chromosomes? Who knows; but M81 is certainly attested to in Egypt. What about XI? Could that be suggestive of an M78 derivative? Plausible, given its high frequencies in sub-Saharan Africa, particularly east Africa. One has to ascertain this plausibility. **To be certain, one would have to be familiar with the specific binary markers that Lucotte et al. would have searched [usually done, once a restrictive digest [by restrictive enzyme] is undertaken to cut DNA into fragments] and amplified [PCR] for haplotype V in the 2001 study above and their 2003 study cited by Keita, to see if they continued to be in the same exact contexts or if variant binary markers were used in respective studies. As noted in the linked discussion, in Lucotte et al.'s case, this doesn't appear to be the case. However, the authors of the current head topic have addressed this issue, reassuring us with relatively more precision, what specific V haplogroups were involved in Lucotte et al.'s several studies.

To be continued.

Link to part 2:
RFLPs: Lucotte et al., A case study — Pt. 2 [clickable]

Skin pigmentation gene alleles — Part 2

Reviewing H. Norton, R. Kittles et al, 2006 - Part 2:

Link to the part 1: Skin pigmentation gene alleles [clickable]

Additional notes:

For those who are curious, the authors of the aforementioned skin pigmentation study [Kittles et al.] don't specifically point out the TMRCAs for the identified genes in question, but apparently ancestral lineages were delineated from their derived counterparts. From extrapolation though, it makes sense that mutations that occurred after divergence of any given groups, would be relatively rare in the common ancestor of these recently diverged groups. On the other hand, certain mutations that were present within the common ancestor may be expressed more acutely later on in one or the other group that diverged from this ancestral population, while dying out or becoming relatively rare in another progeny group. Still these developments are able to assist one in delineating the frequency and mutational particulars of the genes controlled by natural selection and/or the pressure of genetic drift.

As for "Southwest Asian" populations, they generally fall into ranges contained within the Saharo-tropical Africans, while some northerly groups of this region apparently have relatively paler skin shades as a product of more recent migrations into the region. Kittles et al. at least in part, attribute such developments to gene flow from Northern Eurasia and perhaps, in some areas, East Asia. See again, from my last post:

Concerning the "derived" SLC24 A5 gene...

In contrast, the SLC24 A5 11*A-derived allele is found at low frequencies in several sub-Saharan populations including the West African Mandinka and Yoruba, the Southern African San, and South West Bantu.

The relatively **high frequencies** of the derived allele in **Central Asian, Middle Eastern, and North Africa** seem likely to be **due to recent gene flow** with European populations.

Similarly, the presence of the derived allele (albeit at low frequencies) in some sub-Saharan populations may be due to recent gene flow from European and Central Asian populations. —Alternatively, the derived allele may have lost in the ancestors of modern East Asians but retained in the ancestral European populations. The allele then rose to high frequency in Europeans following the divergence of Europeans and East Asian ancestral groups.

Many places outside of Africa, for instance, harbor the 'derivative' counterparts of several "pigmentation" genes [a variety of which have been associated with relatively lighter pigmentation], while ancestral alleles [many of which have generally been associated with relatively darker pigmentation] are commonly found in Africa and amongst direct descendants of earliest out-of-Africa ancestors of modern non-Africans, as is the case with OCA2 gene...

In general, the derived allele (associated with lighter pigmentation) is most common in Europeans and East Asians, and the **ancestral allele** predominates in **sub-Saharan Africa** and **Island Melanesia.**

...and this quite likely applies to "southwest Asians"...in harboring "derived" OCA2 which has been associated with playing a role in lightening skin phenotype, for example.

Lightening effect was apparently a gradual process, as populations started expanding to low UV radiation latitudes. This is readily seen in the intermediary situations between adaptations on opposite poles of empirical tests; see for example:

High Fst values [concerning the three genes TYR, MATP and SLC24A5] between Europeans and darkly pigmented populations such as West Africans and Island Melanesians are not unexpected if these genes have functional effects. However, the notably elevated pairwise Fst values relative to East Asians (the population in our panel that is the most similar to Europeans in pigmentation phenotype) is striking. Populations intermediate in pigmentation (Native Americans and South Asians) also exhibit Fst values falling in the top 5th percentile of their relevant Fst distributions with Europeans for these three loci. In the case of SLC24A5 A111G, South Asian pairwise Fst values also fall in this top 5th percentile when compared to both Europeans (Fst = .389,  p < .01) and East Asians (Fst= .519, p < .01), but not when compared to any other population. At all three loci Europeans have the highest frequency of the derived alleles relative to the other five populations.

