Exploring genotype-phenotype correlations in Sox10 mutations

项目描述：什么样的遗传变异导致什么样的表型变化？SOX蛋白是一个转录因子家族，在胚胎发育和细胞内环境稳定中具有关键功能，这些功能在脊椎动物中高度保守。由于SOX基因在多种细胞类型中的复杂作用，SOX突变体的表型往往是多样的，即使是单个基因。虽然对它们的蛋白质结构和DNA结合特征了解很多，但我们仍然不知道它们的结构与它们的细胞功能之间的关系，即使在充分研究的例子中，基因型-表型相关性也不清楚，例如SOX 10（Pingault et al.，2022，J. Med. Genet. 59，105-114）。SOX 10在神经嵴中广泛表达，神经嵴是高度多能性祖细胞的胚胎群体，SOX 10突变可单独或以各种组合导致色素、听觉、嗅觉和神经表型。表型变异可能反映了突变（例如功能获得）或其他因素（例如修饰基因座的存在）的微妙影响。我们已经显示了Sox 10在斑马鱼和哺乳动物中的保守作用（例如Kelsh，2006，Bioessays 28，788-798）。此外，在由同一团队监督的当前DTP学生项目的过程中，我们已经建立了一种方法，用于产生CRISPR/Cas9诱导的精确基因组修饰，使用化学调节来增强同源性定向修复（Zhang et al.，2018，J.Biol.Chem.293，6611-6622; Aksoy等人，2019，Communications Biology，2，198）。这与斑马鱼胚胎的易接近性和表型特征相结合（例如，Alhashem等人，2022，eLife 11：e73550; Camargo-Sosa等人，2019，PLoS Genetics 15，e1007941），使斑马鱼成为一个理想的系统，以探索在相对受限的遗传背景下特定突变变化的精确影响。为了评估基因型-表型差异，成功的申请人将创建一系列广泛的斑马鱼sox 10等位基因，从与疾病表型相关的人类变异中选择;突变将保持为杂合子，因为大多数可能是纯合致死的。显性和隐性表型，然后将becharterized定量为所有的色素和神经细胞类型。这将使我们能够解开目前模糊的，但基本的，结构与功能的关系，这一重要的发展调节因素。这个跨学科的项目将提供一个机会，发展许多具体的技能，包括斑马鱼遗传学和畜牧业，表型分析，包括原位杂交和免疫荧光，共聚焦和光片显微镜，分子生物学和生物信息学。此外，学生将受益于与小鼠和人类遗传学家在研究所想象（巴黎）的协作互动。

Project description: What genetic variants cause what phenotypic changes? The SOX proteins form a family of transcription factors with keyfunctions in embryonic development and cellular homeostasis, with these functions highly conserved across thevertebrates. Due to their complex roles in multiple cell-types, SOX mutant phenotypes are often diverse, even for a singlegene. Whilst much is known of their protein structure and DNA binding characteristics, we are still ignorant of therelationship between their structure and their cellular functions, with the genotype-phenotype correlation remainingobscure even in well-studied examples, such as SOX10 (Pingault et al., 2022, J. Med. Genet. 59, 105-114). SOX10 isexpressed widely in the neural crest, an embryonic population of highly multipotent progenitors, and SOX10 mutationsmay result in pigment, hearing, olfactory and neural phenotypes, individually or in various combinations. The variation inphenotypes may reflect subtle impacts of the mutations (e.g. gain of function), or other factors (e.g. presence of modifierloci). We have shown the conserved role for Sox10 in zebrafish and mammals (e.g. Kelsh, 2006, Bioessays 28, 788-798).Furthermore, in the course of a current DTP studentship supervised by same team, we have established a method forgenerating CRISPR/Cas9-induced precise genomic modifications, using chemical modulation to enhance Homology-Directed Repair (Zhang et al., 2018, J. Biol. Chem. 293, 6611-6622; Aksoy et al., 2019, Communications Biology, 2, 198).This, combined with the ready accessibility and phenotypic characterisation of zebrafish embryos (e.g. Alhashem et al.,2022, eLife 11:e73550; Camargo-Sosa et al., 2019, PLoS Genetics 15, e1007941), makes the zebrafish an ideal system toexplore the precise impacts of specific mutational changes in a relatively constrained genetic background.To assess the genotype-phenotype discrepancy, the successful applicant will create an extensive series of zebrafishsox10 alleles, selected from amongst the human variants linked to disease phenotypes; mutations will be maintained asheterozygotes, since most are likely to be homozygous lethal. Dominant and recessive phenotypes will then becharacterised quantitatively for all the pigment and neural cell-types. This will enable us to disentangle the currentlyobscure, but fundamental, structure-function relationships for this vital developmental regulatory factor. This interdisciplinary project will give an opportunity to develop numerous specific skillsets, including in zebrafish geneticsand husbandry, phenotypic analysis including by in situ hybridisation and immunofluorescence, confocal and lightsheet microscopy, molecular biology, and bioinformatics. In addition, the student will benefit from collaborativeinteraction with mouse and human geneticists at the Institut Imagine (Paris)