Molecular Basis of Aneuploidy

描述(申请人提供)：这项建议集中于一个重要但尚未探索的问题-类固醇激素，这是众所周知的危险因素，如何与p53突变相互作用，产生非整倍体和恶性肿瘤，以及如何参与染色体分离蛋白分离酶？我们的性激素依赖的p53-小鼠乳腺癌前病变模型允许一个独特的方法来解决这个问题。在这个模型中，在p53突变的乳腺中，类固醇诱导导致染色体不稳定、非整倍体和肿瘤形成，类似于大多数人类乳腺癌。我们提出了一种范式，即存在一组蛋白质，其解除调控促进了非整倍体(称为PR AN；非整倍体的促进者)，包括导致整个或部分染色体丢失或获得的染色体不稳定，并且PRAN蛋白受到类固醇激素和肿瘤抑制基因P53的交互调节。我们发表的和新的初步数据提供了第一个证据，即类固醇激素在参与姐妹染色单体聚集和分离的有丝分裂蛋白的调节中发挥作用。我们认为，抑癌基因P53的突变和类固醇激素的信号传递共同作用，通过影响参与染色体分离的关键蛋白的表达，在乳腺癌中产生非整倍体。我们集中在调节染色体分离的元件，特别是姐妹染色单体凝聚/分离蛋白，作为候选的PRAN蛋白，因为有丝分裂过程中的染色体错误分离可能导致非整倍体。在这一分析中的一个关键基因是ESPL1，它编码一种名为分离酶的内肽酶，在中期到后期的转变中通过裂解粘连蛋白Rad21/SCC1/MCD1来分离连接的姐妹染色单体。假设是荷尔蒙刺激p53缺失的小鼠乳腺导致ESPL1基因的错误表达，从而促进非整倍体和乳腺癌的形成。这一建议适用于体内移植稳定转ESPL1的p53突变和野生型(WT)乳腺细胞，以及ESPL1转基因小鼠模型，以测试激素治疗后的PRAN范例。通过对ESPL1启动子区域的研究，在转录水平上研究了类固醇和P53对ESPL1的调控。这些目标将通过追求两个特定的目标来实现：1)分离酶在非整倍体中的过度表达的功能作用；2)ESPL1基因表达的转录调控。这项拟议的研究不仅阐明了激素诱导非整倍体的潜在机制，这是癌症生物学中一个基本的悬而未决的问题，而且可能识别出一类新的蛋白质，这些蛋白质负责染色体的不稳定和乳腺癌的进展。

DESCRIPTION (provided by applicant): This proposal focuses on an important but unexplored problem - how steroid hormones, which are well known risk factors, interact with p53 mutations to produce aneuploidy and malignancy, and how the chromosomal segregation protein Separase is involved? Our sex-steroid dependent p53-mice preneoplastic breast cancer model allows a unique approach to this problem. In this model, steroidal induction in p53 mutant mammary glands results in chromosomal instability, aneuploidy and tumor formation analogous to that seen in majority of human breast cancers. We propose a paradigm that there is a set of proteins whose deregulation promotes aneuploidy (termed PR AN; Promoter of Aneuploidy) including chromosomal instability which results in loss or gain of whole or parts of chromosomes, and that PRAN proteins are interactively regulated by steroid hormones and the tumor suppressor p53. Our published and new preliminary data provide the first evidence that steroid hormones play a role in the regulation of mitotic proteins involved in sister chromatid cohesion and separation. We propose that the combined effect of mutation of the tumor suppressor p53 and signaling by steroid hormones produces aneuploidy in breast cancer by affecting expression of key proteins involved in chromosomal separation. We focuses on the elements that regulate chromosomal segregation, particularly sister chromatid cohesion/separation proteins, as candidate PRAN proteins, since chromosome missegregation during mitosis can lead to aneuploidy. A key gene in this analysis is ESPL1, which encodes an endopeptidase called Separase that separates joined sister chromatids by cleaving cohesin Rad21/SCC1/MCD1 during the metaphase to anaphase transition. The hypothesis is that hormonal stimulation of p53 null mouse mammary glands results in misexpression of the ESPL1 gene, thus promoting aneuploidy and breast cancer formation. This proposal applies in vivo transplantation of p53 mutant and wild type (WT) mammary cells that are stably transfected with ESPL1, and an ESPL1 transgenic mice model to test the PRAN paradigm following hormone treatment. Steroid and p53 regulation of ESPL1 at the transcriptional level is studied by characterizing the ESPL1 promoter region. These objectives will be accomplished by pursuing two specific aims: 1) Functional role of Separase overexpression in aneuploidy, and 2) Transcriptional regulation of ESPL1 gene expression. The proposed study not only elucidate underlying mechanisms of hormone-induced aneuploidy, a fundamental unresolved question in cancer biology, but also likely to identify a new class of proteins that are responsible for chromosomal instability and breast cancer progression.