Selection, trans-species polymorphism, and locus identification of major histocompatibility complex class IIβ alleles of New World ranid frogs

Abstract

Genes encoded by the major histocompatibility complex (MHC) play key roles in the vertebrate immune system. However, our understanding of the evolutionary processes and underlying genetic mechanisms shaping these genes is limited in many taxa, including amphibians, a group currently impacted by emerging infectious diseases. To further elucidate the evolution of the MHC in frogs (anurans) and develop tools for population genetics, we surveyed allelic diversity of the MHC class II β1 domain in both genomic and complementary DNA of seven New World species in the genus Rana (Lithobates). To assign locus affiliation to our alleles, we used a “gene walking” technique to obtain intron 2 sequences that flanked MHC class IIβ exon 2. Two distinct intron sequences were recovered, suggesting the presence of at least two class IIβ loci in Rana. We designed a primer pair that successfully amplified an orthologous locus from all seven Rana species. In total, we recovered 13 alleles and documented trans-species polymorphism for four of the alleles. We also found quantitative evidence of selection acting on amino acid residues that are putatively involved in peptide binding and structural stability of the β1 domain of anurans. Our results indicated that primer mismatch can result in polymerase chain reaction (PCR) bias, which influences the number of alleles that are recovered. Using a single locus may minimize PCR bias caused by primer mismatch, and the gene walking technique was an effective approach for generating single-copy orthologous markers necessary for future studies of MHC allelic variation in natural amphibian populations.

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References

  1. Arkush KD, Giese AR, Mendonca HL, McBride AM, Marty GD, Hedrick PW (2002) Resistance to three pathogens in the endangered winter-run chinook salmon (Oncorhynchus tshawytscha): effects of inbreeding and major histocompatibility complex genotypes. Can J Fish Aquat Sci 59:966–975

    Article  Google Scholar 

  2. Babik W (2009) Methods for MHC genotyping in non-model vertebrates. Mol Ecol Resour 10:237–251

    Article  Google Scholar 

  3. Babik W, Pabijan M, Radwan J (2008) Contrasting patterns of variation in MHC loci in the Alpine newt. Mol Ecol 17:2339–2355

    PubMed  CAS  Article  Google Scholar 

  4. Berger L, Speare R, Daszak P, Green DE, Cunningham AA, Goggin CL, Slocombe R, Ragan MA, Hyatt AD, McDonald KR, Hines HB, Lips KR, Marantelli G, Parkes H (1998) Chytridiomycosis causes amphibian mortality associated with population declines in the rain forests of Australia and Central America. Proc Natl Acad Sci USA 95:9031–9036

    PubMed  CAS  Article  Google Scholar 

  5. Bernatchez L, Landry C (2003) MHC studies in nonmodel vertebrates: what have we learned about natural selection in 15 years? J Evol Biol 16:363–377

    PubMed  CAS  Article  Google Scholar 

  6. Bevan MJ (1987) Class discrimination in the world of immunology. Nature 325:192–194

    PubMed  CAS  Article  Google Scholar 

  7. Bos DH, DeWoody JA (2005) Molecular characterization of major histocompatibility complex class II alleles in wild tiger salamanders (Ambystoma tigrinum). Immunogenetics 57:775–781

    PubMed  CAS  Article  Google Scholar 

  8. Bos DH, Waldman B (2006) Evolution by recombination and transspecies polymorphism in the MHC class I gene of Xenopus laevis. Mol Biol Evol 23:137–143

    PubMed  CAS  Article  Google Scholar 

  9. Bradley G, Rosen P, Sredl M, Jones T, Longcore J (2002) Chytridiomycosis in native Arizona frogs. J Wild Dis 38:206–212

    Google Scholar 

  10. Brown JH, Jardetzky TS, Gorga JC, Stern LJ, Urban RG, Strominger JL, Wiley DC (1993) Three-dimensional structure of the human class II histocompatibility antigen HLA-DR1. Nature 364:33–39

