Developmental Mechanisms of the Chromodomain Gene Chd7

描述（由申请人提供）：这些研究与提高我们对与Chd7缺陷相关的发育缺陷的理解直接相关。基因表达的精确调控对大多数组织和器官的正常发育至关重要。染色质结构域蛋白以DNA结合和atp依赖的解旋酶结构域为特征，在染色质重塑中发挥的作用和对基因表达的表观遗传影响鲜为人知。在人类中，CHD7（色域蛋白第三亚家族的成员）的单倍性不足会导致CHARGE综合征。CHARGE是一种多发性异常疾病，其特征为眼缺损、心脏缺陷、先天性闭锁、生长发育迟缓、生殖器发育不全以及耳聋和前庭功能障碍等耳部异常。我们为一个被捕获的空等位基因（Chd7Gt/+）产生了小鼠杂合子。这些小鼠表现出许多与人类CHARGE表型相似的特征，包括严重的内耳缺陷和出生后生长迟缓。Chd7Gt/+小鼠在Chd7阳性组织中也表达一种¿-半乳糖苷酶报告基因，可用于追踪Chd7缺陷细胞的命运。人类和小鼠中CHARGE的临床特征是可变的和不完全渗透的，CHD7功能丧失的人类和小鼠中最常见的发育畸形是半规管发育不良或发育不全。小鼠伴有Chd7缺乏症的内耳缺损是复杂的，包括半环管畸形和感觉上皮神经支配。特定的组织和细胞类型似乎对Chd7剂量非常敏感，但细胞中Chd7的表达模式和Chd7缺乏在细胞增殖、分化和存活中的机制尚不清楚。此外，CHD7在耳细胞和神经细胞中的相互作用伙伴和下游靶点尚未确定。这些知识将有助于指导内耳缺陷治疗方法的合理发展。我们的总体假设是CHD7在转录复合体中发挥作用，抑制或激活发育中的耳和神经组织中的下游靶基因。我们有三个具体目标：

DESCRIPTION (provided by applicant): These studies have direct relevance for improving our understanding of the developmental defects associated with Chd7 deficiency. Precise regulation of gene expression is critical for normal development of most tissues and organs. Chromodomain proteins, characterized by DNA binding and ATP-dependent helicase domains, have poorly understood roles in chromatin remodeling and exert epigenetic influences on gene expression. In humans, haploinsufficiency for CHD7, a member of the third subfamily of chromodomain proteins, causes CHARGE syndrome. CHARGE is a multiple anomaly disorder characterized by ocular coloboma, heart defects, atresia of the choanae, retarded growth and development, genital hypoplasia, and ear anomalies including deafness and vestibular disorders. We generated mice heterozygous for a gene trapped null allele (Chd7Gt/+). These mice exhibit many features similar to human CHARGE phenotypes, including severe inner ear defects and postnatal growth delays. Chd7Gt/+ mice also express a ¿-galactosidase reporter in Chd7-positive tissues, and can be used for tracking the fates of Chd7 deficient cells. The clinical features of CHARGE in humans and mice are variable and incompletely penetrant, and the most common developmental malformation in both humans and mice with loss of CHD7 function is semicircular canal dysgenesis or hypoplasia. Inner ear defects with Chd7 deficiency in mice are complex, and include malformations of the semicircular canals and innervation of sensory epithelia. Specific tissues and cell types appear uniquely sensitive to Chd7 dosage, but the cellular Chd7 expression patterns and mechanisms of Chd7 deficiency in cell proliferation, differentiation, and survival are not known. Moreover, CHD7 interacting partners and downstream targets in the ear and neural cells are not characterized. Such knowledge would help guide rational development of therapies for inner ear defects. Our global hypothesis is that CHD7 functions in a transcriptional complex to repress or activate downstream target genes in developing ear and neural tissues. We have three specific aims: (1) Characterize cell proliferation, survival, and differentiation in Chd7 mutant mice, (2) Determine the contributions of CHD7-expressing tissues to inner ear development, and (3) Characterize cultured inner ear and neural stem cells from wildtype and Chd7 mutant embryos, and use them to determine whether CHD7 directly binds the promoters of candidate target genes. Results of proposed experiments will bring novel insights into fundamental processes of chromatin remodeling and transcriptional regulation. Improved understanding of tissue-specific requirements for Chd7 might also improve the diagnosis and treatment of CHARGE-related inner ear and other developmental defects. PUBLIC HEALTH RELEVANCE: This project has direct relevance for understanding the mechanisms of CHD7 deficiency-induced hearing and balance disorders in children and adults with CHARGE syndrome. CHARGE syndrome is a multiple anomaly condition that affects 1 in 10,000 newborn children worldwide. Results of experiments described in this proposal will improve our understanding of genetic and epigenetic factors regulating basic chromatin biology, and their impact on development.