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A genetic mechanism for Tibetan high-altitude adaptation

Nature Genetics volume 46, pages 951–956 (2014)Cite this article

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Abstract

Tibetans do not exhibit increased hemoglobin concentration at high altitude. We describe a high-frequency missense mutation in the EGLN1 gene, which encodes prolyl hydroxylase 2 (PHD2), that contributes to this adaptive response. We show that a variant in EGLN1, c.[12C>G; 380G>C], contributes functionally to the Tibetan high-altitude phenotype. PHD2 triggers the degradation of hypoxia-inducible factors (HIFs), which mediate many physiological responses to hypoxia, including erythropoiesis. The PHD2 p.[Asp4Glu; Cys127Ser] variant exhibits a lower Km value for oxygen, suggesting that it promotes increased HIF degradation under hypoxic conditions. Whereas hypoxia stimulates the proliferation of wild-type erythroid progenitors, the proliferation of progenitors with the c.[12C>G; 380G>C] mutation in EGLN1 is significantly impaired under hypoxic culture conditions. We show that the c.[12C>G; 380G>C] mutation originated ∼8,000 years ago on the same haplotype previously associated with adaptation to high altitude. The c.[12C>G; 380G>C] mutation abrogates hypoxia-induced and HIF-mediated augmentation of erythropoiesis, which provides a molecular mechanism for the observed protection of Tibetans from polycythemia at high altitude.

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Figure 1: Genome-wide allele frequency differentiation between Tibetans, Mongolians and Europeans.
Figure 2: The p.[Asp4Glu; Cys127Ser] PHD2 mutant shows gain of function under hypoxia.
Figure 3: Erythroid colony (BFU-E) assays.
Figure 4: Hypoxia increases the proliferation of control BFU-E colonies but decreases that of colonies with the c.[12C>G; 380G>C] mutation in EGLN1.

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Acknowledgements

P.K. is supported by Academy of Finland grants 120156, 140765 and 218129, the S. Juselius Foundation, the Finnish Cultural Foundation and the Finnish Cancer Organizations. B.O. is supported by US National Institutes of Health (NIH) Mentored Career Development Award HL119355-01. J.X. is supported by the National Human Genome Research Institute (HG005846). G.R.-L. is supported by the National Basic Research Program of China (grant 2012CB518200) and by the Program of International Science and Technology Cooperation of China (grant 2011DFA32720). S.S. is supported by US NIH grant P01CA108671. T.S.S. is supported by the US NIH T32 Postdoctoral Fellowship HL098062. L.B.J. is supported by the University of Utah Seed Grant Program for studies of hypoxic adaptation. G.L.S. is supported by the Johns Hopkins Institute for Cell Engineering. J.T.P. is supported by US NIH grant P01CA108671, a Veterans Affairs Merit Review Award and the University of Utah Seed Grant Program for studies of hypoxic adaptation.

Author information

Author notes

  1. Felipe R Lorenzo, Chad Huff and Mikko Myllymäki: These authors contributed equally to this work.

  2. Peppi Koivunen and Josef T Prchal: These authors jointly directed this work.

Authors and Affiliations

  1. Department of Medicine, University of Utah School of Medicine and George E. Wahlin Veterans Administration Medical Center, Salt Lake City, Utah, USA

    Felipe R Lorenzo, Sabina Swierczek, Tsewang Tashi, Donald A McClain & Josef T Prchal

  2. Eccles Institute of Human Genetics, University of Utah School of Medicine, Salt Lake City, Utah, USA

    Chad Huff, Jinchuan Xing, Tatum S Simonson, Lynn B Jorde & Josef T Prchal

  3. Department of Epidemiology, University of Texas M.D. Anderson Cancer Center, Houston, Texas, USA

    Chad Huff

  4. Biocenter Oulu, Faculty of Biochemistry and Molecular Medicine, Oulu Center for Cell-Matrix Research, University of Oulu, Oulu, Finland

    Mikko Myllymäki & Peppi Koivunen

  5. Division of Cardiovascular Medicine, Department of Medicine, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts, USA

