We previously constructed a sequence tagged site (STS)/Bacterial Artificial Chromosome based physical map in mouse t-complex region and sequenced 43 BACs in conjunction with Mouse Genome Consortium. The mouse t-complex on chromosome 17 has long been a focus of interest for developmental biologists. Because many embryonic lethal mutations are localized in the region, corresponding genes of importance during development are inferred to reside there. ? 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 homozygotic 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 that maps to the same 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 based on its structural domains. Recently ADAMTS10 was found to be a candidate gene for Weill-Marchesani syndrome (OMIM#277600). We have 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 fused a to V5-epitope in a Gateway vector system and transfected into 293 cells. We find that both isoforms are stably expressed and localize to cytoplasm as determined by immunocytochemistry. We are currently completing the characterization of the spatial and temporal expression of ADAMTS10 in the embryo.
|Abe, Kuniya; Yuzuriha, Misako; Sugimoto, Michihiko et al. (2004) Gene content of the 750-kb critical region for mouse embryonic ectoderm lethal tcl-w5. Mamm Genome 15:265-76|
|Kargul, G J; Nagaraja, R; Shimada, T et al. (2000) Eleven densely clustered genes, six of them novel, in 176 kb of mouse t-complex DNA. Genome Res 10:916-23|