In regenerative tissues, one of the strategies to protect stem cells from genetic aberrations, potentially caused by frequent cell division, is to transiently expand the stem cell daughters before further differentiation. However, failure to exit the transit amplification may lead to overgrowth, and the molecular mechanism governing this regulation remains vague. In a Drosophila mutagenesis screen for factors involved in the regulation of germline stem cell (GSC) lineage, we isolated a mutation in the gene CG32364, which encodes a putative RNA-binding protein (RBP) and is designated as tumorous testis (tut). In tut mutant, spermatogonia fail to differentiate and over-amplify, a phenotype similar to that in mei-P26 mutant. Mei-P26 is a TRIM-NHL tumor suppressor homolog required for the differentiation of GSC lineage. We found that Tut binds preferentially a long isoform of mei-P26 3′UTR, and is essential for the translational repression of mei-P26 reporter. Bam and Bgcn are both RBPs that have also been shown to repress mei-P26 expression. Our genetic analyses indicate that tut, bam, or bgcn is required to repress mei-P26 and to promote the differentiation of GSCs. Biochemically, we demonstrate that Tut, Bam, and Bgcn can form a physical complex in which Bam holds Tut on its N-terminus and Bgcn on its C-terminus. Our in vivo and in vitro evidence illustrate that Tut acts with Bam, Bgcn to accurately coordinate proliferation and differentiation in Drosophila germline stem cell lineage.

Hippo pathway and its related genes are required for growth control in various somatic tissues. The mutations of Hippo pathway components lead to tissue overgrowth cell-autonomously. Surprisingly, when we generated germline mutant clones of Hippo-network genes such as fat, expanded, hippo, salvador, and warts, we did not observe any overgrowth of these mutant cells. Consistently, overexpression of the progrowth gene yorkie, which is normally inhibited by Hippo signaling, did not lead to germline overgrowth either. In contrast to previous studies in epithelial tissues, these tumor suppressor genes are dispensable in germline cells for their proliferation control. Furthermore, we demonstrate that expanded functions nonautonomously to regulate spermatogonial proliferation. It appears that expanded acts from the somatic support cells surrounding the germline to restrict spermatogonial amplification. Developmental Dynamics 237:270 -275, 2008.