关键词: CG8005基因, 果蝇睾丸, 生殖细胞, 自我更新与分化, 增殖与凋亡

Abstract:

The processes of self-renewal and differentiation of germ cells are crucial to the development of male infertility and germ cell tumors. CG8005 gene is one of the regulatory factors of the testicular germ stem cells in Drosophila melanogaster, and its functional mechanism is still unknown. To explore the biological function(s) of CG8005 gene in the germ cell niche of Drosophila testis, first, the UAS-gal4 system was used to drive the expression of UAS-CG8005 RNAi in Drosophila testicular germ cells and cyst cells. Fertility tests were then performed to determine the fertility rate of male flies. Second, the expression patterns of Vasa, IBI, Zn finger homeodomain 1 (Zfh1), eyes absent (Eya), DE-cad, FasIII and Phospho-Histone H3(PH3), and TUNEL were analyzed by immunofluorescence staining in both control and CG8005 RNAi testes. Lastly, small interfering RNA (siRNA) was used to silence the CG8005 gene expression in Drosophila S2 cells; and PH3 was used to detect the proliferation ability of Drosophila S2 cells in the control group and CG8005 siRNA group. Apoptosis of Drosophila S2 cells was analyzed with TUNEL and flow cytometry. To observe the relative expression of the spliceosome, the mRNA levels of the spliceosome subunits were detected by fluorescence quantitative RT-PCR. As compared with the control group, the results showed that deletion of the CG8005 gene in the germ cells and cyst cells of the testis reduced or even completely abolished the fertility of male fruit flies. In addition, nos-gal4 driven UAS-CG8005 RNAi led to loss of fusomes and reduce the proliferative ability of germ cells. Noticeably, tj-gal4-directed UAS-CG8005 RNAi knockdown of CG8005 gene in the testis led to germ cell tumor development. Knockdown of CG8005 gene in Drosophila S2 cells resulted in increase in apoptosis and inhibition of proliferation. Further, the silencing of the CG8005 gene in Drosophila S2 cells caused increases in the mRNA levels of the spliceosome subunits. Hence, CG8005 gene is essential for the self-renewal and differentiation of germ cells in Drosophila testis. Its deletion may lead to restricted germ cell survival and the formation of germ cell-like tumors. CG8005 gene can participate in the regulation of proliferation and apoptosis of Drosophila S2 cells, which is essential for the maintenance of cell life, and might competitively regulate the mRNA levels of spliceosome subunits.

Key words: CG8005, Drosophila testis, germ cells, self-renewal and differentiation, proliferation and apoptosis