The South Asians being referred to here, comprised of east Indian samples, which are claimed to be 'intermediate' along with the Native American samples. The phenomenon described above, seems to suggest that the alleles at the three said loci in the said 'intermediate' groups predate those attained in both East Asians and Europeans; as noted, their Fst values are not quite as high when compared with any other population [which would essentially be the dark populations]. Apparently, the pigmentation distribution in these 'intermediate' groups reflect demographic events distinct from those that produced the more dramatic pigment-oriented phenotypic manifestations in Europeans and East Asians respectively; being that they possess alleles that post-date OOA migration events, and yet those that predate extreme pigment-related adaptations sported by Europeans and East Asians, they are bound to report intermediary patterns. One might recall that the Native American OCA2-derived allele frequency was said to be comparatively lower than that of East Asians...

Interestingly, derived allele frequencies at this locus are quite different between Native American (15%) and East Asian populations (45%), suggesting that perhaps the derived allele at this locus did not reach very high frequencies in East Asians until after the colonization of the Americas

And might also recall that KhoiSans on the other hand, reported high frequencies of OCA2-derived...

The lightly pigmented hunter-gatherer San populations of Southern Africa is exceptional in having a high frequency of the derived allele relative to geographically proximate and more darkly pigmented African populations (Jablonski and Chaplin 2000), further supporting the importance of OCA2 in regulating normal variation in pigmentation. The widespread distribution of the derived allele in the CEPH-Diversity Panel suggests that it is not necessarily a new mutation, nor has it been restricted to a specific geographic area.

So yes, the derived version of OCA2 likely predates the often-talked about Upper Paleolithic OOA migration in varying frequencies in different populations, but likely did not pick up in distribution dramatically in East Asian and European populations, until after a section of central-East Asian had left for America, in a wave(s) following that of the first Paleo-Americans. This suggests that one drift episode [perhaps amongst the earliest for this type] raised its frequency considerably in at least one African group [the Sans], another drift episode raised its distribution in vicinity of central and/or east Asia to reasonably visible levels, but yet another drift episode raised its distribution even further in east Asia at a later time. All this paints gradual evolution in skin pigmentation relaxation temporally, in tandem with territorial shifts amongst populations.

And recalling...

The discordance between our Fst -based divergence values and allele frequencies in the Melanesian CEPH populations at ASIP largely stem from the relatively low frequency of the ancestral allele in the 2 CEPH Island Melanesian populations relative to our original Island Melanesian sample. These discrepancies make it difficult to determine if ASIP truly underlies broad pigmentation differences between darkly and lightly pigmented populations or instead inter-population variation at this locus can largely be explained by differences between Africans and non-Africans…

The answer is rather obvious, no? It reflects the basal phylogenetic position of Melanesians, which is why they'd share ancestral ASIP alleles with continental Africans. The difference then here, would be one of the basal phylogenetic position of Africans vis-a-vis OOA-derived populations, with the deepest-clade bearers of all OOA-derived groups carrying over basal African alleles outside of Africa.

More on the "derived" SLC24 A5 gene...

On the above piece, in one personal encounter, a question had come up along the lines of:

...because one of the authors says not enough time has passed for mutations? And just how is it the author would know this? Since the author, or one of the authors didn't specifically say how much time has to pass for mutations then I'm asking you...to explain to me what you know they meant by this.

The natural answer to that question, as the present author put forth, was this:

Yes, the authors reckon that "not enough time has passed for mutations" and don't specify "how much time has to pass for [new] mutations" to occur, nor need to, because they determined this from the fact that the DNA flanking the gene in question lacked variation in the samples they studied; the tacit idea here, is that the DNA locus in question not only indicates selective pressure of the gene SLC24A5, where by the flanking DNA in question must have been part of a selective sweep, but its lack of variation suggests that not enough time has accumulated since such a selective sweep would have occurred; otherwise, more variation, however modest, would be expected of a designated DNA locus that has been around for a considerable length of time. And oh, it must be suggestive of some linkage disequilibrium in the inheritance of this assemblage of DNA.

On Jablonski :

The weaker the ultraviolet light, the fairer the skin. Jablonski went on to show that people living above 50 degrees latitude have the highest risk of vitamin D deficiency. "This was one of the last barriers in the history of human settlement," Jablonski says. " Only after humans learned fishing, and therefore had access to food rich in vitamin D, could they settle these regions." — The evolution of race was as simple as the politics of race is complex, By Gina Kirchweger

And to that, the present author says:

"Absolutely"!

