Non-hominid TP63 lacks retroviral LTRs but contains a novel conserved upstream exon

We have recently identified novel isoforms of human p63, with specific expression in testicular germ cells. The synthesis of these p63 mRNA species is driven by the long terminal repeat (LTR) of the endogenous retrovirus ERV9. This LTR was inserted upstream of the previously known TP63 exons roughly 15 million years ago, leading to the expression of novel exons and the synthesis of germline-specific transactivating p63 (GTAp63) isoforms in humans and great apes (Beyer et al. Proc Natl Acad Sci USA 2011; 108:3624-9). However, this study did not reveal whether similar upstream exons can also be found in the TP63 genes of non-hominid animals. Here we performed rapid amplification of cDNA ends (RACE) to identify a novel upstream exon of murine TP63, located in the 5' position from the previously described start of transcription. This exon, termed "exon U3" in our previous publication, is conserved within a broad range of mammalian species, including hominids. However, in contrast to the human TP63 gene structure, the murine exon U3 represented the most upstream transcribed sequence of TP63. Murine exon U3 is then alternatively spliced to acceptor sites within exon 1 or upstream of exon 2, resulting in two different available translational start sites. p63 mRNAs comprising exon U3 are detectable in various tissues, with no particular preference for testicular cells. Thus, whereas the retroviral LTR in hominid species results in strictly germline-associated p63 isoforms, the upstream exon in non-hominids fails to confer this tissue specificity. This notion strongly supports the concept that the synthesis of a testis-specific p63 isoform is a recently acquired, unique feature of humans and great apes.