Novel targets of CRL4 ligase within Cohesinopathy pathways

CRL4 连接酶在粘连病途径中的新靶标

• 批准号: 10699961
• 负责人: ROBERT SKIBBENS
• 金额: $ 7.86万
• 依托单位: LEHIGH UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-13 至 2024-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10699961
• 关键词: Affect; Bruck-de Lange syndrome; Caring; Cells; Child; Cleft Palate; Complex; Congenital Abnormality; Craniofacial Abnormalities; Cullin Proteins; Data; Defect; Development; Disease; Embryo; Emotional; Ensure; Exhibits; Exposure to; Eye; Family; Financial Hardship; Frequencies; Future; Gastrointestinal tract structure; Gene Mutation; Genes; Genetic; Genetic Transcription; Goals; Growth; Healthcare Systems; Heart Abnormalities; Infant; Infant Mortality; Injections; Intellectual functioning disability; Intervention; Ligase; Limb structure; Messenger RNA; Microcephaly; Modeling; Mutate; Mutation; Organ; Pathway interactions; Patients; Pharmacological Treatment; Phenotype; Phocomelia; Proteins; Regulation; Respiratory System; Roberts-SC phocomelia syndrome; Role; Set protein; Severities; Syndrome; Teratogens; Testing; Thalidomide; Transcriptional Regulation; Ubiquitination; Zebrafish; cohesin; disease phenotype; family burden; genome integrity; hearing impairment; infant death; innovation; insight; knock-down; liquid chromatography mass spectrometry; novel; novel strategies; overexpression; pharmacologic; programs; protein expression; reduce symptoms; ubiquitin ligase

Every year, approximately ~120,000 infants are born with birth defects that require special care and interventions to survive. In fact, birth defects are the leading cause of infant death. Thus, developmental maladies represent a significant emotional and financial burden on both families and the health care system. Transcription dysregulation, or gene mutations affecting developmental programs, account for the majority of birth defects. While numerous gene mutations that underlie developmental maladies have been identified, this often does not provide insight into treatments or symptom amelioration through pharmacological approaches. Here, we leverage our expertise on two related genetic syndromes, Roberts Syndrome (RBS) and Cornelia de Lange Syndrome (CdLS). RBS and CdLS patients exhibit a range of severe phenotypes that include craniofacial defects (microcephaly, eye defects, hearing loss), reduced limb size (phocomelia), abnormalities of the heart, GI and respiratory tracts, and intellectual disabilities. The genetic basis of RBS and CdLS are known - both arise through mutation of a cohesin-based pathway that regulates gene transcription and ensures genome integrity. We recently discovered that the cohesin pathway regulates the transcription of ddb1 - encoding a key component of the Cullin4 Ring Ligase (CRL4) ubiquitination complex. We hypothesize that RBS (esco2 mutated) and CdLS (smc3 mutated), and likely other developmental maladies, arise in large part through reduced CRL4 activity. In support of this hypothesis, exogenous expression of ddb1 reduces the severity of developmental defects that otherwise arise in smc3 knockdown zebrafish embryos. We performed liquid chromatography–mass spectrometry on embryos knocked down for esco2 (RBS), smc3 (CdLS) and ddb1. We obtained a prioritized list of candidates, common across all treatments, that we predict are downstream of CRL4 activity and involved in RBS/CdLS phenotypes. In Specific Aim 1 of this proposal, we validate the LC-MS data and further test the extent to which knockdown of these targets impact RBS/CdLS-type developmental defects in zebrafish embryos simultaneously reduced in either esco2 or smc3 expression. In Specific Aim 2, we test our hypothesis that exogenous expression of these candidates is teratogenic. In combination, these studies will reveal new targets through which birth defect severity can be reduced.

每年，大约约120,000名婴儿出生有先天缺陷 需要特殊护理和干预措施才能生存。实际上，先天缺陷是领先的 婴儿死亡的原因。那就是发展性疾病代表着一种重要的情感 以及家庭和医疗保健系统的财务燃烧。转录 失调或影响发展计划的基因突变，解释了 大部分出生缺陷。而众多基因突变是发展的基础 已经确定了疾病，这通常无法提供有关治疗或 症状通过药物方法进行amélioration。 在这里，我们利用有关两个相关遗传综合症的专业知识，罗伯茨 综合征（RB）和Cornelia de Lange综合征（CDLS）。 RB和CDLS患者 展示了包括颅面缺陷的一系列严重表型（小头畸形， 眼睛缺陷，听力损失），肢体大小减小（凤梨素），心脏异常， 胃肠道和呼吸道，以及肠道残疾。 RB和 CDL是已知的 - 两者都是通过基于粘着蛋白的途径的突变而产生的 调节基因转录并确保基因组完整性。我们最近发现 粘蛋白途径调节DDB1的转录 - 编码一个关键组件 Cullin4环连接酶（CRL4）泛素化复合物。我们假设苏格兰皇家银行 （ESCO2突变）和CDL（SMC3突变），以及其他可能的发育疾病， CRL4活性在很大程度上出现。为了支持这一假设， DDB1的外源表达降低了发育缺陷的严重程度 否则在SMC3敲低斑马鱼胚胎中出现。 我们在爆可的胚胎上进行了液相色谱 - 质谱法 向ESCO2（RB），SMC3（CDLS）和DDB1下降。我们获得了优先的清单 我们预测的所有治疗中常见的候选人是CRL4的下游 活性并参与RBS/CDLS表型。在本提案的特定目的1中，我们 验证LC-MS数据并进一步测试这些目标的敲除 斑马鱼胚胎中的RBS/CDLS型发育缺陷 在ESCO2或SMC3表达中降低。在特定目标2中，我们检验假设 这些候选者的外源表达具有致变性。结合这些 研究将揭示新的目标，可以减少先天缺陷的严重性。