Side notes:
The very "relaxed" eumelanin concentration in the skin of 'pale skin' individuals is the expression of their relatively "recessive" alleles, vis-a-vis the more "dominant" counterparts of those that instruct for more production, to produce the considerable skin eumelanin concentration of dark skin individuals. The present author has come across comical claims about the aforementioned "recessive" counterparts "masking" the effects of the more "dominant" skin pigmentation alleles, no doubt from individuals who are in the dark about the basics of genetics. The case in humans, whereby one comes across skin tone gradients, from extreme dark to extreme paleness, can best be described as one of the interplay of "incomplete" dominance of the 'wild types' over their "recessive" counterparts in subjects of "intermediate" skin tones, via polygenic inheritance—wherein the individual effects of "dominant" or "wild" alleles that produce greater eumelanin dosage, will mask those of the relatively "recessive" counterparts in normal "heterozygous" [so to speak, for simplification purposes] subjects, while the "wild" or "dominant" allele types will simply instruct for considerable skin eumelanin in "homozygous" dark skin subjects, and that of the relatively "recessive" allele types instruct for little dosages in "homozygous" pale skin subjects. Now of course, common sense should tell one that these terms "recessive" vs. "dominant" are relative terms, for we know that even in individuals where 'pale skin' is product of natural selection, as opposed to genetic mishap or disorder, the alleles which instruct for only modest eumelanin dosage, if we had two copies of the same alleles from each parent, then neither is dominant or recessive to the other; however, one such allele in the presence of a "wild type" from a darker skin parent, will tend to be "recessive" relative to the said "wild type". All the said alleles in this case, will instruct for eumelanin dosage, but the 'wild type", and hence more "dominant" type, will instruct for bigger dosages than the other allele, the "recessive" counterpart. For those needing basic illustrative demostration, check this site out: http://waynesword.palomar.edu/lmexer5.htm

Gist: If one is recessive, it has to be recessive relative to another, and likewise, if one is "dominant", it has to be so over another. It is just common sense.

Skin pigmentation gene alleles

Reviewing H. Norton, R. Kittles et al, 2006:

Besides the variations in the “SLC24A5” gene, as mentioned in the intro article, the “TYR” gene, the “OCA2“, the “ASIP“, and to some extent those seen in the MC1R gene, Kittles et al. have noted other genes "MATP C374G", “ADAM17“, “ATRN“, and “DCT” the mutations of which are deemed to have to had influence in promoting paleness…

Taken together (with the results of previous admixture mapping studies), these results point to the importance of several genes in shaping the pigmentation phenotype and a complex evolutionary history involving strong selection. Polymorphisms in 2 genes, ASIP and OCA2, may play a shared role in shaping light and dark pigmentation across the globe, whereas SLC24A5, MATP, and TYR have a predominant role in the evolution of light skin in Europeans but not in East Asians. These findings support a case for the recent convergent evolution of a lighter pigmentation in Europeans and East Asians…

Pairwise Fst estimates for the ASIP A8818G and OCA2 A355G SNPs tentatively suggest a pattern of divergence between 4 populations (Europeans, East Asians, Native Americans, and South Asians) and the relatively more darkly pigmented populations of West Africa and Island Melanesia, or possibly only between West Africans and all other populations. At both loci, West Africans and Island Melanesians have higher frequencies of the ancestral alleles than the other 4 populations. Pairwise locus-specific Fst values falling in the top 5% of the empirical distributions are observed between West Africans and 3 other populations (South Asians, Native Americans, and Europeans) at ASIP A8818G. Fst values between West Africans and East Asians at this locus are elevated but do not reach our cutoff value of 5% (Fst = .489, P = .065). At OCA2 A355G, only West Africans and Europeans show Fst values falling into the top fifth percentile of relevant comparisons (Fst = .516, P<.05). The low pair wise Fst values and higher frequency of ancestral alleles at both SNPs studied in these loci between West Africans and Island Melanesians hint that dark pigmentation associated with both loci in these populations may have a common evolutionary origin (Mean Fst (WA-IM) = .182; ASIP A8818G Fst (WA-IM) = .260, P = .282; OCA2 A355G Fst (WA-IM) = .101, P=.525).

Continuing with regards to OCA2 gene, we are told…

In contrast, the ancestral allele associated with dark pigmentation has a shared high frequency in sub-Sharan African and Island Melanesians. A notable exception is the relatively lightly pigmented San population of Southern Africa where the derived allele predominates (93%), although this may be simply due to small sample size (n=14).

