Nuclear Transport Factors in Chromosomal Instability

描述(申请人提供)：大多数人类癌症的特征是染色体数目改变。引发或促进整个染色体的丢失或获得的分子缺陷在很大程度上仍不清楚。在一些染色体不稳定的癌症中，已经发现编码有丝分裂检查点蛋白的基因突变，如Bub1、BubR1和MAD2。这些蛋白质是一个复杂的分子机器的一部分，该机器阻止后期的开始，直到所有的染色体都正确地连接到有丝分裂纺锤体上并在中期平板上排列。在有丝分裂时核膜解体后，核质运输因子Ran和Importinα/β成为有丝分裂纺锤体组装的关键调节因子。Nup98是另一种核运输因子，经常成为急性白血病基因改变的目标。Nup98包含一个被称为GLEBS的基序，它直接与假定的mRNA输出因子RAE1结合。我们最近发现在有丝分裂检查点蛋白Bub1和BubR1中存在一个GLEBS基序，它是RAe1相关的有丝分裂检查点蛋白Bub3的结合位点。采用基因敲除的方法，我们已经确定Nup98和Rae1不仅与Bub蛋白有序列同源性，而且是有丝分裂检查点功能所必需的。这项提案的总体目标是确定Nup98及其结合伙伴RAE1在维持染色体稳定和肿瘤形成中的作用。在第一个目标中，我们将确定Nup98调节有丝分裂检查点功能和染色体稳定性的机制。在第二个目标中，我们将通过对小鼠体内Nup98的组织特异性破坏来研究Nup98在正常和肿瘤生长中的作用。在第三个目标中，我们将重点研究RAE1在有丝分裂检查点功能、维持染色体稳定和肿瘤形成中的作用。从这些研究中获得的信息将为理解核运输因素导致遗传不稳定和癌症的机制提供一个框架。

DESCRIPTION (provided by applicant): Most human cancers are characterized by chromosome number alterations. Molecular defects that initiate or promote losses or gains of whole chromosomes remain largely unknown. In some cancers with chromosomal instability, mutations have been identified in genes that encode for mitotic checkpoint proteins, such as Bub1 BubR1 and Mad2. These proteins are parts of an intricate molecular machine that blocks the onset of anaphase until all chromosomes are properly attached to the mitotic spindle apparatus and aligned at the metaphase plate. Following disassembly of the nuclear envelope at mitosis, the nucleocytoplasmic transport factors Ran and importin alpha/beta become key regulators of mitotic spindle assembly. Nup98, another nuclear transport factor, is a frequent target of genetic alterations in acute leukemias. Nup98 contains a motif, termed GLEBS, that directs binding to the putative mRNA export factor Rae1. We have recently shown that a GLEBS motif is present in the mitotic checkpoint proteins Bub1 and BubR1, where it serves as a binding site for the Rae1-related mitotic checkpoint protein Bub3. Using a gene-knockout approach, we have determined that Nup98 and Rae1 not only share sequence homology with Bub proteins, but are required for mitotic checkpoint function as well. The overall goal of this proposal is to define the role of Nup98 and its binding partner Rae1 in the maintenance of chromosomal stability and tumor formation. In the first aim, we will determine the mechanism by which Nup98 regulates mitotic checkpoint function and chromosomal stability. In the second aim, we will examine the role of Nup98 in normal and neoplastic growth by tissue-specific disruption of Nup98 in mice. In the third aim, we will focus on the role of Rae1 in mitotic checkpoint function, maintenance of chromosomal stability, and tumor formation. Information gained from these studies will provide a framework for understanding the mechanisms by which nuclear transport factors contribute to genetic instability and cancer.