Melanocortin 4 receptor signaling and puberty onset regulation in Xiphophorus swordtails.

Fish of the genus Xiphophorus provide a prominent example of genetic control of male body size and reproductive tactics. In X.nigrensis and X.multilineatus, puberty onset and body length are determined by melanocortin 4 receptor (Mc4r) allelic and copy number variations which were proposed to fine-tune the signaling output of the system. Accessory protein Mrap2 is required for growth across species by affecting Mc4r signaling. The molecular mechanism how Mc4r signaling controls puberty regulation in Xiphophorus and whether the interaction with Mrap2 is also involved was so far unclear. Hence, we examined Mc4r and Mrap2 in X.nigrensis and X.multilineatus, in comparison to a more distantly related species, X.hellerii. mc4r and mrap2 transcripts co-localized in the hypothalamus and preoptic regions in large males, small males and females of X.nigrensis, with similar signal strength for mrap2 but higher expression of mc4r in large males. This overexpression is constituted by wild-type and one subtype of mutant alleles. In vitro studies revealed that Mrap2 co-expressed with Mc4r increased cAMP production but did not change EC50. Cells co-expressing the wild-type and one mutant allele showed lower cAMP signaling than Mc4r wild-type cells. This indicates a role of Mc4r alleles, but not Mrap2, in puberty signaling. Different from X.nigrensis and X.multilineatus, X.hellerii has only wild-type alleles, but also shows a puberty onset and body length polymorphism, despite the absence of mutant alleles. Like in the two other species, mc4r and mrap2 transcripts colocalized and mc4r is expressed at substantially higher levels in large males. This demonstrates that puberty and growth regulation mechanism may not be identical even within same genus.