The distributions of the derived and ancestral alleles at TYR A192C, MAPT C374G, and SLC24A5 A111G are consistent with Fst results suggesting strong European specific divergence at these loci. The derived allele at TYR, 192*A (previously linked with lighter pigmentation [Shriver et al. 2003]), has a frequency of 38% among European populations but a frequency only 14% among non-Europeans. The differences between Europeans and non-Europeans for the MAPT 374*G and SLC24A5 111*A alleles (both derived alleles associated with lighter pigmentation) were even more striking (MAT [European] = 87%; MATP [non-European] = 17%; SLC24A5 [European] = 100%; SLC24A5 [non-European] = 46%). The frequency of the SLC24A5 111*A allele outside of Europe is largely accounted for by high frequencies in geographically proximate populations in northern Africa, the Middle East, and Pakistan (ranging from 62% to 100%).

By way of negative Tajima D values, which when strongly negative, indicate selective pressure, or more specifically—“directional selection”, especially when taken into account with both high locus-specific branch length and strongly negative heterozygosity values, the authors continue...

These data confirm the unusual European-specific patterns at MATP and SLC24A5. Both genes display long range (consecutive windows) and significant indications of positive selection for all 3 statistics. In contrast, there is little evidence of a European-specific pattern in the TYR locus although the non-synonymous TYR A192C SNP does individually show a strongly significant CEU-LSBL (P<.003) in the HapMap data as in our original findings. The contrast may be explained by the limitations of our HapMap sliding windows analyses, whereby adjacent SNPs are averaged using a method that does not consider Haplotype structure.

East Asians showed relatively stronger selection for a different set of genes…

…In particular, 2 genes (ADAM17 and ATRN) showed East Asian-specific signatures comparable in strength with those observed for MATP and SLC24A5 in Europeans.

While…

The ADTB3A gene also shows a strong and focused signature of positive selection in Africans...

Many hypotheses predict that natural selection will eliminate genetic variants associated with lighter skin in the regions of high UVR as a protection against photo damage (e.g., sunburn, melanoma, and basal and squamous cell carcinomas) (Blum 1961; Kollias et al. 1991) and folic acid photo degradation (Branda and Eaton 1978; Jablonski and Chaplin 2000). The photo protective properties of a highly melanized skin and the recent African origin of modern humans suggest that the ancestral phenotype is one of the relatively dark skin (Jablonski and Chaplin 2000; Rogers et al. 2004). If dark skin is the ancestral phenotype, then we may assume that the first migrants out of Africa were relatively darkly pigmented…

There are 2 primary explanations for the evolution of lighter skin in regions of low UVR:

—1)The first suggests that light skin is merely due to the relaxation of functional constraint and that derived alleles associated with lighter pigmentation may have simply drifted to high frequency in the absence of strong purifying selection (Brace 1963).

—2)The second explanation suggests that in lower UVR regions, positive selection would have favored mutations leading to lighter skin as a way to maximize cutaneous vitamin D synthesis (Rana et al. 1999; Jablonski and Chaplin 200). Given the relatively recent arrival and divergence of humans in and across Europe and Asia, the most parsimonious evolution of light skin would involve such mutations arising in a proto-Eurasian population soon after humans left Africa.

Consequently, these mutations should be shared between modern Asian and European populations. Alternatively, if separate existing functional variants were driven to high frequency in East Asian and Europeans or independent de novo mutations arose and were selected in each population after divergence of Europeans and Asians, then these would be obvious as high allele frequency differences between modern European and East Asian populations. Reduced levels of heterozygosity surrounding the SLC24A5 A111G polymorphism in the European, but not East Asian, HapMap populations support the latter hypothesis (Lamason et al. 2005), as do reduced polymorphism levels based on full resequencing data from MATP in populations of European descent (Soejima et al. 2005).

So basically, while “SLC24A5, MATP, and TYR have a predominant role in the evolution of light skin in Europeans,” the ADAM17, ATRN, and DCT appear to play a dominant role in the evolution of light skin in East Asians.

