RNA helicase D1PAS1 resolves R-loops and forms a complex for mouse pachytene piRNA biogenesis required for male fertility
To investigate the function of RNA helicase D1PAS1 in mouse spermatogenesis, we established D1Pas1 knockout (KO) and D1Pas1-FLAG knockin (KI) lines in which the D1Pas1 gene has been knocked out by deleting the transcription start site and the 3XFLAG sequences added at the end of C-term by CRISPR/Cas9. We then performed gene expression profiling analysis using data obtained from RNA-seq of 3 different biological replicates at three different developmental stages. Investigated small RNA abundance from WT and KO testis via small RNA-seq. To profile the genome-wide D1PAS1 RNA helicase dependent R-loop loci, we performed BisMapR sequencing analysis with D1PAS1 KI and KO spermatocytes. Overall design: Expression profiling of differentially expressed genes by RNA-seq at three different developmental stages, small RNA abundance by small RNA-seq, genome-wide RNA:DNA hybrids (R-loop) profiling by BisMapR, and D1PAS1-bound RNA by eCLIP-sequencing.