Studies on the mechanisms for nucleocytoplasmic transport and localization of mRNAs using the temperature sensitive mutants in the fisson yeast

mRNA的核质转运是真核生物基因表达的重要环节之一。然而，mRNA转运的机制大多是未知的。为了阐明mRNA输出的分子机制，我们用与mRNA的poly A尾杂交的oligo dT探针进行原位杂交，筛选了裂殖酵母Schizosacccharomyces pombe中对生长温度敏感的突变体库，并鉴定了7个在非允许温度下在核中积累mRNA的突变体。这些突变都是隐性的，分为七个不同的组，命名为ptr 1 -7（poly（A）^+ RNA转运）。通过使用粟酒裂殖酵母基因组文库的功能互补，我们已经克隆了与每个突变互补的基因。对克隆基因的分析表明，ptr 1 ^+、ptr 2 ^+、ptr 3 ^+、ptr 4 ^+和ptr 6 ^+分别编码泛素连接酶E3样蛋白、RNA/TC 4的鸟嘌呤核苷酸交换因子、泛素激活酶E3、染色体分离所需的Cut 1 p和人TAFII 55的同源物。发现ptr 5 ^+和ptr 7 ^+在粟酒裂殖酵母中编码新蛋白。这些结果表明，泛素化系统在mRNA转运途径中起着重要作用，并且mRNA转运与其他细胞事件如基因转录和细胞周期进程之间可能存在紧密联系

Nucleocytoplasmic transport of mRNA is one of the essential steps in the eukaryotic gene expression. However, a mechanism of mRNA transport is mostly unknown. To elucidate a molecular mechanism for mRNA export, we have screend a bank of mutants temperature sensitive for growth in fission yeast Schizosacccharomyces pombe by using in situ hybridization with an oligo dT probe that hybridizes to the poly A tail of mRNAs, and identified seven mutants that accumulate mRNA in the nuclei at the nonpermissive temperature. Those mutations are all recessive and fall into seven different groups designated as ptr1-7 (poly(A)^+ RNA tranport). By functional complementation using an S.pombe genomic library, we have cloned the genes complementing each mutation. Analyzes of the cloned genes revealed that ptr1^+, ptr2^+, ptr3^+, ptr4^+ and ptr6^+ encode the ubiquitin ligase E3-like protein, the guanine nucleotide exchange factor for Rna/TC4, ubiquitin activating enzyme E3, Cut1p required for the chromosome segregation, and the homologue for human TAFII55, respectively. ptr5^+ and ptr7^+ were found to encode novel proteins in S.pombe. These results suggest that the ubiquitination system plays an important role in the mRNA transport pathway, and that there might be a tight linkage between the mRNA transport and the other cellular events, such as the gene transcription and cell cycle progression