Through mutagenesis by P-element transposition, we identified a series of mutants with deletions in topoisomerase 3β gene (top3β) and an adjacent, previously uncharacterized gene CG15897, here named wuho (wh). Whereas top3β truncation does not affect viability or fertility, wh null mutants display male sterile and female semi-sterile phenotypes. Furthermore, wh mutants can be fully rescued by wh transgenes, but not by top3β transgenes, suggesting that the fertility phenotypes are caused by wh deletion. The alignment of WH protein sequence with other eukaryotic putative homologues shows they are evolutionarily conserved proteins with 5 WD40 repeats in the middle portion of the protein, and a bipartite nuclear localization signal at the carboxyl terminus. Yeast homologue with 5 WD40 repeats, Trm82, is the non-catalytic subunit of a tRNA methylase. Immunostaining shows that WH has the highest expression in hub cells, a niche for germline stem cells of testis. However, WH is not required for the maintenance of hub cells or the germline stem cells. In wh mutant males, spermatogenesis is arrested at the elongating stage of the developing spermatids, resulting in an absence of mature sperms in the seminal vesicles. The decreased fertility in wh mutant females is mostly due to defects in oogenesis. There are abnormal egg chambers present in the mutant females, in which the cystocytes fail to arrest their cell division at the fourth mitotic cycle, resulting in more than 16 cells in a single egg chamber. Additionally, these abnormal cystocytes do not undergo multiple rounds of endoreplication as the nurse cells do in a normal egg chamber. Therefore, the cytological analyses demonstrate that wh has a critical function in cellular differentiation for germline cells during gametogenesis.
Bibliographical noteFunding Information:
We thank Haifan Lin for helpful discussion, a gift of vasa antibody, and the use of a fluorescence microscope, Larry Lee and Carrie Reardon for technical assistance. The use of Confocal Microscope Facility and the microinjection by Model System Genomics Facility are acknowledged here. This work is supported by a grant from National Institute of Health (GM29006).
- Hub cells
- Male sterile
- WD40 repeats