Structural and Functional Characterization of BRCA1/BARD1

BRCA1/BARD1 的结构和功能表征

• 批准号: 7881691
• 负责人: Rachel E Klevit
• 金额: $ 37.44万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-12-21 至 2013-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7881691
• 关键词: Affinity; Ankyrin Repeat; BARD1 gene; BRCA1 gene; Binding; Binding Proteins; Biochemical; Biological; Biological Assay; Breast; C-terminal; Cell physiology; Cells; Chromosome abnormality; Complex; Crystallography; DNA Damage; Enzymes; Estrogen Receptors; Estrogens; Event; Functional disorder; Future; Genes; Genome Stability; Goals; Grant; Health; Human; Human Ubiquitin; In Vitro; Inherited; Investigation; Knock-in Mouse; Knock-out; Lead; Learning; Ligase; Link; Maintenance; Malignant Neoplasms; Malignant neoplasm of ovary; Mass Spectrum Analysis; Methods; Molecular; Mono-S; Multienzyme Complexes; Mus; Mutation; N-terminal; Nature; Outcome; Play; Polyubiquitin; Progesterone Receptors; Protein Binding; Proteins; Regulation; Resolution; Role; Substrate Interaction; System; TP53 gene; Techniques; Testing; Tumor Suppressor Proteins; Ubiquitin; Ubiquitin-Conjugating Enzymes; Ubiquitination; Woman; Work; Yeasts; in vivo; insight; lifetime risk; malignant breast neoplasm; mutant; protein complex; public health relevance; research study; response; three dimensional structure; tool; ubiquitin ligase; ubiquitin-protein ligase; yeast two hybrid system

DESCRIPTION (provided by applicant): The breast and ovarian cancer tumor suppressor protein, BRCA1, and its obligate protein partner, BARD1 are critical to proper functioning of fundamental cellular processes that support genomic stability. To date, a single biochemical activity has been identified for BRCA1/BARD1, namely, they function together as a ubiquitin E3 ligase. Although it is believed that the ability of BRCA1/BARD1 to modify specific cellular proteins with ubiquitin is fundamental to its role as a tumor suppressor, the details and ramifications of this relationship remain to be elucidated. During the past grant period, we discovered that BRCA1/BARD1 can function with ten human ubiquitin-conjugating enzymes (E2s) and that the ultimate product generated by BRCA1/BARD1 on substrates depends on the E2 present. We also identified several new substrates, including the estrogen receptor. In the next grant period we will expand our focus to include the essential subunit, BARD1, more explicitly and will participate in a collaborative effort to develop new molecular insights and tools for another RING E3 ligase that is critical in cancers, Mdm2/MdmX. The overall goals for the next grant period are 1) elucidate structural and functional determinants of mono-ubiquitin transfer and poly-ubiquitin chain formation by BRCA1/BARD1 and its interacting E2s, 2) investigate the molecular and structural determinants of BRCA1/BARD1-substrate interactions, 3) characterize BARD1 and its interactions, and 4) identify the human E2s that interact with Mdm2/MdmX, the ligase responsible for p53 regulation in vivo. A broad experimental approach will be undertaken, including biochemical, structural, molecular biological, and cellular techniques. Results from these studies will provide new insights into BRCA1/BARD1 function and will contribute to the general understanding of protein ubiquitination. PUBLIC HEALTH RELEVANCE: The breast and ovarian cancer tumor suppressor protein, BRCA1, plays a role in the maintenance of genomic stability and its loss or dysfunction leads to widespread chromosomal abnormalities. Inheritance of a mutant form of BRCA1 increases a woman's lifetime risk of developing breast cancer from 1 in 8 to greater than 1 in 2. Some of the most common inherited mutations abrogate BRCA1's function as a ubiquitin ligase, implying that this function is central to the health of a cell. A full description of the molecular interactions that are critical to BRCA1 function will provide new insight into the early events associated with loss of BRCA1 that lead to tumorogenesis.

描述（由申请人提供）：乳腺癌和卵巢癌肿瘤抑制蛋白 BRCA1 及其专性蛋白伴侣 BARD1 对于支持基因组稳定性的基本细胞过程的正常运行至关重要。迄今为止，已鉴定出 BRCA1/BARD1 的单一生化活性，即它们作为泛素 E3 连接酶共同发挥作用。尽管人们认为 BRCA1/BARD1 用泛素修饰特定细胞蛋白的能力对其作为肿瘤抑制因子的作用至关重要，但这种关系的细节和后果仍有待阐明。在过去的资助期内，我们发现 BRCA1/BARD1 可以与十种人类泛素结合酶 (E2) 一起发挥作用，并且 BRCA1/BARD1 在底物上产生的最终产物取决于 E2 的存在。我们还鉴定了几种新的底物，包括雌激素受体。在下一个资助期内，我们将扩大我们的重点，更明确地包括基本亚基 BARD1，并将参与合作，为另一种在癌症中至关重要的 RING E3 连接酶 Mdm2/MdmX 开发新的分子见解和工具。下一个资助期的总体目标是 1) 阐明 BRCA1/BARD1 及其相互作用的 E2 形成单泛素转移和多泛素链的结构和功能决定因素，2) 研究 BRCA1/BARD1 底物的分子和结构决定因素相互作用，3) 表征 BARD1 及其相互作用，以及 4) 识别与连接酶 Mdm2/MdmX 相互作用的人类 E2负责体内p53的调节。将采用广泛的实验方法，包括生物化学、结构、分子生物学和细胞技术。这些研究的结果将为 BRCA1/BARD1 功能提供新的见解，并将有助于对蛋白质泛素化的一般理解。公共健康相关性：乳腺癌和卵巢癌肿瘤抑制蛋白 BRCA1 在维持基因组稳定性方面发挥着重要作用，其缺失或功能障碍会导致广泛的染色体异常。 BRCA1 突变形式的遗传会使女性终生患乳腺癌的风险从八分之一增加到二分之一以上。一些最常见的遗传突变会废除 BRCA1 作为泛素连接酶的功能，这意味着该功能对于乳腺癌至关重要。细胞的健康。对 BRCA1 功能至关重要的分子相互作用的完整描述将为了解导致肿瘤发生的 BRCA1 缺失相关的早期事件提供新的见解。