| According to "Out of Africa" hypothesis, modern human expanded to all of the world during the past 100,000 years. Human evolution, dynamics, migration history and pressure of nature selection have driven Human into different specific sub-groups. One of them is High-altitude populations, such as Tibetan population. The world's highest and largest plateau stands in Qinghai-Tibet Plateau. The local Tibetan population and its branches have been living in the Tibetan Plateau at least 20,000-30,000 years. The Tibetan population is the best choice to study the adaptation of high altitude, because of their unique language and culture, homogeneous genetic background, as well as the natural barrier of the Tibetan Plateau.Hypoxia is a main characteristic of High altitude. For living in this environment and undergoing the natural selection pressure, Tibetan population has taken physical changes, such as oxygen content, pulmonary ventilation and the oxidation-sensitive. The molecular genetic mechanisms of adaptation to hypoxia have been studied in Tibetan population, and some polymorphic alleles of hypoxic-associated genes were found, such as ACE, EPO, NOS3. However, previous studies comparing low altitude people with high altitude people found the difference in the non-coding loci. These studies are difficult to rule out differences in genetic background.Hypoxia inducible factor (HIF) is a very important oxygen regulator in mammals, which actives hypoxia-related genes in hypoxia. HIF constitutes withαsubunit andβsubunit.αsubunit is regulatory and half-life of it in normoxic condition is very short. Only in hypoxic conditions,αandβare the dimerization, which combine with hypoxic response element in down-stream genes, mediate hypoxia-related gene's transcription, and result in adaptive physiological changes to maintain the steady-state oxygen balance. Up to now, three isoforms of HIF-αhave been found. HIF-1αand HIF-2αis the most important regulation factors in hypoxic responses.In current study, we selected three Tibetan groups according to the different altitude, who are from Tibet province Langkazi county(4454m on average), Qinghai province Guinan county(3100m on average), and Yunnan province Gongshan county(2500m on average). According to the database of NCBI and HAPMAP, we genotyped 10 SNPs (rs3783752,C111A,rs61755705,rs4899056,rsl957757,rs10873142,rs41508050,rs11549465,rs11549467,rs1802821)and a GT dinucleotide repeats in HIF-1αgene and 14 SNPs(rs11125068,rs1867785,rs7571879,rs10193827, rs13419896,rs2881324,rs1562453,rs7589621,rs4953360,rs4953361,rs7594912,rs11675441,rs7571218,rs7598371) in the HIF-2a gene using restriction fragment length polymorphism (RFLP) and gene scan methods. The genotype and allele of populations was compared by software SPSS 17.0χ2 test module, SNP loci linkage disequilibrium analysis (D') and haplotype frequencies were estimated with software SHEsis (http://analysis.bio-x.cn/myAnalysis.php), P<0.05 was considered significant difference.Nature selection is mainly directed to the functional site, but the intron which are cis-elements in gene also have regulation function. These SNPs in intron were linkage disequilibrium with cis-regulatory regions.Our results showed there were four non-synonymous mutation SNPs site of HIF-1αgene in these populations. There were no significant difference of other SNPs among three Tibetan groups, except for SNP rs11549465 between Tibet Tibetan and Yunnan Tibetan, as well as between Qinghai Tibetan and Yunnan Tibetan. Frequencies of genotype and allele in rs4899056, rsl957757, rs10873142, rs3783752 had significant difference between Tibet Tibetan and Yunnan Tibetan, and between Qinghai Tibetan and Yunnan Tibetan. We observed these differences among three Tibetan groups were negative correlation with the altitude. In our study, we found a new C/T SNP adjacent with rs11549467 by sequencing, which affected result of RFLP of rs11549467, so there is no statistical data about rs11549467. This new SNP in the coding region is a synonymous mutation and not reported before. We constructed haplotype of rs11549465 and 4 introns SNPs and found haplotype B (C-C-C-T) prevailed in Yunnan Tibetan, haplotype G (T-T-T-C) in Tibet Tibetan and Qinghai Tibetan. Results of GT dinucleotide repeats showed that 14 repeats allele prevailed in Tibet Tibetan and Qinghai Tibetan,15 repeats allele prevailed in Yunnan Tibetan. The allele of 14 repeats was significant difference between Tibet Tibetan and Yunnan Tibetan, and Qinghai Tibetan and Yunnan Tibetan, but no difference between Tibet Tibetan and Qinghai Tibetan. So did 15 repeats allele.In the HIF-2 genes, except rs2881324, rs7594912, the other 12 alleles frequencies and genotype frequencies between the three groups were almost significantly difference. The difference is correlated with elevation. We constructed haplotype of 14 SNPs and found some haplotypes were significantly difference among three groups or between any two groups, especially between Yunnan Tibetan and Qinghai Tibetan, Yunnan Tibetan and Tibet Tibetan. Haplotype 1 (A-A),5 (C-G),9 (C-A-G-G), 16 (C-A-G),17 (C-G-G) were dominate in Yunnan Tibetan; haplotype2 (G-A),4 (C-A) in Tibet Tibetan; haplotype 3 (G-G),6 (T-A),12 (T-G-A-A),20 (T-G-C) in Tibet Tibetan and Qinghai Tibetan; haplotype 8 (C-A-G-A) in Yunnan Tibetan and Qinghai Tibetan.HAPMAP data suggested the differences of genotype frequency and allele frequency related with ethnic groups backgroups, such as Africans, Europeans and Asians. Why are significant differences among three Tibetan groups? According to different altitude of the three Tibetan groups, and HIF-la, HIF-2a gene features, we considered hypoxia caused by the high altitude environment is likely play a selection to HIF-la and HIF-2a gene, and the latter was more effective.Our data show that in three Tibetan groups, Yunnan Tibetan is very unique. Gongshan is located in the middle mountain range, Tibetan in there are relative isolation. We think genetic drift might influence the genetic background of the group. |
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