    PubMed  CAS  Article  Google Scholar 

  11. Carey C, Cohen N, Rollins-Smith LA (1999) Amphibian declines: an immunological perspective. Dev Comp Immunol 23:459–472

    PubMed  CAS  Article  Google Scholar 

  12. Cereb N, Hughes AL, Yang SY (1997) Locus-specific conservation of the HLA class I introns by intra-locus homogenization. Immunogenetics 47:30–36

    PubMed  CAS  Article  Google Scholar 

  13. Cottage A, Yang A, Maunders H, de Lacy RC, Ramsay NA (2001) Identification of DNA sequences flanking T-DNA insertions by PCR-walking. Plant Mol Biol Rep 19:321–327

    CAS  Article  Google Scholar 

  14. Daszak P, Strieby A, Cunningham AA, Longcore JE, Brown CC, Porter D (2004) Experimental evidence that the bullfrog (Rana catesbeiana) is a potential carrier of chytridiomycosis, and emerging fungal disease of amphibians. Herpetol J 14:201–207

    Google Scholar 

  15. Du Pasquier L, Flajnik MF (1990) Expression of MHC class II antigens during Xenopus development. Dev Immunol 1:85–95

    PubMed  Article  Google Scholar 

  16. Du Pasquier L, Schwager J, Flajnik MF (1989) The immune system of Xenopus. Annu Rev Immunol 7:251–275

    PubMed  Article  Google Scholar 

  17. Ellis SA, Bontrop RE, Antczak DF, Ballingall K, Davies CJ, Kaufman J, Kennedy LJ, Robinson J, Smith DM, Stet SMJ, RJM WMJ, Walter L, Marsh SGE (2006) ISAG/IUIS-VIC Comparative MHC Nomenclature Committee report, 2005. Immunogenetics 57:953–958

    PubMed  Article  Google Scholar 

  18. Fernandez-Soria VM, Morales P, Castro MJ, Suarez B, Recio MJ, Moreno MA, Paz-Artal E, Arnaiz-Villena A (1998) Transcription and weak expression of HLA-DRB6: a gene with anomalies in exon 1 and other regions. Immunogenetics 48:16–21

    PubMed  CAS  Article  Google Scholar 

  19. Flajnik MF, Du Pasquier L (1990) The major histocompatibility complex of frogs. Immunol Rev 113:47–63

    PubMed  CAS  Article  Google Scholar 

  20. Frost DR, Grant T, Faivovich J, Bain RH, Haas A, Haddad CFB, De Sa RO, Channing A, Wilkinson M, Donnellan SC, Raxworthy CJ, Campbell JA, Blotto BL, Moler P, Drewes RC, Nussbaum RA, Lynch JD, Green DM, Wheeler WC (2006) The amphibian tree of life. Bull Am Mus Nat Hist 297:1–370

    Article  Google Scholar 

  21. Garner TW, Perkins M, Govindarajulu P, Seglie D, Walker S, Cunningham AA, Fisher MC (2006) The emerging amphibian pathogen Batrachochytrium dendrobatidis globally infects introduced populations of the North American bullfrog, Rana catesbeiana. Biol Lett 2:455–459

    PubMed  Article  Google Scholar 

  22. Gray MJ, Miller DL, Hoverman JT (2009) Ecology and pathology of amphibian ranaviruses. Dis Aquat Org 87:243–266

    PubMed  Article  Google Scholar 

  23. Green DE, Converse KA, Scgrader AK (2003) Epizootiology of sixty-four amphibian morbidity and mortality events in the USA, 1996–2001. Annals New York Acad Sci 969:323–339

    Article  Google Scholar 

  24. Grimholt U, Larsen S, Nordmo R, Midtlyng P, Kjoeglum S, Storset A, Saebø S, Stet RJM (2003) MHC polymorphism and disease resistance in Atlantic salmon (Salmo salar); facing pathogens with single expressed major histocompatibility class I and class II loci. Immunogenetics 55:210–219

