Srlp is crucial for the self-renewal and differentiation of germline stem cells via RpL6 signals in Drosophila testes

Abstract

Self-renewal and differentiation in germline stem cells (GSCs) are tightly regulated by the stem cell niche and via multiple approaches. In our previous study, we screened the novel GSC regulatory gene Srlp in Drosophila testes. However, the underlying mechanistic links between Srlp and the stem cell niche remain largely undetermined. Here, using genetic manipulation of the Drosophila model, we systematically analyze the function and mechanism of Srlp in vivo and in vitro. In Drosophila , Srlp is an essential gene that regulates the self-renewal and differentiation of GSCs in the testis. In the in vitro assay, Srlp is found to control the proliferation ability and cell death in S2 cells, which is consistent with the phenotype observed in Drosophila testis. Furthermore, results of the liquid chromatography-tandem mass spectrometry (LC-MS/MS) reveal that RpL6 binds to Srlp. Srlp also regulates the expression of spliceosome and ribosome subunits and controls spliceosome and ribosome function via RpL6 signals. Collectively, our findings uncover the genetic causes and molecular mechanisms underlying the stem cell niche. This study provides new insights for elucidating the pathogenic mechanism of male sterility and the formation of testicular germ cell tumor.

RpS13 controls the homeostasis of germline stem cell niche through Rho1‐mediated signals in the Drosophila testis

“…20,21 And ribosomal subunits could act as an intermediary to affect GSC self-renewal and differentiation in the Drosophila testis. 22 We showed that both RpS13 and Rho1 could regulate the expression of ribosome subunits. Consequently, it is highly possible that Therefore, these results indicate that RpS13, together with the small GTPase Rho1, are crucial for GSC self-renewal and differentiation.…”

“…Immunofluorescence assays were carried out as described previously. 22 Testes were dissected in 1 × PBS and fixed for 30 minutes in 4% paraformaldehyde (PFA), then washed 3 times with 0.3% PBS-Triton X-100 (PBST) and blocked for 1 hour in 5% bovine serum albumin (BSA) (Sangon Biotech, Shanghai, China). Next, the testes were incubated overnight at 4°C with primary antibodies (Table S3) diluted in 5% BSA.…”

Precursor RNA processing 3 is required for male fertility, and germline stem cell self-renewal and differentiation via regulating spliceosome function in Drosophila testes

“…S2 cells were collected after transfection for 48 h. Cell apoptosis was determined using the TUNEL assay according to the manufacturer’s protocols 37 . The TUNEL BrightRed Apoptosis Detection Kit was obtained from Vazyme (A113, Nanjing, China).…”

“…S2 cells were collected after transfection for 48 h. Flow cytometry was performed using an Annexin V/PI Apoptosis Assay Kit (FMSAV647-100, FcMACS, Nanjing, China) 37 . After transfection for 48 h, S2 cells were washed with ice-cold PBS.…”

A second hybrid-binding domain modulates the activity of Drosophila ribonuclease H1

“…Raw data for each replicate is shown in Table S2. In each case, we retained target proteins that were identified in every replicate experiment with the given bait protein (n=5 in each case), but which and were absent from all controls (45). The primary screen revealed a list of 63 proteins that we subdivided into two main groups, nuclear (Table 4) and mitochondrial candidates ( Table 5), based on the major subcellular location of the protein as currently annotated in Flybase (www.flybase.org).…”