Application of transcriptional and biological network analyses in mouse germ-cell transcriptomes

Serial analysis of gene expression (SAGE) provides a global analysis platform for profiling mRNA populations present in cells of interest without the constraint of gene selection and the ambiguous nature of data obtained. However, most of the reports on SAGE and germ cell development are limited to descriptive analyses. Here, we report a series of bioinformatic analyses using recently published SAGE data on the transcriptome of mouse type A spermatogonia (Spga), pachytene spermatocytes (Spcy), and round spermatids (Sptd). Tags with a total count of ≥20 in three SAGE libraries were examined. Our aim was to identify and discover potential transcriptional regulators and pathways involved at different stages of spermatogenesis. Unsupervised hierarchical clustering based on tag expression and Gene Ontology analysis were applied to identify genes and biological processes overrepresented at a particular stage of development. The 5′ cis-regulatory elements were examined for common regulators in different functional clusters. Potential biological networks were also constructed to reveal the link between the gene candidates. Biological pathways related to the three germ cell stages were constructed. A number of known transcription regulators in spermatogenesis, including NF-κB, SP1, AP-1, and EGR, were identified. Novel promoter elements such as the E box in Spga-specific genes, GATA in Spcy-specific genes, and GKLF in Sptd-specific genes were also observed. Taken together, our approach is reliable and provides a foundation for the generation of novel biological hypotheses for studying spermatogenesis.