Advertisement

Individual Variation in Pheromone Isoform Ratios of the Red-Legged Salamander, Plethodon shermani

  • Adam J. Chouinard
  • Damien B. Wilburn
  • Lynne D. Houck
  • Richard C. Feldhoff
Chapter

Abstract

For more than 15 years, pheromone signalling in plethodontid ­salamanders has served as an important amphibian system for studying how molecules can influence reproductive behaviour. In the red-legged salamander (Plethodon shermani), males utilise proteinaceous pheromones from a submandibular (mental) gland to affect female behaviour and reduce the duration of courtship. The two major proteins, Plethodontid Receptivity Factor (PRF) and Plethodontid Modulating Factor (PMF), exist as multiple isoforms within the gland of an individual male. While previous research has focused on biochemical characterisation of these proteins, this study explores the degree of intraspecific variability of pheromone isoform ratios among males of a single population. Biochemical analyses were performed on the mental gland extracts of individual male salamanders (n = 108) via high-performance liquid chromatography to quantify components of the pheromone mixture and establish the extent of individual variability. The results of these individual analyses revealed that the two main proteins (PMF and PRF) comprise over 80% of the total mixture, with a 5:3 ratio (PMF:PRF). Numerous minor peaks also appeared in the elution range of PRF, which could account for an excess of mRNA transcripts that have not been proteomically characterised. Lastly, these data demonstrate a remarkable diversity of isoform ratios among males. Taken together, these aspects of pheromone complexity and diversity could have profound functional effects on reproductive behaviour in this family of salamanders.

Keywords

Major Isoforms Accessory Olfactory Bulb Ternary Plot Multicomponent Signal Pheromone Composition 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Notes

Acknowledgements

We thank Kathleen Bowen, Pamela Feldhoff, Kari Leichty, Sarah Eddy and Josef Uyeda for help in the lab or field and/or for comments on the manuscript. We also thank James Costa and the staff of the Highlands Biological Station (Highlands, NC). Funding was supported by National Science Foundation grants to LDH (IOS-0818554) and RCF (IOS-0818649).

