Comparative structure and characterization of a CDKN2 gene in a Xiphophorus fish melanoma model

We have cloned, sequenced, and characterized the RNA expression properties of a fish CDKN2 gene from Xiphophorus us helleri and X. maculatus. This gen e, termed CDKN2X, shows a high degree of amino acid sequence similarity to members of the mammalian CDKN2 gene family, which includes the tumor suppre ssor loci CDKN2A (P16) and CDKN2B (P15). Comparative sequence analysis sugg ests that fish CDKN2X is similarly related to all four mammalian gene famil y members, and may represent a descendant of an ancestral prototypic CDKN2 gene. CDKN2X was mapped to a region on autosomal Xiphophorus linkage group V (LG V) known to contain the DIFF gene that acts as a tumor suppressor of melanoma formation in X. helleri/X. maculatus backcross hybrids. Thus, CDKN 2X may be a candidate for the tumor suppressor DIFF gene. Here we have sequ enced CDKN2X in both Xiphophorus species and have characterized its express ion in normal. and melanotic tissues within control and backcross hybrid fi sh. A simultaneous expressional analysis of the Xmrk-2 tyrosine kinase rece ptor gene, which is strongly implicated in melanomagenesis in this system, was also performed. RT-PCR analyses revealed that both genes were highly ex pressed in melanomas. For CDKN2X, this result contrasts numerous findings i n human tumors including human melanoma in which either CDKN2A (P16) deacti vation or LOH was observed.