    PubMed  CAS  Article  Google Scholar 

  25. Hauswaldt JS, Stuckas H, Pfautsch S, Tiedemann R (2007) Molecular characterization of MHC class II in a nonmodel anuran species, the fire-bellied toad Bombina bombina. Immunogenetics 59:479–491

    PubMed  CAS  Article  Google Scholar 

  26. Hedrick PW, Kim TJ (2000) Genetics of complex polymorphisms: parasites and maintenance of the major histocompatibility complex variation. In: Krimbas CB (ed) Evolutionary genetics: from molecules to morphology. Cambridge University Press, Cambridge, pp 203–234

    Google Scholar 

  27. Hedrick PW, Parker KM, Lee RN (2001) Using microsatellite and MHC variation to identify species, ESUs, and MUs in the endangered Sonoran topminnow. Mol Ecol 10:1399–412

    PubMed  CAS  Article  Google Scholar 

  28. Hill WG, Robertson A (1968) Linkage disequilibrium in finite populations. Theor Appl Genet 38:226–231

    Article  Google Scholar 

  29. Hughes A, Nei M (1992) Maintenance of MHC polymorphism. Nature 355:402–403

    PubMed  CAS  Article  Google Scholar 

  30. International Union for Conservation of Nature (2010) IUCN red list of threatened species. International Union for Conservation of Nature. http://www.iucnredlist.org/, accessed January 27, 2010

  31. Kaufman J, Salomonsen J, Flajnik MF (1994) Evolutionary conservation of MHC class I and class II molecules—different yet the same. Semin Immunol 6:411–424

    PubMed  CAS  Article  Google Scholar 

  32. Klein J (1987) Origin of major histocompatibility complex polymorphism: the trans-species hypothesis. Hum Immunol 19:155–62

    PubMed  CAS  Article  Google Scholar 

  33. Klein J, Horejsi V (1997) Immunology. Blackwell Science, Oxford

    Google Scholar 

  34. Klein J, Bontrop RE, Dawkins RL, Erlich HA, Gyllensten UB, Heise ER, Jones PP, Parham P, Wakeland EK, Watkins DI (1990) Nomenclature for the major histocompatibility complexes of different species: a proposal. Immunogenetics 31:217–219

    PubMed  CAS  Google Scholar 

  35. Ko W-Y, David R, Akashi H (2003) Molecular phylogeny of the Drosophila melanogaster species subgroup. J Mol Evol 57:562–573

    PubMed  CAS  Article  Google Scholar 

  36. Kobari F, Sato K, Shum BP, Tochinai S, Katagiri M, Ishibashi T, Du Pasquier L, Flajnik MF, Kasahara M (1995) Exon–intron organization of Xenopus MHC class II beta chain genes. Immunogenetics 42:376–385

    PubMed  CAS  Article  Google Scholar 

  37. Langefors Å, Lohm J, Grahn M, Andersen O, von Schantz T (2001) Association between major histocompatibility complex class IIB alleles and resistance to Aeromonas salmonicida in Atlantic salmon. Proc R Soc Lond B 268:479–485

    CAS  Article  Google Scholar 

  38. Lewontin RC (1964) The interaction of selection and linkage. I. Genetic considerations; heterotic models. Genetics 49:49–67

    PubMed  CAS  Google Scholar 

  39. Lips KR, Brem F, Brenes R, Reeve JD, Alford RA, Voyles J, Carey C, Livo L, Pessier AP, Collins JP (2006) Emerging infectious disease and the loss of biodiversity in a neotropical amphibian community. Proc Natl Acad Sci USA 103:3165–3170

    PubMed  CAS  Article  Google Scholar 

  40. Marsh SGE, Parham P, Barber LD (2000) The HLA factsbook. Academic, San Diego

    Google Scholar 

  41. May S, Beebee TJC (2009) Characterisation of major histocompatibility complex class II alleles in the natterjack toad, Bufo calamita. Conservation Genetics Resources 1:415–417