    Benjamin Olenchock

  6. Sickle Cell Center, University of Illinois, Chicago, Illinois, USA

    Victor Gordeuk

  7. Research Center for High-Altitude Medicine, Qinghai University, Xining, People's Republic of China

    Tana Wuren & Ge Ri-Li

  8. Icahn School of Medicine at Mount Sinai, New York, New York, USA

    Tahsin M Khan

  9. Sher-i-Kashmir Institute of Medical Sciences, Srinagar, India

    Parvaiz A Koul

  10. Regional Centre for Biotechnology, Gurgaon, India

    Prasenjit Guchhait

  11. Department of Pathology, University of Utah, Salt Lake City, Utah, USA

    Mohamed E Salama

  12. ARUP Laboratories, Hematopathology, Salt Lake City, Utah, USA

    Mohamed E Salama

  13. Department of Genetics, Rutgers, State University of New Jersey, Piscataway, New Jersey, USA

    Jinchuan Xing

  14. Vascular Program, Institute for Cell Engineering, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA

    Gregg L Semenza

  15. Department of Medical Oncology, Dana-Farber Cancer Institute, Harvard Medical School, Boston, Massachusetts, USA

    Ella Liberzon & William G Kaelin Jr

  16. Howard Hughes Medical Institute, Chevy Chase, Maryland, USA

    Ella Liberzon & William G Kaelin Jr

  17. Departmant of Family and Preventive Medicine, University of Utah School of Medicine, Salt Lake City, Utah, USA

    Andrew Wilson

  18. Division of Physiology, University of California San Diego School of Medicine, La Jolla, California, USA

    Tatum S Simonson

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  1. Felipe R Lorenzo
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Contributions

F.R.L. screened and discovered the PHD2 alteration, designed the experiment, performed expression studies and data analysis, and drafted and edited the manuscript. M.M. and P.K. performed the kinetic studies and wrote and edited the manuscript. C.H., J.X., T.S.S. and L.B.J. analyzed array data, characterized and estimated the origin of the variants and edited the manuscript. P.A.K. and G.R.-L. hosted F.R.L., T.T. and J.T.P. in their countries, assisted with the recruitment of subjects in India and China, and obtained necessary local regulatory and institutional review board documents. T.W. helped organize and assisted with the collection and phenotyping of samples from the Tibetan plateau. P.G. helped organize collection and assisted in the preparation and extraction of DNA in India. S.S. performed the erythroid colony assays, interpreted their results and edited the manuscript. M.E.S. performed the quantification of mean size and hemoglobinization of BFU-Es. A.W. designed and performed the statistical analysis of changes in BFU-E hypersensitivity to EPO. V.G. and D.A.M. critically revised the concept, contributed to the intellectual content and design, and gave final approval of the manuscript. G.L.S. critically revised the concept, contributed to the intellectual content and wrote the manuscript. B.O., W.G.K., E.L. and T.M.K. performed the knockdown study and edited the manuscript. J.T.P. conceived and designed the study, analyzed the data, and wrote and edited the manuscript.

Corresponding author

Correspondence to Josef T Prchal.

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The authors declare no competing financial interests.

Integrated supplementary information

Supplementary Figure 1 Genetic relationship between local Tibetans and other East Asian populations.

(A) Principal-components analysis. The first two principal components (PCs) are shown. Each individual is represented by one dot, and color corresponds to the population. The percentage of variance explained by each PC is shown on the axis. (b) Individual grouping inferred by ADMIXTURE. Results from k = 4 are shown. Each individual’s genome is represented by a vertical bar composed of colored sections, where each section represents the proportion of an individual’s ancestry derived from one of the k ancestral populations. Individuals are arrayed horizontally and grouped by population as indicated. JPT, Japanese; CHB, Chinese; Local Tibetan, Tibetans collected in this study. The two Local Tibetans who have the wild-type D4E variant (TU09 and TU12) are indicated in the plots.

Supplementary Figure 2 Analysis of purified wild-type PHD2 enzyme, the D4E and C127S single mutants, and the D4E and C127S double mutant.

Recombinant enzymes were expressed in insect cells and were affinity purified exploiting their C-terminal Flag tags in anti-Flag affinity gel columns. The near-to-homogeneity purified enzymes were analyzed by 10% SDS-PAGE followed by Coomassie Blue staining. Molecular weight markers (kDa) are shown on the left.

Supplementary Figure 3 Expression study of hypoxia and several HIF target genes.

(a) Expression of hypoxia-regulated genes in granulocytes from Tibetan (n = 4) versus control (n = 6) subjects. (b) BFU-E expression of selected HIF target genes in Tibetan (n = 1) versus control (n = 2) erythroid progenitors grown at optimal EPO levels (3,000 mU). Two independent experiments (technical replicates) from BFU-Es were pooled together and utilized the statistical randomization test using the Relative Expression Software Tool (REST, Qiagen). Error bars represent s.d.

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Lorenzo, F., Huff, C., Myllymäki, M. et al. A genetic mechanism for Tibetan high-altitude adaptation. Nat Genet 46, 951–956 (2014). https://doi.org/10.1038/ng.3067

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