Gene Candidates for Embryonic Lethals in the The Mouse T-complex

小鼠 T 复合体中胚胎致死的候选基因

• 批准号: 7592023
• 负责人: Ramaiah Nagaraja
• 金额: $ 6.15万
• 依托单位: NATIONAL INSTITUTE ON AGING
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/7592023
• 关键词: Attention; Bacterial Artificial Chromosomes; Candidate Disease Gene; Cells; Chromosome Mapping; Chromosomes, Human, Pair 17; Classification; Complex; Cytoplasm; Databases; Deposition; Development; Embryo; Embryonic Development; Embryonic Lethal Mutation; Epitopes; Genes; Goals; Homologous Gene; Human; Kidney; Length; Localized; Lung; Maps; Metalloendopeptidases; Metalloproteases; Mus; Mutate; Mutation; Open Reading Frames; Oryctolagus cuniculus; Physical Map of the Human Genome; Play; Protein Isoforms; RNA Splicing; Role; Sequence Tagged Sites; Staging; Syndrome; Tissues; Translations; Zinc; base; immunocytochemistry; intercellular communication; interest; mouse genome; physical mapping; polyclonal antibody; protein function

From a sequence tagged site (STS)/Bacterial Artificial Chromosome based physical map in mouse t-complex region 43 BACs were representing a minimal tiling path were selected, sequenced, annotated and deposited into the public databases. Based on the physical map the tw5 embryonic lethal locus mutation was refined to a single, 175 Kb BAC. Histological studies have demonstrated that mice homozygous for tw5 die at the gastrulation stage. The candidate BAC rescues the tw5 lethality, restoring viability. Our collaborator Dr. K. Abe is currently focusing on the search for the gene mutated in the tw5 embryonic lethality. Currently, we have focused our attention on the mouse homolog of human ADAMTS10 gene, which maps to the interval. Several metalloproteases play critical roles in cell-cell signal transduction. ADAMTS-10 gene, a zinc metalloendopeptidase with thrombospondin domains, is a potential candidate for important functions during development. Recently ADAMTS10 was found to be a candidate gene for Weill-Marchesani syndrome (MIM#277600). Previously we found that mouse ADAMTS-10 is alternatively spliced in a tissue-specific manner and encodes protein isoforms that either include or exclude the thrombospondin domains by truncation of the open reading frame. We have cloned and characterized 3 major isoforms from embryo, kidney and lung, and find that the embryo, for example, expresses an isoform that has a truncated translation product. The truncated embryonic and full-length kidney isoforms were transfected into 293 cells. We find that both isoforms are stably expressed and localize to the cytoplasm, as determined by immunocytochemistry. Rabbit polyclonal antibodies were generated targeted to epitopes that could distinguish between different isoforms and are being purified and characterized for further use to define the protein function.

从小鼠t复合体区基于序列标记位点(STS)/细菌人工染色体的物理图谱中选择43个代表最小平铺路径的BACs，进行测序、注释并存入公共数据库。根据物理图谱，将tw5胚胎致死位点突变细化为单个175 Kb BAC。组织学研究表明，纯合子为tw5的小鼠在原肠胚期死亡。候选BAC挽救了tw5的杀伤力，恢复了生存能力。我们的合作者K. Abe博士目前正专注于寻找导致胚胎死亡的基因突变。目前，我们主要关注的是人类ADAMTS10基因的小鼠同源性，该同源性映射到区间。几种金属蛋白酶在细胞-细胞信号转导中起重要作用。ADAMTS-10基因是一种具有血小板反应蛋白结构域的锌金属内肽酶，在发育过程中具有重要的潜在功能。最近，ADAMTS10被发现是Weill-Marchesani综合征（MIM#277600）的候选基因。先前我们发现小鼠ADAMTS-10以组织特异性的方式选择性拼接，并通过截断开放阅读框编码包含或排除凝栓蛋白结构域的蛋白质同工型。我们从胚胎、肾脏和肺中克隆并鉴定了3种主要的异构体，发现胚胎中表达的异构体具有截断的翻译产物。将截短的胚胎肾和全长肾同种异构体转染293细胞。我们发现这两种异构体都稳定地表达并定位于细胞质，这是由免疫细胞化学确定的。兔多克隆抗体是针对可以区分不同同种异构体的表位产生的，并正在纯化和表征，以进一步用于确定蛋白质的功能。

项目成果

Gene content of the 750-kb critical region for mouse embryonic ectoderm lethal tcl-w5.

小鼠胚胎外胚层致死 tcl-w5 的 750 kb 关键区域的基因内容。

• DOI: 10.1007/s00335-003-2329-1
• 发表时间: 2004
• 期刊: Mammalian genome : official journal of the International Mammalian Genome Society
• 作者: Abe,Kuniya;Yuzuriha,Misako;Sugimoto,Michihiko;Ko,MinoruSH;Brathwaite,Mgavi;Waeltz,Paul;Nagaraja,Ramaiah
• 通讯作者: Nagaraja,Ramaiah

Eleven densely clustered genes, six of them novel, in 176 kb of mouse t-complex DNA.

176 kb 的小鼠 T 复合体 DNA 中有 11 个密集聚集的基因，其中 6 个是新基因。

• DOI: 10.1101/gr.10.7.916
• 发表时间: 2000
• 期刊: Genome research
• 影响因子: 7
• 作者: Kargul,GJ;Nagaraja,R;Shimada,T;Grahovac,MJ;Lim,MK;Nakashima,H;Waeltz,P;Ma,P;Chen,E;Schlessinger,D;Ko,MS
• 通讯作者: Ko,MS