TSSK homologue regulates the expression ofprotamineandmosquito testes specificgenes inAnopheles stephensi

The testis-specific serine/threonine-protein kinase ( tssk ) gene is known to play an important role in spermiogenesis in mammals. This study identifies tssk homologues, As_tssk3 and Aea_tssk1 in Anopheles stephensi and Aedes aegypti , respectively. Both As_tssk3 and Aea_tssk1 were found to express in a male-specific manner throughout the development. Efficient knockdown in the expression of As_tssk3 gene was observed upon feeding An. stephensi larvae with target dsRNA. As_tssk3 knockdown led to a reduced sperm reservoir in male adults. Additionally, the knockdown of As_tssk3 resulted in the reduced expression of two male specifically expressed genes, protamine & mosquito testes specific ( mts- a sperm-specific marker) homologues of An. stephensi . In-silico interaction analysis suggests that As_TSSK3 protein interacts with the Nucleosome Assembly Protein1 like-4 (NAP1like-4), a putative transcription factor. We hypothesize that As_TSSK3 regulates the transcription of protamine and mts genes via NAP, as both NAP and protamine have previously been shown to function in the establishment and maintenance of chromatin dynamics, in independent studies. The study presented here is the first report of characterisation of tssk homologues in any mosquito species. The knockout studies and phosphorylation assays would provide more insights to explore the mechanism by which As_tssk3 regulates protamine and mts genes in Anopheles stephensi . Highlights This study identifies the tssk homologues in Ae. aegypti and An. stephensi and showed their male-specific expression in both the mosquito species. dsRNA mediated knockdown of As_TSSK3 resulted into the males with a reduced size of sperm reservoir. dsRNA mediated knockdown of As_TSSK3 led to the reduction in the expression of As_prot and As_mts , homologues of protamine and mosquito testis specific, genes in An. stephensi . In-silico studies suggest that As_TSSK3 regulates the expression of As_prot and As_mts genes via NAP.