    Article  Google Scholar 

  42. Mayer WE, O’Huigin C, Klein J (1993) Resolution of the HLA-DRB6 puzzle: a case of grafting a de novo generated exon on an existing gene. Proc Natl Acad Sci USA 90:10720–10724

    PubMed  CAS  Article  Google Scholar 

  43. McVean G, Awadalla P, Fearnhead P (2002) A coalescent-based method for detecting and estimating recombination from gene sequences. Genetics 160:1231–1241

    PubMed  CAS  Google Scholar 

  44. Nonaka M, Namikawa C, Kato Y, Sasaki M, Salter-Cid L, Flajnik MF (1997) Major histocompatibility complex gene mapping in the amphibian Xenopus implies a primordial organization. Proc Natl Acad Sci USA 94:5789–579

    PubMed  CAS  Article  Google Scholar 

  45. Nozawa M, Suzuki Y, Nei M (2009) Reliabilities of identifying positive selection by the branch-site and the site-prediction methods. Proc Natl Acad Sci USA 106:6700–6705

    PubMed  CAS  Article  Google Scholar 

  46. Ohta Y, Goetz W, Hossain MZ, Nonaka M, Flajnik MF (2006) Ancestral organization of the MHC revealed in the amphibian Xenopus. J Immunol 176:3674–3685

    PubMed  CAS  Google Scholar 

  47. Paterson S, Wilson K, Pemberton JM (1998) Major histocompatibility complex variation associated with juvenile survival and parasite resistance in a large unmanaged ungulate population (Ovis aries L.). Proc Natl Acad Sci USA 95:3714–3719

    PubMed  CAS  Article  Google Scholar 

  48. Pond SLK, Frost SDW (2005) Not so different after all: a comparison of methods for detecting amino acid sites under selection. Mol Biol Evol 22:1208–1222

    CAS  Article  Google Scholar 

  49. Pounds AJ, Bustamante MR, Coloma LA, Consuegra JA, Fogden MPL, Foster PN, La Marca E, Masters KL, Merino-Viteri A, Puschendorf R, Ron SR, Sanchez-Azofeifa GA, Still CJ, Young BE (2006) Widespread amphibian extinctions from epidemic disease driven by global warming. Nature 439:161–167

    PubMed  CAS  Article  Google Scholar 

  50. Richman AD, Herrera G, Reynoso VH, Méndez G, Zambrano L (2007) Evidence for balancing selection at the DAB locus in the axolotl, Ambystoma mexicanum. Int J Immunogenet 34:475–478

    PubMed  CAS  Article  Google Scholar 

  51. Richmond JQ, Savage AE, Zamudio KR, Rosenblum EB (2009) Toward immunogenetic studies of amphibian chytridiomycosis: linking innate and acquired immunity. BioScience 59:311–320

    Article  Google Scholar 

  52. Ronquist F, Huelsenbeck JP (2003) MrBayes 3: Bayesian phylogenetic inference under mixed models. Bioinformatics 19:1572–1574

    PubMed  CAS  Article  Google Scholar 

  53. Sato K, Flajnik M, Du Pasquier L, Katagiri M, Kasahara M (1993) Evolution of the MHC: isolation of class II beta-chain cDNA clones from the amphibian Xenopus laevis. J Immunol 150:2831–2843

    PubMed  CAS  Google Scholar 

  54. Scheffler K, Martin DP, Seoighe C (2006) Robust inference of positive selection from recombining coding sequences. Bioinformatics 22:2493–2499

    PubMed  CAS  Article  Google Scholar 

  55. Schierup MH, Hein J (2000) Consequences of recombination on traditional phylogenetic analysis. Genetics 156:879–891

    PubMed  CAS  Google Scholar 

  56. Tajima F, Nei M (1984) Estimation of evolutionary distance between nucleotide sequences. Mol Biol Evol 1:269–285

    PubMed  CAS  Google Scholar 

  57. Takahata N (1990) A simple genealogical structure of strongly balanced allelic lines and trans-species evolution of polymorphism. Proc Natl Acad Sci USA 87:2419–2423

