Piwi regulates the usage of alternative transcription start sites in the Drosophila ovary

Transcription from alternative transcription start sites (TSSs) is prevalent among mammalian genes and has important biological functions. However, how TSSs are selected remains elusive. We identified 87 genes with altered TSS usage (ATU) in Piwi-deficient Drosophila ovaries using the cap-analysis gene expression sequencing (CAGE-seq) method. Piwi regulates TSS usage of ATU genes in germ cells and somatic cells in ovaries as well as in cultured ovarian somatic cells (OSCs). Correspondingly, RNA Pol II initiation and elongation at TSSs of ATU genes were affected in germline-piwi-knockdown ovaries and piwi-knockdown OSCs. We identified a FACT complex component, Ssrp, as a novel interactor of Piwi in the nucleus. Temporally controlled knockdown of ssrp affected TSS usage of ATU genes whereas overexpression of ssrp partially rescued the TSS usage of ATU genes in piwi mutant ovaries. Thus, Piwi may interact with Ssrp to regulate TSS usage in Drosophila ovaries by affecting Pol II initiation and elongation. To investigate how Piwi affected Ssrp in germ cells, chromatin immunoprecipitation DNA-sequencing (ChIP-seq) was conducted to profile the occupancy of Ssrp in young germline-egfp-knockdown and germline-piwi-knockdown ovaries.