Structural and Functional Characterization of BRCA1/BARD1

BRCA1/BARD1 的结构和功能表征

• 批准号: 7931268
• 负责人: Rachel E Klevit
• 金额: $ 19.83万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-30 至 2011-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7931268
• 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）：乳腺癌和卵巢癌抑制蛋白BRCA1及其强制性蛋白质伴侣Bard1对于支持基本稳定性的基本细胞过程至关重要。迄今为止，已经确定了用于BRCA1/BARD1的单一生化活性，即它们作为泛素E3连接酶一起起作用。尽管认为BRCA1/BARD1用泛素修饰特定的细胞蛋白的能力对于其作为肿瘤抑制剂的作用至关重要，但这种关系的细节和后果仍然待阐明。在过去的赠款期间，我们发现BRCA1/BARD1可以与十种人的泛素偶联酶（E2S）一起起作用，并且BRCA1/BARD1在底物上产生的最终产物取决于E2。我们还确定了包括雌激素受体在内的几个新底物。在下一个赠款期间，我们将更加明确地将重点扩展到包括基本亚基BARD1，并将参与协作的努力，以开发新的分子见解和工具，以针对MDM2/MDMX至关重要的另一个环E3连接酶。下一个赠款期的总体目标是1）BRCA1/BARD1及其相互作用的E2S阐明单泛素转移和多泛素链形成的结构和功能决定因素，2）研究BRCA1/BARD1-SUBSTRATE ITOCTOTION的分子和结构确定性，并识别与人类相互作用的BRCA1/BARD1-SUBSTRATE相互作用，以及4） MDM2/MDMX，负责体内p53调节的连接酶。将采用广泛的实验方法，包括生化，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，并有助于对蛋白质泛素化的一般理解。公共卫生相关性：乳腺癌和卵巢癌抑制蛋白BRCA1在维持基因组稳定性及其丧失或功能障碍的过程中起作用，导致广泛的染色体异常。 BRCA1突变形式的遗传会使女性患乳腺癌的终生风险从8中的1增加到大于2中的1。一些最常见的遗传突变消除了BRCA1作为泛素连接酶的功能，这意味着该功能是细胞健康的核心。对BRCA1功能至关重要的分子相互作用的完整描述将为与BRCA1丢失相关的早期事件提供新的见解，从而导致肿瘤发生。