Zebrafish model of Peters-plus syndrome

描述（申请人提供）：彼得斯加综合征（PPS）是一种常染色体隐性遗传的可变复杂疾病。PPS常见的特征有：眼前段缺损、身材矮小、趾短、唇裂和/或腭裂、各种颅面缺损、脑、心、泌尿生殖系统异常。最近发现PPS综合征是由编码糖基转移酶的B3GALTL基因突变引起的。糖基转移酶在各种生物过程中发挥着重要作用，是催化糖分子附着到特定受体位点的酶。PPS患者有两个突变的B3GALTL等位基因，通常要么是基因的完全缺失，要么是核苷酸的改变，预计会导致缺乏催化结构域的截断蛋白。B3GALTL突变似乎可以解释大多数经典PPS，但在具有高度相似疾病的人类患者中尚未发现该基因突变。目前尚不清楚PPS的发育机制及相关情况，也未建立b3galtl缺乏症/PPS的动物模型。研究者的初步研究已经在斑马鱼中发现了两个b3galtlb基因，它们的表达模式和敲低表型（对于b3galtlb）与人类的PPS特征一致。由于PPS具有高度可变的表型，因此生成斑马鱼PPS模型对于该病的遗传解剖和鉴定其他致病或修饰位点/基因至关重要。在本申请中，她计划在Specific Aim 1中重点确定斑马鱼b3galtl基因的功能序列和表达模式，包括b3galtl基因的转录本、基因组结构、功能调控区域和表达模式的表征；在Specific Aim 2中，通过分析morpholino介导的单基因和双基因敲低，研究与b3galtl基因缺陷相关的表型。通过锌指核酸酶技术生成的斑马鱼系将显示出强大的表型，以创建脊椎动物模型，研究与B3GALTL/ B3GALTL缺乏相关的复杂和可变特征。

DESCRIPTION (Provided by Applicant): Peters-plus syndrome (PPS) is a variable complex disorder with autosomal recessive inheritance. The following features are commonly seen in PPS: ocular anterior segment defects, short stature, brachydactyly, cleft lip and/or cleft palate, various craniofacial defects, and brain, heart, as well as genitourinary anomalies. PPS syndrome was recently discovered to be caused by mutations in the B3GALTL gene that encodes a glycosyltransferase. Glycosyltransferases are known to play important roles in various biological processes, and represent enzymes that catalyze the attachment of a sugar molecule to specific acceptor sites. Patients with PPS have two mutated alleles of B3GALTL, typically either a complete deletion of the gene or nucleotide changes predicted to result in a truncated protein lacking its catalytic domain. B3GALTL mutations appear to explain the majority of classic PPS, but no mutations in this gene have been identified in human patients with highly similar disorders. At this time, the developmental mechanisms of PPS and related conditions are not known, and no animal models have been established for B3GALTL-deficiency/PPS. The investigator's preliminary studies have identified two b3galtl genes in the zebrafish that demonstrate expression patterns and knockdown phenotype (for b3galtlb) consistent with PPS features in humans. Since PPS is characterized by a highly variable phenotype, generation of the zebrafish model of PPS is essential to genetic dissection of this disease and identification of additional causative or modifying loci/genes. In this application, she plans to focus, in Specific Aim 1, to determine the functional sequences and expression patterns of zebrafish b3galtl genes, which involves characterization of transcripts, genomic structures, functional regulatory regions, and expression patterns of the b3galtl genes; and in Specific Aim 2, to examine phenotypes associated with deficiency for the b3galtl genes through analyses of morpholino-mediated single as well as double gene knockdowns. Generation of zebrafish lines via zinc-finger nuclease technology will be initiated for gene(s) that demonstrate a robust phenotype to create vertebrate model(s) to study the complex and variable features associated with B3GALTL/b3galtl deficiency. NARRATIVE: Peters-plus syndrome (PPS) is a complex disorder that involves abnormal development of the eye, head, brain, hands, heart, kidney, and stature. Mechanism(s) of PPS and related debilitating conditions are largely unknown. Mutations in B3GALTL gene were shown to explain a significant portion of patients with these phenotypes. Studies in animal models can provide a better understanding of underlying developmental mechanisms and involvement of additional factors. In this application, a classic vertebrate developmental model, zebrafish, will be explored to gain insight into developmental processes associated with orthologous b3galtl genes in zebrafish to better understand mechanisms of PPS and related conditions in humans.