The yeast gene controlling over the initiation of sexual reproduction

裂解酵母ste13^基因编码依赖于ATP的RNA解旋酶，是响应氮饥饿信号而启动有性繁殖所必需的。通过对ste13基因及其产物的分析，验证了我们的假设，即ste13在转录后水平上调控了基因在性周期中的表达，得到了以下结果：(1)ste13的体外诱变：ste13缺失突变体在交配和减数分裂过程中存在缺陷，但能够在丰富的培养基中生长。在RNA解旋酶的共同模体中引入定点突变导致了不育的表型。这些事实表明，ste13基因对有丝分裂细胞的分裂不是必需的，但对性周期的诱导是必不可少的。(2)ste13蛋白的细胞内定位：GST-ste13融合蛋白在大肠杆菌中得到了高效表达。以亲和纯化的融合蛋白为抗原，制备多克隆抗体。Western印迹分析表明，该抗体可识别庞氏葡萄球菌细胞提取物中的58 kDa多肽。免疫荧光显微镜显示，Ste13蛋白未定位于胞核，而分布于细胞质。

The fission yeast ste13^+ gene which encodes a putative ATP-dependent RNA helicase is essential for initiation of sexual reproduction in response to nitrogen-starvation signal. The ste13^+ gene and its products were analyzed to verify our hypothesis that Ste13 regulates gene expression during sexual cycle at a post-transcriptional level and the following results were obtained.(1)In vitro mutagenesis of ste13 :The ste13 null mutant was defective in mating and meiosis but able to grow in rich medium. Site-directed mutagenesis introduced into the consensus motifs of RNA helicases caused the sterile phenotype. These facts indicate that the ste13 gene is not essential for mitotic cell division but indispensable for induction of sexual cycle.(2)Intracellular localization of the ste13 protein :The GST-ste13 fusion proteins were overproduced in E.coli. The affinity-purified fusion protein was used as the antigen to prepare a polyclonal antibody. Western blot analysis revealed that this antibody recognized a 58 kDa polypeptide in S.pombe cell extracts. Immunofluorescence microscopy with anti-ste13 showed that the ste13 protein was not localized in the nucleus and distributed in the cytoplasm.