ROLES OF THE MFH-1 AND WT1 GENES IN KIDNEY DEVELOPMENT

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We generated the knockout mice of the MIFH-1 gene. The MIFH-1 null mice displayed phenotypes of cardiovascular system and skeleton. The formers were interruption of the aortic arch and ventral septal defect, and the latters were defects arid malformation of the vertebrates and craniofacial bones. These results indicate that the MFH-1 gene is essential for aortic arch formation and skeletogenesis.The MFH-1 gene is also expressed in the kidney. In the MFH-1 knockout mice, the kidneys were a little small and the pelvis was malformed. However, the structures of glomerulus and renal tubules had no abnormalities. We interpret that other forkhead family genes, for example BF-1 and MF-1, might compensate the function of the MFH-1 gene.Next we investigated the target genes of MFH-1 gene. We screened the cell lines with monoclonal anti-mouse MFH-1 antibody in order to study whether the MFH-1 gene is expressed in the cells. We found that the MFH-1 protein was expressed in the Wilms tumor cell line, osteosarcoma cells, chondrosarcoma cells, aortic endothelial cells, aortic smooth muscle cells and fibroblast cells, NIH/3T3 and Balb/3T3. In order to investigate the molecular mechanism governing the transcriptional control of the MFH-1 gene in the kidney, we transfected the 3.Okbp of promoter and 5'-flanking region of the MFH-1 gene which was connected to the reporter gene luciferase into the Wilms tumor cell line. The construct which contains 3.0kbp flanking region showed a strong transcriptional activity in the Wilms tumor cell line. This result indicates that the 3.0 kbp region has elements regulating the expression of the MFH-1 gene positively in the nephroblastoma cells. Currently, the WT1-binding sites within the 3.0kbp region have been tried to identify by gel shift assay and cotransfection with the WT1-expression vector.