Current archeological evidence suggests human presence in Island Melanesia by at least 40ky ago and in other parts of Sahul by at least 45ky ago (O’Connell and Allen 2004). If the original migrants to Oceania arrived there via a corridor of relatively high UVR, then we might expect their descendants to share ancestral pigmentation variants with African populations. However, if the ancestors of modern day Island Melanesians spent a significant amount of time in low-UVR, then it is possible that mutations associated with lighter pigmentation could have accumulated and a readaptation to high-UVR conditions would have been necessary, leading to potential divergence between Island Melanesians and Africans at functional pigmentation loci. In actuality, both of these scenarios may apply, as we know that modern Island Melanesian populations are descended broth early migrants (arriving 40ky ago) as well as later proto-Austronesian-speaking peoples from a southeast Asian homeland ~ 3,200 years ago (Spriggs 1997).

The discordance between our Fst -based divergence values and allele frequencies in the Melanesian CEPH populations at ASIP largely stem from the relatively low frequency of the ancestral allele in the 2 CEPH Island Melanesian populations relative to our original Island Melanesian sample. These discrepancies make it difficult to determine if ASIP truly underlies broad pigmentation differences between darkly and lightly pigmented populations or instead inter-population variation at this locus can largely be explained by differences between Africans and non-Africans. The discordance between the frequencies of the ASIP ancestral allele in our original Island Melanesian sample and the Melanesian samples from the CEPH panel may be indicative of both the complex demographic history of Island Melanesia (involving several migratory events (Spriggs 1997) and probable extensive genetic drift (Friendlaender 1975, 1987) as well as the importance of multiple loci in determining pigmentation phenotype…

Thus possible further extensions of variations detected amongst Melanesians can be explained by successive demographic events After their African ancestors migrated over 40ky ago. The “original Melanesian sample” appears to have more ancestral pigmentation genes in common with tropical Africans, which is to be expected given that they are direct descendants of the earliest Eurasians, as demonstrated as follows with the OCA2 gene…

In general, the derived allele (associated with lighter pigmentation) is most common in Europeans and East Asians, and the ancestral allele predominates in sub-Saharan Africa and Island Melanesia.

The mutations in the OCA2 gene may well have implications on imparting paleness, as demonstrated in the south African San people…

The lightly pigmented hunter-gatherer San populations of Southern Africa is exceptional in having a high frequency of the derived allele relative to geographically proximate and more darkly pigmented African populations (Jablonski and Chaplin 2000), further supporting the importance of OCA2 in regulating normal variation in pigmentation. The widespread distribution of the derived allele in the CEPH-Diversity Panel suggests that it is not necessarily a new mutation, nor has it been restricted to a specific geographic area.

While it seems plausible that the “derived” OCA2 gene came to being before the out-of-Africa migration that give rise to modern Eurasians, it doesn’t appear that this derived allele was necessarily widespread, and may well have been later on selected for in European and East Asians…

Interestingly, derived allele frequencies at this locus are quite different between Native American (15%) and East Asian populations (45%), suggesting that perhaps the derived allele at this locus did not reach very high frequencies in East Asians until after the colonization of the Americas

Contrast the situation with OCA2 gene with that of the MATP 374*G allele…

The virtual absence of MATP 374*G-derived allele in the sub-Saharan African populations that we examined in the CEPH-Diversity Panel is consistent with the origin of this mutation outside of Africa AFTER the divergence of modern Asians and Europeans.

Contrasting that of the “derived” SLC24 A5 [as in the case with the “derived” OCA2 allele], where two possible scenarios arise…

In contrast, the SLC24 A5 11*A-derived allele is found at low frequencies in several sub-Saharan populations including the West African Mandinka and Yoruba, the Southern African San, and South West Bantu. *The presence of the derived allele (albeit at low frequencies) in some sub-Saharan populations may be due to recent gene flow from European and Central Asian populations...

—1)The relatively high frequencies of the derived allele in Central Asian, Middle Eastern, and North Africa seem likely to be due to recent gene flow with European populations.

—2)Alternatively, the derived allele may have lost in the ancestors of modern East Asians but retained in the ancestral European populations. The allele then rose to high frequency in Europeans following the divergence of Europeans and East Asian ancestral groups.

The different mechanism of the evolution of light skin in Europeans and East Asians apparent from genetic examination, supports the understanding that evolution of pale skin came very late, because if had occurred prior to the divergence of the Europeans and East Asians, then it seems highly plausible that they would share more in common with one another the dominating alleles in playing a role in skin lightening…but as demonstrated, different set of alleles play dominating role in the lightening effect of the skin in Europeans and East Asians…

These results simultaneously and strongly suggest that Europeans and East Asians have evolved lighter skin independently and via distinct genetic mechanism, as there is an absence of any unusual pattern of diversity at SLC24A5, MATP, and TYR in East Asians.