    PubMed  CAS  Article  Google Scholar 

  58. Takahata N, Nei M (1990) Allelic genealogy under overdominant and frequency-dependent selection and polymorphism of major histocompatibility complex loci. Genetics 124:967–978

    PubMed  CAS  Google Scholar 

  59. Tamura K, Dudley J, Nei M, Kumar S (2007) MEGA4: molecular evolutionary genetics analysis (MEGA) software version 4.0. Mol Biol Evol 24:1596–1599

    PubMed  CAS  Article  Google Scholar 

  60. Teacher AGF, Garner TWJ, Nichols RA (2009) Evidence for directional selection at a novel major histocompatibility class I marker in wild common frogs (Rana temporaria) exposed to a viral pathogen (Ranavirus). PLoS ONE 4:e4616

    PubMed  Article  Google Scholar 

  61. Tong J, Bramson J, Kanduc D, Chow S, Sinha A, Ranganathan S (2006) Modeling the bound conformation of Pemphigus vulgaris-associated peptides to MHC class II DR and DQ alleles. Immunome Res 2:1

    PubMed  Article  Google Scholar 

  62. van Oosterhout C, Joyce DA, Cummings SM, Blais J, Barson NJ, Ramnarine IW, Mohammed RS, Persad N, Cable J (2006) Balancing selection, random genetic drift, and genetic variation at the major histocompatibility complex in two wild populations of guppies (Poecilia reticulata). Evolution 60:2562–2574

    PubMed  Article  Google Scholar 

  63. Vyas JM, Van der Veen AG, Ploegh HL (2008) The known unknowns of antigen processing and presentation. Nature Rev Immunol 8:607–618

    CAS  Article  Google Scholar 

  64. Wilson DJ, McVean G (2006) Estimating diversifying selection and functional constraint in the presence of recombination. Genetics 172:1411–1425

    PubMed  CAS  Article  Google Scholar 

  65. Zeisset I, Beebee TJC (2009) Molecular characterization of major histocompatibility complex class II alleles in the common frog, Rana temporaria. Mol Ecol Resour 9:738–745

    CAS  Article  Google Scholar 

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Acknowledgments

We thank J. D. Austin and E. Rittmeyer for assistance with specimen collection. This study was supported by Population and Evolutionary Process National Science Foundation Grants DEB-0815315 (to KRZ) and DEB-0909013 (to KRZ and AES). Members of the Zamudio laboratory group made helpful comments on the earlier versions of this manuscript. The core facilities at Cornell University (Evolutionary Genetics Core Facility and Life Sciences Core Laboratories) provided the infrastructure for data collection.

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Correspondence to Karen M. Kiemnec-Tyburczy.

Electronic supplementary material

Additional supporting material for this manuscript is provided online.

Online Resource 1

Locality data for the seven ranid species from which MHC class IIβ alleles were amplified (DOC 31 kb)

Online Resource 2

Text file containing anuran MHC class IIβ sequences used for detecting selection on amino acids (TXT 7 kb)

Online Resource 3

Nucleotide sequences of partial exon 2 and intron 2 obtained from gene walking and subsequent PCR. The reverse complement of the MHC-5R primer sequence is positioned above the alignment. The intron/exon boundary is indicated by the vertical line. The splice donor site is shaded, the dashed box corresponds to the annealing site of the primer MHC-5R, and “–” represent gaps. The boxes outline the sections of the last two sequences in the alignment, Xela-DAB (GenBank accesssion no. D50039) and Raca-D2B*01, that could not be aligned with the other ranid introns (GIF 185 kb)

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Kiemnec-Tyburczy, K.M., Richmond, J.Q., Savage, A.E. et al. Selection, trans-species polymorphism, and locus identification of major histocompatibility complex class IIβ alleles of New World ranid frogs. Immunogenetics 62, 741–751 (2010). https://doi.org/10.1007/s00251-010-0476-6

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Keywords

  • Amphibia
  • Beta chain
  • Gene walking
  • Lithobates
  • Positive selection
  • Rana