MOUSE MINUTES: A GENETIC STUDY OF RIBOSOMAL PROTEINS

小鼠分钟：核糖体蛋白的遗传学研究

• 批准号: 6941361
• 负责人: Thomas M. Glaser
• 金额: $ 30.49万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-09-01 至 2008-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6941361
• 关键词: cellular pathology; complementary DNA; embryonic stem cell; gene expression; gene expression profiling; gene mutation; genetic disorder; genetic models; genetic transduction; genetically modified animals; hematopoiesis; laboratory mouse; molecular pathology; phenotype; polysomes; protein structure function; retinal ganglion; ribosomal proteins; tissue /cell culture

DESCRIPTION (provided by applicant): The spontaneous mouse mutation Bst (belly spot and tail) causes midventral spotting, vertebral anomalies (tail kinks), preaxial polydactyly, a reduction in retinal ganglion cell (RGC) number, optic nerve aplasia, colobomata, and subretinal neovascularization (Rice et al. 1997; Smith et al. 2000). Bst homozygotes die before implantation. We have identified Bst as a deletion within Rpl24, the gene encoding the large subunit ribosomal protein L24. The mutation causes aberrant splicing in 80% of transcripts, leading to truncation of the L24 polypeptide. Mouse BAC and human cDNA transgenes correct the Bst/+ phenotype. Almost nothing is known about ribosomal protein (RP) gene mutations in vertebrates. In Drosophila, numerous ribosomal gene mutations, termed Minutes, are dispersed across the genome (Lambertsson 1998). They are homozygous lethal and have similar heterozygous phenotypes consisting of smaller, thinner bristles and profoundly delayed development (Schultz 1929). Their effects are not additive when combined. Minutes were instrumental in pioneering studies that defined concepts of cell autonomy, clonal lineage and developmental compartmentation, as Minute cells compete poorly with wild type clones in somatic mosaics (Simpson and Morata 1981). In humans, Diamond-Blackfan anemia (hereditary red blood cell aplasia with variable congenital anomalies) is caused by mutations in RPS19, which encodes a small subunit riboprotein (Willig et al. 1999). In view of the universal cellular requirement for ribosomes, the tissue specificity of mouse Rpl24 and human RPS19 phenotypes is puzzling. Our findings suggest that riboprotein defects may underlie common human malformation syndromes that have a suspected genetic etiology but no single target locus (Lalani et al. 2003). Fibroblasts from Bst/+ embryos grow slower than wild-type MEFs, remain longer in G1 phase, and exhibit decreased rates of protein synthesis. Although complete phenotypic surveys are lacking, the majority of tissues compensate for this cellular growth defect, so that organogenesis proceeds and Bst/+ mice approach normal adult size. While extraribosomal functions for L24 are possible, unique Bst phenotypes can also be explained by a dyschronic mechanism, in tissues such as the retina where proliferation and the sequence of cell fate determination are relatively uncoupled. In this proposal, we aim to: (1) complete characterization of Bst molecular, cellular and tissue developmental effects; (2) investigate the selective growth disadvantage of Rpl24 deficient cells in ROSA26 lacZ <-> Bst/+ blastocyst chimeras and somatic mosaics, as predicted by classical Minute studies; and (3) define the spectrum of riboprotein phenotypes by comparing Bst/+ mice to selected knockout mice with other RP mutations, including several that are available as targeted ES cells.

描述(申请人提供)：小鼠自发突变BST(腹斑和尾巴)会导致中腹区斑点、脊椎异常(尾部扭结)、轴前多指、视网膜神经节细胞(RGC)数量减少、视神经再生障碍性疾病、结缔组织瘤和视网膜下新生血管(Rice等人)。1997年；Smith等人。2000)。BST纯合子在植入前死亡。我们已经确定BST是编码大亚基核糖体蛋白L24的基因Rpl24中的一个缺失。该突变导致80%的转录本发生异常剪接，导致L24多肽被截断。小鼠BAC和人cDNA3基因可纠正BST/+表型。脊椎动物的核糖体蛋白(RP)基因突变几乎一无所知。在果蝇中，许多核糖体基因突变，称为分钟，分散在整个基因组中(Lambertsson 1998)。它们是纯合子致命性的，具有相似的杂合子表型，包括更小、更薄的刚毛和严重延迟的发育(Schultz 1929)。当它们结合在一起时，它们的影响不是相加的。在定义细胞自主性、克隆谱系和发育分区概念的开创性研究中，会议记录起到了重要作用，因为在体细胞嵌合体中，微小细胞与野生型克隆竞争很差(Simpson和Morata 1981)。在人类中，钻石-黑粉贫血(遗传性红细胞再生障碍性贫血伴各种先天性异常)是由RPS19突变引起的，RPS19编码一个小亚单位核蛋白(William等人。1999年)。鉴于细胞对核糖体的普遍需求，小鼠RPL24和人RPS19表型的组织特异性令人费解。我们的发现表明，核蛋白缺陷可能是常见的人类畸形综合征的基础，这些综合征有可疑的遗传病因，但没有单一的靶点(Lalani等人)。2003年)。 来自BST/+胚胎的成纤维细胞生长慢于野生型MEF，停留在G1期的时间更长，并且蛋白质合成速率降低。尽管缺乏完整的表型调查，但大多数组织弥补了这种细胞生长缺陷，因此器官发生进行，BST/+小鼠接近正常成年大小。虽然L24的体外功能是可能的，但在像视网膜这样的组织中，独特的BST表型也可以用动态机制来解释，在这些组织中，增殖和细胞命运决定的序列相对独立。 在这项建议中，我们的目标是：(1)完成BST分子、细胞和组织发育效应的表征；(2)研究Rp24缺陷细胞在rosa26 Lacz&lt；-&gt；-&gt；BST/+囊胚嵌合体和体细胞嵌合体中的选择性生长劣势；(3)通过比较BST/+小鼠和具有其他RP突变的选定敲除小鼠(包括几种可用作靶向ES细胞的基因敲除小鼠)，确定核蛋白表型的谱。