High-Resolution Profiling of Novel Transcribed Regions During Rat Spermatogenesis
Mammalian spermatogenesis is a complex and highly orchestrated combination of processes in which male germline proliferation and differentiation result in the production of mature spermatozoa. If recent genome-wide studies have contributed to the in-depth analysis of the male germline protein-coding transcriptome, little effort has yet been devoted to the systematic identification of novel unannotated transcribed regions expressed during mammalian spermatogenesis. We report high-resolution expression profiling of male germ cells in the rat using Illumina next-generation sequencing technology and highly enriched testicular cell populations. Among 20,424 high-confidence transcripts reconstructed, we defined a stringent set of 1,419 long multi-exonic unannotated transcripts expressed in the testis, named TUTs. These were classified into seven groups with different expression patterns. The vast majority of TUTs share many of the characteristics of vertebrate long non-coding RNAs (lncRNAs). We also markedly reinforced the finding that TUTs and known lncRNAs accumulate during the meiotic and postmeiotic stages of spermatogenesis in mammals, and that X-linked meiotic TUTs do not escape the silencing effects of meiotic sex chromosome inactivation. Importantly, we discovered that TUTs and known lncRNAs with a peak expression during meiosis define a distinct class of non-coding transcripts that exhibit exons twice as long as those of other transcripts. Our study provides new insights in transcriptional profiling of the male germline and represents a high-quality resource of novel loci expressed during spermatogenesis that significantly contributes to the rat genome annotation. A graphical display of the data is conveniently accessible through the ReproGenomics Viewer (RGV) at http://rgv.genouest.org. Overall design: Genome-wide expression profiling analysis using Illumina next-generation sequencing technology