MOLECULAR BIOLOGY OF MITOSIS IN ASPERGILLUS NIDULANS

有丝分裂及其调控对于理解 细胞生物学、癌症和人类发育。形态上的和 有丝分裂的生化特征在真核生物中高度保守 有机体。为此，使用低级真核生物，例如酵母和 真菌对有丝分裂的遗传学研究取得了很高的成果。在这里我们 建议对曲霉的bimE基因及其基因产物进行研究 在以前的实验中已经被证明是一个重要的负面影响 有丝分裂的调节者，似乎是调节系统的一部分， 如果DNA合成尚未完成，则阻止有丝分裂。我们将生成 并使用针对bimE/E.coliTrype或LacZ融合蛋白的抗体来 BimE基因产物在细胞周期中的同步化 细胞，寻找其合成、生化结构的变化，后 翻译修饰、本地化及其与他人的互动 蛋白质。我们还建议表征具有bimE的蛋白质 间接抑制子分析和解链分析可能与蛋白质相互作用 非互补突变。我们将尝试了解bime如何监管 通过检测bimE对转录的影响来检测nimA基因的表达水平 和/或NIMA基因的稳定性。最后，与约翰博士合作 加州大学圣地亚哥分校的Newport，我们将在 非洲爪哇研究bimE基因产物对小鼠有丝分裂的影响 非洲爪哇卵的细胞质无细胞系统。

Mitosis and its regulation are of fundamental importance to understanding cell biology, cancer, and human development. The morphological and biochemical features of mitosis are highly conserved among eukaryotic organisms. For this reason the use of lower eukaryotes, e.g. yeasts and fungi to study the genetics of mitosis has been highly productive. Here we propose to study the bimE gene of Aspergillus and its gene product, which has been shown in previous experiments to be an important negative regulator of mitosis and appears to be part of a regulatory system that prevents mitosis if DNA synthesis has not been completed. We will generate and use antibodies to bimE/E. coli trypE or lacZ fusion proteins to characterize the bimE gene product through the cell cycle in synchronized cells, looking for changes in its synthesis, biochemical structure, post- translational modification, localization and interactions with other proteins. We also propose to characterize proteins with which the bimE protein might interact by analysis of indirect suppressor and unlinked noncomplementing mutations. We will attempt to learn how bimE regulates the level of nimA mRNA by determining the effect of bimE on transcription and/or stability of nimA mRNA. Finally, in collaboration with Dr. John Newport of UC-San Diego we will look for a functional homolog of bimE in Xenopus laevis nad study the effect of the bimE gene product on mitosis in the Xenopus egg cytoplasm cell-free system.