References

  1. Andersson M (1994) Sexual selection. Princeton University Press, PrincetonGoogle Scholar
  2. Arnold SJ (1976) Sexual behavior, sexual interference, and sexual defense in the salamanders Ambystoma maculatum, Ambystoma tigrinum and Plethodon jordani. Z Tierpsychol 42:247–300CrossRefGoogle Scholar
  3. Beynon RJ, Veggerby C, Payne CE, Robertson DH, Gaskell SJ, Humphries RE, Hurst JL (2002) Polymorphism in major urinary proteins in mouse liver: a biochemical study. Biochem Genet 28(7):1429–1445Google Scholar
  4. Calenge C (2006) The package adehabitat for the R software: a tool for the analysis of space and habitat use by animals. Ecol Model 197:516–519CrossRefGoogle Scholar
  5. Dawley EM (1984a) Identification of sex through odors by male red-spotted newts, Notopthalmus viridescens. Herpetologica 40(2):101–105Google Scholar
  6. Dawley EM (1984b) Recognition of individual, sex, and species odours by salamanders of the Plethodon glutinosusP. jordani complex. Anim Behav 32(2):352–361CrossRefGoogle Scholar
  7. Dawley EM, Bass AH (1988) Organization of the vomeronasal organ in a plethodontid salamander. J Morphol 198:243–355CrossRefGoogle Scholar
  8. Dawley EM, Bass AH (1989) Chemical access to the vomeronasal organs of a plethodontid salamander. J Morphol 200:163–174CrossRefGoogle Scholar
  9. Feldhoff RC, Rollmann SM, Houck LD (1999) Chemical analyses of courtship pheromones in a plethodontid salamander. In: Johnston RE, Muller-Schwarze D, Sorenson PW (eds) Advances in chemical signals in vertebrates. Kluwer Academic/Plenum Publishers, New York, pp 117–125CrossRefGoogle Scholar
  10. Haga S, Hattori T, Sato T, Sato K, Matsuda S, Kobayakawa R, Sakano H, Yoshihara Y, Touhara K (2010) The male mouse pheromone ESP1 enhances female sexual receptive behaviour through a specific vomeronasal receptor. Nature 466:118–122PubMedCrossRefGoogle Scholar
  11. He J, Ma L, Kim S, Schwartz J, Santilli M, Wood C, Durnin MH, Yu CR (2010) Distinct signals conveyed by pheromone concentrations to the mouse vomeronasal organ. J Neurosci 30(22):7473–7483PubMedCrossRefGoogle Scholar
  12. Hefetz A, Graur D (1988) The significance of multicomponent pheromones in denoting specific compositions. Biochem Sys Ecol 16(6):557–566CrossRefGoogle Scholar
  13. Houck LD (2009) Pheromone communication in amphibians and reptiles. Annu Rev Physiol 73:161–176CrossRefGoogle Scholar
  14. Houck LD, Palmer CA, Watts RA, Arnold SJ, Feldhoff PW, Feldhoff RC (2007) A new vertebrate courtship pheromone, PMF, affects female receptivity in a terrestrial salamander. Anim Behav 73:315–320CrossRefGoogle Scholar
  15. Houck LD, Watts RA, Arnold SJ, Bowen KE, Kiemnec KM, Godwin HA, Feldhoff PW, Feldhoff RC (2008) A recombinant courtship pheromone affects sexual receptivity in a plethodontid salamander. Chem Senses 33:623–631PubMedCrossRefGoogle Scholar
  16. Hurst JL (2009) Female recognition and assessment of males through scent. Behav Brain Res 200:295–303PubMedCrossRefGoogle Scholar
  17. Hurst JL, Beynon RJ (2004) Scent wars: the chemobiology of competitive signalling in mice. Bioessays 26:1288–1298PubMedCrossRefGoogle Scholar
  18. Hurst JL, Payne CE, Nevison CM, Marie AD, Humphries RE, Robertson DH, Cavaggioni A, Beynon RJ (2001) Individual recognition in mice mediated by major urinary proteins. Nature 414:631–634PubMedCrossRefGoogle Scholar
  19. Jaeger RG (1981) Dear enemy recognition and the costs of aggression between salamanders. Am Nat 117:962–974CrossRefGoogle Scholar
  20. Kiemnec-Tyburczy KM, Watts RA, Gregg RC, von Borstel D, Arnold SJ (2009) Evolutionary shifts in courtship pheromone composition revealed by EST analysis of plethodontid salamander mental glands. Gene 432:75–81PubMedCrossRefGoogle Scholar
  21. Kiemnec-Tyburczy KM, Woodley SK, Watts RA, Arnold SJ, Houck LD (2012) Expression of vomeronasal receptors and related signaling molecules in the nasal cavity of a caudate amphibian (Plethodon shermani). Chem Senses 37:335–346PubMedCrossRefGoogle Scholar
  22. Kikuyama S, Toyoda F, Ohmiya Y, Matsuda K, Tanaka S, Hayashi H (1995) Sodefrin: a female-attracting peptide pheromone in newt cloacal glands. Science 267:1643–1645PubMedCrossRefGoogle Scholar
  23. Laberge F, Feldhoff RC, Feldhoff PW, Houck LD (2008) Courtship pheromone induced c-Fos-like immunolabeling in the female salamander brain. Neuroscience 151:329–339PubMedCrossRefGoogle Scholar
