Mistargeting of Elastase in Bone Marrow Failure

骨髓衰竭中弹性蛋白酶的误定位

• 批准号: 7105072
• 负责人: MARSHALL S. HORWITZ
• 金额: $ 36.62万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-09-30 至 2009-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7105072
• 关键词: African American; antibody; catalyst; clinical research; cofactor; congenital blood disorder; elastases; enzyme substrate; family genetics; gene mutation; genetic promoter element; human genetic material tag; human subject; immunologic substance development /preparation; intracellular transport; laboratory mouse; molecular pathology; neutropenia; protein localization; protein transport; site directed mutagenesis; transport proteins

DESCRIPTION (provided by applicant): Neutrophils are phagocytic white blood cells, and a deficiency of their numbers ("neutropenia") predisposes to bacterial and fungal infection. The two principal inherited forms of human neutropenic bone marrow failure are cyclic neutropenia (CN) and leukemia-predisposing, severe congenital neutropenia (SCN). "Benign ethnic neutropenia" is common to individuals of African descent and may also have health consequences. Mutations of the gene ELA2, encoding neutrophil elastase (NE), cause CN and are the most frequent cause of SCN. Recent findings that canine CN is caused by mutations of the AP3 transporter and that some SCN may result from ELA2 promoter variants or mutations of the transcriptional repressor Gfi1, both of which lead to over-expression of NE, suggest a hypothesis for neutropenia in these and other bone marrow failure syndromes: NE is a transmembrane "cargo" protein for AP3. Mutation of NE's transmembrane domains causes too much NE to accumulate in granules, resulting in CN. Mutation of NE's AP3-recognition signal, or of AP3 itself, mislocalizes NE to the plasma membrane. Similarly, over-expression of NE consequent to ELA2 promoter variants or Gfi1 mutations overwhelms normal AP3-mediated trafficking pathways and diverts NE to the plasma membrane. Three Specific Aims are directed toward testing this hypothesis, determining if a common ELA2 promoter variant in the African-American population associates with benign ethnic neutropenia, and identifying additional neutropenia genes: 1 Characterize NE's membrane relationship: 1.1 Develop antibodies to predicted cytoplasmic and luminal surfaces of NE; 1.2 Test membrane-bound NE for altered catalysis on model and biological substrates; 1.3 Examine the evolutionary conservation of AP3 interactions in other species and for other proteases. 2 Evaluate mistrafficking of NE as a general mechanism for neutropenia: 2.1 Identify potential substrates and cofactors of mislocalized NE; 2.2 Inspect NE trafficking in other neutropenic disorders. 3 Discover other mutations that may cause neutropenia through over-expression of NE: 3.1 Determine if ELA2 promoter variation contributes to neutropenia; 3.2 Measure the frequency of the ELA2 C-199A allele in individuals of African descent with benign ethnic neutropenia; 3.3 Identify new neutropenia genes.

描述（由申请人提供）： 中性粒细胞是吞噬白细胞，其数量不足（“中性粒细胞减少”）易于细菌和真菌感染。人类中性粒细胞减少性骨髓衰竭的两种主要遗传形式是环状中性粒细胞减少（CN）和白血病 - 可抑制，严重的先天性中性粒细胞减少症（SCN）。 “良性族裔中性粒细胞减少症”是非洲血统个体共同的，也可能会带来健康后果。编码中性粒细胞弹性蛋白酶（NE）的基因ELA2的突变引起CN，是SCN的最常见原因。犬CN犬CN的最新发现是由AP3转运蛋白的突变引起的，并且某些SCN可能是由ELA2启动子变体或转录抑制剂GFI1的突变引起的，这两者都会导致NE的过表达，这表明在这些和其他骨髓失败综合症中中性粒细胞增多症的假设：Ne ne a apbrane opbrane opbrane“ cargo of cargo” NE跨膜结构域的突变导致NE过多无法在颗粒中积聚，从而导致CN。 NE的AP3识别信号或AP3本身的突变使NE误入为质膜。同样，对ELA2启动子变体或GFI1突变导致NE的过表达压倒了正常AP3介导的运输途径，并将NE转移到质膜上。三个具体的目的是针对检验这一假设的，确定非裔美国人人口与良性种族中性粒细胞减少症的共同ELA2启动子的变体，并确定了其他中性粒细胞基因：1表征了NE的膜关系：1.1从NE NE NE NE NE NE NE NE NE NE表面进行预测的细胞胞质和脉络膜表面的抗体； 1.2测试膜结合的NE，用于改变模型和生物底物的催化作用； 1.3检查其他物种和其他蛋白酶中AP3相互作用的进化保护。 2评估NE作为中性粒细胞减少症的一般机制的错误贸易：2.1识别NE的潜在底物和辅助因子； 2.2检查其他中性粒细胞减少疾病的NE贩运。 3发现其他可能导致中性粒细胞减少症的突变：3.1确定ELA2启动子变异是否有助于中性粒细胞减少。 3.2测量具有良性族裔中性粒细胞减少良性的非洲血统中ELA2 C-199A等位基因的频率； 3.3鉴定新的中性粒细胞减少基因。