The interesting part of the study, is this about the MC1R gene about its…

The MCIR gene was the only locus examined in detail that did not show any signal of potential positive selection. Previous sequence-based studies have reached conflicting conclusions about whether or not MC1R has been subject to positive selection outside of Africa (Rana et al. 1999; Harding et al. 2000; Makova et al. 2001).

Although MC1R’s association with red hair, fair skin, freckles, and melanorma risk in European and European-derived populations primarily from the British Isles (Box et al. 1997; Smith et al. 1998a; Schioth et al. 1999; Flanagan et al. 2000; Bastiaens et al. 2001) clearly demonstrates the important regional role that it plays in pigmentation, MC1R may have (with some exceptions [John et al. 2003; Nakayama et al. 2006]) little effect on variation outside of Europe (Myles et al. 2006). Consequently, no signal will be detected using our approaches.

Although the 2 SNPs that we typed in MC1R are not strongly associated with the red hair and fair skinned phenotype for which MC1R is so well known (Sturm et al. 2003), both are polymorphic in global surveys of populations (Rana et al. 1999; Harding et al. 2000). In addition, the MC1r G92A SNP may have a ”mild” effect on pigmentation phenotype (Motokawa et al. 2006). The 92*A allele at this site is known to have a lower affinity for alpha-MSH than wild-type MC1R alleles (Xu et al. 1996), which suggests that it may contribute to **normal** variation in pigmentation. However, if positive directional selection has acted on MC1R, we would expect variation at linked sites to be affected. As such, even if have not assayed the relevant SNP, we should still have observed some signal selection, especially given the small size (~3 kb) of this gene.

So polymorphisms in the MC1R gene seem to have had relatively more impact in Europeans than other populations. Perhaps this might have something to do with the effects of MCIR mutations in Europeans having an "exacerbating effect", i.e. in addition to those of other “pigmentation”-influencing alleles therein…or maybe to some degree, tenuously linked to the effects of one or the other, or a few of those lightening alleles in Europeans.

Finally, the seem to be a strong case for the ASIP and OCA2 genes in playing a role as a tale-teller [by way of ‘ancestral‘ genes and their ‘derived’ counterparts ] of the derivation of non-Africans from Africans, the populations wherein polymorphisms at these loci could well have played a role in skin tone variation to some degree or another…

The pattern of diversity at ASIP 8818*G allele (the ancestral allele associated with darker pigmentation) indicates a role primarily in African/non divergence (sub-Saharan African frequency; 66%, all other populations; 14%) rather than between darkly and lightly pigmented populations. At OCA2 355, the derived allele (linked with lighter pigmentation) occurs at its highest frequencies across Europe and Asia, but is also relatively common among Native American populations (18-34%) and is present at much lower frequencies (0-10%) among Bantu-speaking African groups. In contrast, the ancestral allele associated with **dark** pigmentation has a shared **high frequency** in sub-Saharan African and Island Melanesians...

Observed patterns of global skin pigmentation diversity and their correlation with environmental UV exposure suggest an adaptive response. Although we cannot rule out a role for sexual selection, our results support multiple genetic mechanisms for evolution of skin color. We provide evidence that at least 2 genes, ASIP and OCA2, probably played a shared role in shaping light and dark pigmentation across the globe.

Aside from non-sequitur about the need for “uniformity” in dark hue in ancestral humans, considering that not even a single immediate family or household will necessarily pass for such a ridiculous test, all in all, Kittles et al.’s analysis lend strong support to the claims made by the likes of Jablonski, about dark skin being the original or default state of Homo Sapien Sapiens!

As a matter of fact, this paper discredits Frank Sweet's claim on his "Onedroprule" site, about the "default" human skin tone being light brown of the likes of Khoisan, and the "supposed dark tone of Bantus being more recent", as others and the present author himself have demonstrated in "Egyptsearch.com" discussions. There is no evidence that Africans in their ancestral skin tone state were uniformly dark skin, but preponderance of evidence does show that dark skin was the ancestral state of human skin pigmentation. As noted already, the ancestral alleles appear to be shared between dark skin populations like Melanesisans and tropical Africans.

This posting above, is itself a slightly modified repro of earlier posting in the following link: White race very young [clickable Egyptsearch link]

Referenced source: Genetic Evidence for the Convergent Evolution of Light Skin in Europeans and East Asians, by Rick Kittles et al. , 2006.

Link to part 2: Skin pigmentation gene alleles — Part 2 [clickable]