  24. Leinders-Zufall T, Lane AP, Puche AC, Weidong M, Novotny MV, Shipley MT, Zufall F (2000) Ultrasensitive pheromone detection by mammalian vomeronasal neurons. Nature 405:792–796PubMedCrossRefGoogle Scholar
  25. Linn CE, Campbell MG, Roelofs WL (1985) Male moth sensitivity to multicomponent pheromones: critical role of female-released blend in determining the functional role of components and active space of the pheromone. J Chem Ecol 12(3):659–668CrossRefGoogle Scholar
  26. Linn CE, Campbell MG, Roelofs WL (1987) Pheromone components and active spaces: what do moths smell and where do they smell it? Science 237:650–652PubMedCrossRefGoogle Scholar
  27. Meyer D, Zeileis A, Hornik K (2006) The strucplot framework: visualizing multi-way contingency tables with vcd. J Stat Softw 17(3):1–48Google Scholar
  28. Palmer CA, Hollis DM, Watts RA, Houck LD, McCall MA, Gregg RG, Feldhoff PW, Feldhoff RC, Arnold SJ (2007a) Plethodontid modulating factor, a hypervariable salamander courtship pheromone in the three-finger protein superfamily. FEBS J 274:2300–2310PubMedCrossRefGoogle Scholar
  29. Palmer CA, Watts RA, Houck LD, Picard AL, Arnold SJ (2007b) Evolutionary replacement of components in a salamander pheromone signaling complex: more evidence for phenotypic-molecular decoupling. Evolution 61(1):202–215PubMedCrossRefGoogle Scholar
  30. Palmer CA, Watts RA, Hastings AP, Houck LD, Arnold SJ (2010) Rapid evolution of Plethodontid Modulating Factor, a hypervariable salamander courtship pheromone, is driven by positive selection. J Mol Evol 70:427–440PubMedCrossRefGoogle Scholar
  31. Rollmann SM, Houck LD, Feldhoff RC (1999) Proteinaceous pheromone affecting female receptivity in a terrestrial salamander. Science 285:1907–1909PubMedCrossRefGoogle Scholar
  32. Rollmann SM, Houck LD, Feldhoff RC (2000) Population variation in salamander courtship pheromones. J Chem Ecol 26(12):2713–2724CrossRefGoogle Scholar
  33. Schmidt A, Roth G (1990) Central olfactory and vomeronasal pathways in salamanders. J Hirnforsch 31(5):543–553PubMedGoogle Scholar
  34. Simon G, Madison DM (1984) Individual recognition in salamanders: cloacal odours. J Herpetol 12(3):397–406Google Scholar
  35. Smith PK, Krohn RI, Hermanson GT, Mallia AK, Gartner FH, Provenzano MD, Fujimoto EK, Goeke NM, Olson BJ, Klenk DC (1985) Measurement of protein using bicinchoninic acid. Anal Biochem 150:76–85PubMedCrossRefGoogle Scholar
  36. Swanson WJ, Vacquier VD (2002) The rapid evolution of reproductive proteins. Nat Rev Gen 3:137–144CrossRefGoogle Scholar
  37. Watts RA, Palmer CA, Feldhoff RC, Feldhoff PW, Houck LD, Jones AG, Pfrender ME, Rollmann SM, Arnold SJ (2004) Stabilizing selection on behavior and morphology masks positive selection on the signal in a salamander pheromone signaling complex. Mol Biol Evol 21(6):1032–1041PubMedCrossRefGoogle Scholar
  38. Wilburn DB, Bowen KE, Gregg RG, Cai J, Feldhoff PW, Houck LD, Feldhoff RC (2012) Proteomic and UTR analyses of a rapidly evolving hypervariable family of vertebrate pheromones. Evolution 66:2227–2239PubMedCrossRefGoogle Scholar
  39. Wirsig-Wiechmann C, Houck LD, Feldhoff PW, Feldhoff RC (2002) Pheromonal activation of vomeronasal neurons in plethodontid salamanders. Brain Res 952:335–344PubMedCrossRefGoogle Scholar
  40. Wirsig-Wiechmann C, Houck LD, Wood JM, Feldhoff PW, Feldhoff RC (2006) Male pheromone protein components activate female vomeronasal neurons in the salamander Plethodon shermani. BMC Neurosci 7:26–34PubMedCrossRefGoogle Scholar
  41. Wyatt TD (2003) Pheromones and animal behaviour: communication by smell and taste. Cambridge University Press, CambridgeCrossRefGoogle Scholar
  42. Wyatt TD (2010) Pheromones and signature mixtures: defining species-wide signals and variable cues for identity in both invertebrates and vertebrates. J Comp Physiol A 196:685–700CrossRefGoogle Scholar
  43. Zufall F, Kelliher KR, Leinders-Zufall T (2002) Pheromone detection by mammalian vomeronasal neurons. Microsc Res Techniq 58:251–260CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2013

Authors and Affiliations

  • Adam J. Chouinard
    • 1
  • Damien B. Wilburn
    • 2
  • Lynne D. Houck
    • 1
  • Richard C. Feldhoff
    • 2
  1. 1.Department of ZoologyOregon State UniversityCorvallisUSA
  2. 2.Department of BiochemistryUniversity of LouisvilleLouisvilleUSA

Personalised recommendations