Identification of a promoter region generating Sry circular transcripts both in germ cells from male adult mice and in male mouse embryonal gonads.

The mouse testis determining gene Sry is expressed in somatic cells of the differentiating male gonad as a linear transcript, encoding a transcription factor containing an HMG box. In the adult mouse testis, Sry expression occurs in meiotic and postmeiotic germ cells. The mouse genomic Sry locus is characterized by two arms of a large inverted repeat, flanking a unique region that, between an acceptor and a donor splice site, contains a single exon encoding the Sry protein. In germ cells from the adult mouse testis, Sry RNA is a circular molecule, which is generated by an inverted splicing event that utilizes the above-mentioned splice sites. Thus, a circular exon is spliced out starting from a large linear RNA precursor containing both arms of the inverted repeat, which pair and generate a large stem-loop structure. Using reverse transcription-polymerase chain reaction and an RNase protection assay, we have now mapped the 5' end of this precursor RNA in the 5' arm of the inverted repeat. Gel mobility shift assay and in vitro transcription with nuclear extracts from adult germ cells further confirm that a region immediately 5' upstream of two transcriptional initiation sites of the precursor RNA contains a promoter sequence in which two consensus Sry binding sequences are specifically recognized by nuclear factors present in adult germ cells but not in Sertoli cells. We also show that the linear precursor of the Sry circular transcript and its splicing product are specifically expressed not only in adult germ cells but also in male embryonal gonads between 11.5 and 13.5 days postcoitum, immediately after the expression of the linear transcript starting from the unique region.