The thermo-sensitive gene expression signatures of spermatogenesis

Background: Spermatogenesis in most mammals (including human and rat) occurs at similar to 3 degrees C lower than body temperature in a scrotum and fails rapidly at 37 degrees C inside the abdomen. The present study investigates the heat-sensitive transcriptome and miRNAs in the most vulnerable germ cells (spermatocytes and round spermatids) that are primarily targeted at elevated temperature in a bid to identify novel targets for contraception and/or infertility treatment. Methods: Testes of adult male rats subjected to surgical cryptorchidism were obtained at 0, 24, 72 and 120 h post-surgery, followed by isolation of primary spermatocytes and round spermatids and purification to >90% purity using a combination of trypsin digestion, centrifugal elutriation and density gradient centrifugation techniques. RNA isolated from these cells was sequenced by massive parallel sequencing technique to identify the most-heat sensitive mRNAs and miRNAs. Results: Heat stress altered the expression of a large number of genes by >= 2.0 fold, out of which 594 genes (286 up arrow;308 down arrow) showed alterations in spermatocytes and 154 genes (105 up arrow;49 down arrow) showed alterations in spermatids throughout the duration of experiment. 62 heat-sensitive genes were common to both cell types. Similarly, 66 and 60 heat-sensitive miRNAs in spermatocytes and spermatids, respectively, were affected by >= 1.5 fold, out of which 6 were common to both the cell types. Conclusion: The study has identified Acly, selV, SLC16A7(MCT-2), Txnrd1 and Prkar2B as potential heat sensitive targets in germ cells, which may be tightly regulated by heat sensitive miRNAs mo-miR-22-3P, mo-miR-22-5P, mo-miR-129-5P, mo-miR-3560, mo-miR-3560 and mo-miR-466c-5P.