Three new collagen VI genes; COL6A4, COL6A5 and COL6A6

三个新的 VI 型胶原蛋白基因；

• 批准号: 8077311
• 负责人: Jamie Fitzgerald
• 金额: $ 29.64万
• 依托单位: OREGON HEALTH & SCIENCE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-07-30 至 2014-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8077311
• 关键词: 3q23; Address; Affect; Anabolism; Basement membrane; Biochemical; C-terminal; Candidate Disease Gene; Cells; Childhood; Chromosomal Breaks; Clinical; Collagen; Collagen Gene; DAG/PE-Binding Domain; Development; Discrimination; Disease; Elements; Extracellular Matrix; Family; Fibroblasts; Frameshift Mutation; Gene Mutation; Genes; Genetic Counseling; Genotype; Goals; Grant; Health; Human; Human Chromosomes; In Vitro; Individual; Knockout Mice; Light; Messenger RNA; Microfibrils; Modeling; Molecular; Mus; Muscle; Muscle Development; Muscular Dystrophies; Mutation; Myopathy; Names; Nonsense Codon; Patients; Phenotype; Proteins; Recombinants; Role; Skeletal Muscle; Staging; Structure; Testing; Tissues; base; congenital muscular dystrophy; extracellular; gene therapy; in vivo; member; molecular assembly/self assembly; mouse development; neonate; novel therapeutic intervention

DESCRIPTION (provided by applicant): Collagen VI is an extracellular matrix component that forms microfibrillar structures in virtually all tissues. To date, three collagen VI chains have been described; a1(VI), a2(VI) and a3(VI) which assemble in a 1:1:1 ratio. We have identified three additional collagen VI genes at a single locus on human chromosome 3q23 and named them COL6A4, COL6A5 and COL6A6 which encode the a4(VI), a5(VI) and a6(VI) chains, respectively. The COL6A4 gene is disrupted in humans and likely to be non-functional. Mutations in COL6A1, COL6A2 and COL6A3, result in two childhood muscle disorders; the relatively mild Bethlem myopathy (BM) and the more severe Ullrich congenital muscular dystrophy (UCMD). We identified a homozygous frameshift mutation in COL6A6 that results in a premature stop codon and is predicted to result in the complete absence of a6(VI) protein. The affected individual died as a neonate of a muscular dystrophy-like disorder that is more severe than BM or UCMD. We hypothesize that the a6(VI) chain is essential for development. Further, we hypothesize that the COL6A6 gene harbors mutations in individuals with severe congential muscular dystrophy distinct from BM and UCMD. The discovery of three additional collagen VI chains indicates that collagen VI assembly and extracellular microfibril composition is more complicated than previously thought. We hypothesize that the three new chains can substitute for a3(VI) to co-assemble with the a1(VI) and a2(VI) chains to form new molecular assemblies of collagen VI. The Specific Aims are: 1) To identify and characterize COL6A6 mutations in patients with congenital muscular dystrophy. Following identification of mutations, the biochemical consequences on collagen VI assembly and extracellular secretion will then be assessed in vitro in patient fibroblasts. 2) To analyze the function of the a6(VI) chain in vivo. We will generate a line of col6a6-null mice to serve as a model for the human mutation we identified. The effect of the lack of a6(VI) chains on mouse development will be examined in biochemical, immunohistochemical and ultrastructural analyses. 3) To understand the mechanism of a4(VI), a5(VI) and a6(VI) assembly into collagen VI. The potential for the new chains to co-assemble with a1 and a2 to form collagen VI heterotrimers will be assessed in vitro. Then, individual domains will be deleted systematically, and the effect on collagen VI biosynthesis, intracellular assembly and secretion determined. Our finding that the loss of a6(VI) protein leads is lethal in humans extends the severe end of the collagen VI disease spectrum and demonstrates that the a6(VI) chain is essential for normal development. This grant will explore the developmental and pathophysiological role of a6(VI) through the analysis of patient cells and of mice deficient for the col6a6 gene. In addition, we will define the mechanism of assembly of the new chain and explore whether they can form heterogeneous collagen VI structures. PUBLIC HEALTH RELEVANCE: We have identified a mutation in a new gene for collagen VI in a patient with a severe muscular dystrophy. This discovery will change the genetic counseling patients and their families with these type of congenital muscular dystrophies receive. In addition, understanding the full spectrum of genes and mutations involved in muscular dystrophy is vital if novel therapeutic approaches, such as gene therapy, are feasible.

描述(申请人提供)：VI型胶原是一种细胞外基质成分，在几乎所有组织中形成微纤维结构。到目前为止，已经描述了三个VI型胶原链：A1(VI)、A2(VI)和A3(VI)，它们以1：1：1的比例组装。我们在人类染色体3q23上的一个位点上发现了另外三个VI型胶原基因，分别命名为COL6A4、COL6A5和COL6A6，它们分别编码A4(VI)、A5(VI)和A6(VI)链。COL6A4基因在人类中被破坏，很可能是无功能的。COL6A1、COL6A2和COL6A3的突变导致两种儿童肌肉疾病：相对轻微的Bethlem肌病(BM)和较严重的Ullrich先天性肌营养不良(UCMD)。我们在COL6A6中发现了一个纯合子移码突变，它导致了一个过早的终止密码子，并被预测导致a6(VI)蛋白的完全缺失。受影响的人死于一种肌肉营养不良样疾病，这种疾病比BM或UCMD更严重。我们假设a6(VI)链对发育是必不可少的。此外，我们假设COL6A6基因在严重的先天性肌营养不良症患者中存在突变，与BM和UCMD不同。另外三条VI型胶原链的发现表明，VI型胶原的组装和细胞外微原纤维的组成比之前认为的要复杂得多。我们假设这三个新的链可以取代A3(VI)与A1(VI)和A2(VI)链共同组装，形成新的VI型胶原分子组合。其具体目的是：1)鉴定和鉴定先天性肌营养不良患者COL6A6突变。在确定突变后，将在体外评估患者成纤维细胞中对VI型胶原组装和细胞外分泌的生化影响。2)分析a6(VI)链在体内的功能。我们将产生一组COL6A6缺失的小鼠，作为我们识别的人类突变的模型。缺乏a6(VI)链对小鼠发育的影响将在生化、免疫组织化学和超微结构分析中进行检验。3)了解A4(VI)、A5(VI)和A6(VI)组装成VI型胶原的机制。体外实验将评估这些新链与A1和A2共同组装形成VI型胶原杂三聚体的可能性。然后，系统地删除单个结构域，并测定其对VI型胶原生物合成、细胞内组装和分泌的影响。我们的发现是，a6(VI)蛋白的丢失对人类是致命的，延长了VI型胶原疾病谱的严重末端，并证明了a6(VI)链对正常发育是必不可少的。这笔赠款将通过对患者细胞和COL6A6基因缺陷小鼠的分析，探索a6(VI)的发育和病理生理学作用。此外，我们还将确定新链的组装机制，并探索它们是否能形成异相的VI型胶原结构。公共卫生相关性：我们在一名患有严重肌营养不良症的患者中发现了一种新的VI型胶原基因突变。这一发现将改变患有这些类型先天性肌营养不良的患者及其家人接受的遗传咨询。此外，如果基因治疗等新的治疗方法可行，了解肌营养不良症所涉及的基因和突变的全谱是至关重要的。