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AG-NOR PROTEINS USED FOR CANCER DIAGNOSIS

AG-NOR 蛋白用于癌症诊断

基本信息

批准号：
2112291
负责人：
BENIGNO C VALDEZ
金额：
$ 10万
依托单位：
BAYLOR COLLEGE OF MEDICINE
依托单位国家：
美国
项目类别：
财政年份：
1995
资助国家：
美国
起止时间：
1995-09-30 至 1998-09-29
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/2112291
关键词：
chromosomes neoplasm /cancer diagnosis nucleoproteins silver impregnation

项目摘要

This is a Shannon award providing partial support for the research projects that fall short of the assigned Institute's funding range but are in the margin of excellence. The Shannon award is intended to provide support to test the feasibility of the approach; develop further tests and refine research techniques; perform secondary analysis of available data sets; or conduct discrete projects that can demonstrate the PI's research capabilities or lend additional weight to an already meritorious application. The abstract below is taken from the original document submitted by the principal investigator. DESCRIPTION: Ag-NOR proteins are silver-stainable proteins located in the nucleolar organizer region (NOR) of a eukaryotic cell. These proteins are used for cancer diagnosis to distinguish malignant from benign tumors: malignant lesions contain more Ag-NOR proteins than benign lesions. Identification and characterization of the Ag-NOR proteins have both biological and clinical significance. They will be useful in understanding the mechanisms of two important cellular processes: nucleolar formation which begins at the NOR during cell division, and transcription of ribosomal RNA. The investigators will also provide useful information for improved cancer diagnosis. Only nucleolin/C23 and nucleophosmin/B23 are known to belong to this group of proteins. However, their specific activities in terms of silver staining are yet to be compared to the specific activities of other unidentified Ag-NOR proteins. The long-term objectives of this proposed project are to identify other Ag-NOR proteins, to characterize them and to determine the mechanism through which they are silver stained. To accomplish these goals, a human cDNA library will be screened by Ag-NOR staining protocol which identifies silver stainable proteins on nitrocellulose filters and by hybridization technique using a P32-labeled DNA probe which codes for the second acidic domain of nucleophosmin/B23 shown to be responsible for the nucleophosmin/B23 silver staining property. Isolated cDNA clones corresponding to the silver staining proteins will be sequenced and expressed in bacteria. Localization in HeLa cells will be done by immunostaining using antibodies raised against these proteins. Known nucleolar proteins with acidic domains such as UBF1, HMG1, and HMG2 will be tested for their ability to be silver stained. The elucidation of the mechanism of Ag-NOR staining will be based on their previous finding that aspartate residues are responsible for silver staining of nucleophosmin/B23. Mutations will be introduced to find if the same mechanism holds true for other Ag-NOR poteins and to determine the minimum number of aspartic acid residues required for silver staining. Nucleolar proteins that bind to Ag-NOR proteins will also be identified. The high proliferative activity of cancer cells may be related to their high level of ribosomal RNA expression. Since Ag-NOR proteins are involved in ribosomal RNA synthesis, the results form this study may explain in detail the relationship between Ag-NOR proteins and malignancy. This project may also lead to the development of better cancer diagnostic tools like using antibodies against Ag-NOR proteins, and in designing drugs that may inhibit cancer cell growth by disrupting the interaction of Ag-NOR proteins wiht other nucleolar proteins.

这是一个香农奖提供部分支持的研究项目不属于指定的研究所的资金范围，但都处于优秀的边缘。香农奖旨在提供支持测试方法的可行性;开发进一步的测试并完善研究技术;对现有的数据集;或进行离散的项目，可以证明PI的研究能力或增加已经有价值的应用程序.以下摘要摘自原始文件由主要研究者提交。描述：Ag-NOR蛋白是位于真核细胞的核仁组成区。这些蛋白质用于癌症诊断以区分恶性肿瘤和恶性肿瘤。良性肿瘤：恶性病变含有更多的Ag-NOR蛋白，良性病变Ag-NOR的鉴定和表征蛋白质具有生物学和临床意义。他们将有助于理解两种重要的细胞机制，过程：核仁的形成，在细胞分裂过程中开始于NOR 分裂和核糖体RNA的转录。调查人员还将为改善癌症诊断提供有用的信息。只已知核仁素/C23和核磷蛋白/B23属于这组 proteins.然而，就银染色而言，它们的特异性活性还有待与其他身份不明的 Ag-NOR蛋白。这项拟议计划的长远目标是鉴定其他Ag-NOR蛋白，表征它们，它们被银染色的机制。完成这些目的：通过Ag-NOR染色筛选人cDNA文库其识别硝酸纤维素过滤器上的银蛋白，通过使用P32标记的DNA探针的杂交技术，核磷蛋白/B23的第二个酸性结构域被证明是负责用于核磷蛋白/B23银染色性质。分离的cDNA克隆对应于银染色蛋白质的蛋白质将被测序，在细菌中表达。在HeLa细胞中的定位将通过使用针对这些蛋白质产生的抗体进行免疫染色。已知具有酸性结构域的核仁蛋白如UBF 1、HMG 1和HMG 2将测试其被银染色的能力。的说明， Ag-NOR染色的机制将基于他们先前的发现，天冬氨酸残基负责银染色，核磷蛋白/B23。将引入突变以确定是否相同机制适用于其他Ag-NOR poteins，并确定银染色所需的天冬氨酸残基的最小数量。还将鉴定与Ag-NOR蛋白结合的核仁蛋白。癌细胞的高增殖活性可能与其高水平核糖体RNA表达。由于Ag-NOR蛋白是参与核糖体RNA合成，本研究的结果可能详细解释Ag-NOR蛋白与恶性肿瘤这个项目也可能导致更好的发展癌症诊断工具，如使用针对Ag-NOR蛋白的抗体，在设计药物时， Ag-NOR蛋白与其它核仁蛋白的相互作用。