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: 6823786
• 负责人: Thomas M. Glaser
• 金额: $ 30.95万
• 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/6823786
• 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（由申请人提供）：自发小鼠突变BST（腹部和尾部）引起室内中间斑点，椎骨异常（尾部扭结），预性多态度，视网膜神经节细胞（RGC）数量降低，数量降低，光神经，光神经aplasia，colobomata，colobomata，colobomata，collebomatical and rectertactal Neovearnic Neoveal nerevalsalial n. 1997； BST纯合子在植入前死亡。我们已经将BST确定为RPL24中的缺失，该基因编码了大型亚基核糖体蛋白L24。该突变导致80％的转录本中引起异常剪接，从而导致L24多肽的截断。小鼠BAC和人cDNA转基因纠正BST/+表型。关于脊椎动物中核糖体蛋白（RP）基因突变几乎一无所知。在果蝇中，大量的核糖体基因突变分别在基因组中分散（Lambertsson 1998）。它们是纯合致死的，具有相似的杂合表型，包括较小，较薄的刷毛和深刻延迟的发育（Schultz 1929）。合并后，它们的效果不是加性的。会议记录在开创性的研究中发挥了作用，该研究定义了细胞自主，克隆谱系和发育隔室的概念，因为微小细胞与体细胞镶嵌的野生型克隆竞争不佳（Simpson and Morata 1981）。在人类中，钻石黑粉刺贫血（具有可变先天性异常的遗传性红细胞性植物）是由RPS19中的突变引起的，RPS19编码了一个小的亚基核糖蛋白（Willig etal。1999）。鉴于核糖体的通用细胞需求，小鼠RPL24和人RPS19表型的组织特异性令人困惑。我们的发现表明，核蛋白缺陷可能是人类畸形综合症的基础，该综合症状综合症具有怀疑的遗传病因，但没有单个靶点基因座（Lalani等，2003）。 来自BST/+胚胎的成纤维细胞比野生型MEF慢，在G1期保持较长，并且蛋白质合成速率降低。尽管缺乏完整的表型调查，但大多数组织会补偿这种细胞生长缺陷，因此会进行器官发生，而BST/+小鼠接近正常的成人大小。尽管可能是L24的外虫体功能，但在诸如视网膜等组织中，唯一的BST表型也可以通过功能障碍的机制来解释，其中增殖和细胞命运测定的序列相对取消耦合。 在此提案中，我们的目标是：（1）BST分子，细胞和组织发育效果的完整表征； （2）根据经典的分钟研究预测，研究Rosa26 Lacz <-> BST/+胚泡嵌合体和体细胞镶嵌的RPL24缺陷细胞的选择性生长劣势； （3）通过将BST/+小鼠与选定的基因敲除小鼠与其他RP突变进行比较，定义了核蛋白表型的光谱，包括几种可作为靶向ES细胞可用的。