Abstract
Drosophila spermatocytes grow up to 25 times their original volume before the onset of male meiosis. Several insulin-like peptides and their cognate receptors (InR) are essential for the cell growth process in Drosophila. Here, we aimed to identify additional signaling pathways and other regulatory factors required for germline cell growth in Drosophila males. Spermatocyte-specific expression of the dominant-negative form of InR inhibits cell growth. Conversely, constitutively active forms of signaling factors downstream of InR suppress growth inhibition. Furthermore, hypomorphic mutations in the target of rapamycin (Tor) inhibit spermatocyte growth. These data indicate that the insulin/TOR pathway is essential for the growth of premeiotic spermatocytes. RNA interference (RNAi) screening for the identification of other novel genes associated with cell growth showed that the silencing of each of the five members of heat shock cognate 70 (Hsc70) genes significantly inhibited the process. Hsc70-silenced spermatocytes showed Akt inhibition downstream of the insulin signaling pathway. Our pleckstrin homology domain-green fluorescent protein (PH-GFP) reporter studies indicated that PI3K remained activated in Hsc70-4-silenced cells, suggesting that the Hsc70-4 protein possibly targets Akt or Pdk1 acting downstream of PI3K. Moreover, each of the Hsc70 proteins showed different subcellular localizations. Hsc70-2 exhibited cytoplasmic colocalization with Akt in spermatocytes before nuclear entry of the kinase during the growth phase. These results indicated the involvement of Hsc70 proteins in the activation of various steps in the insulin signaling pathway, which is essential for spermatocyte growth. Our findings provide insights into the mechanism(s) that enhance signal transduction to stimulate the growth of Drosophila spermatocytes.
