The goal of our research is to define the genes responsible for the lack of kidneys in an insertional mouse mutant that we have recently produced and to understand their role in kidney development. The mutant locus has been termed kdl for kidney: dysgenic, neonatal lethal (kdl). This mutation was identified as an insertional mutation due to a transgene integration. Homozygous mice die within 24 hours of birth, with abnormally developed kidneys. Gross histological examination reveals no abnormalities in other tissues. We have isolated sequences flanking the insertion site of the transgene and identified a gene encoding a novel G-protein coupled receptor, which has been disrupted by the insertion. We have cloned the cDNA of the receptor from rat and sequenced it entirely. This gene (hereby referred to as KDLR) is expressed predominantly in the adult mouse kidney and is identified in whole mouse embryo RNA as early as day 7. The amino terminus of the kdlr gene is deleted and kdl mice. A PAC clone which spans the deleted region has been isolated from the wild type mouse KDLR region and will be evaluated for other candidate genes. Preliminary mapping of the insert indicates that the maximum deleted region is less than 100 kb, and consequently likely to contain only KDLR or possibly a limited number of other genes.
The specific aims of our study include: 1) Complete the mapping of the kdl insertion site and the mKDLR wild type genomic loci: scanning, identification, and evaluation of other candidate genes; 2) Provide a detailed analysis of defects in kdl mutant mice and KDLR null mice including immunohistochemical, ultrastructural and morphometric characterizations. 3) Produce mice with targeted ablation of KDLR and other candidate genes and compare with the kdl phenotype; KDLRr is an excellent candidate, hence it will be tested concurrently with the mapping of the region. 4) Analysis of tissue specific and developmental expression of candidate genes including KDLR; and determine the human chromosomal location for KDLR. 5) Investigate the functional characterization of ligand binding and cellular activity of KDLR or other candidate genes. These studies will lead to an understanding of how the functions of new genes affect kidney development and provide an insight into the process of nephrogenesis.
