| Budget Amount *help |
¥3,700,000 (Direct Cost: ¥3,700,000)
Fiscal Year 2004: ¥1,400,000 (Direct Cost: ¥1,400,000)
Fiscal Year 2003: ¥2,300,000 (Direct Cost: ¥2,300,000)
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| Research Abstract |
Sex determination is an essential step to give the fetus the sex specific defferntiation program. Previous research investigating gene cascade underlining mouse gonadal development revealed that many transcriptional factors, growth factors, and chromatin proteins were necessary for adequate sex differentiation. Among these factors, in this project, we focused on the relation ship of transcription factors Ad4BP/SF-1 and Sox9, and chromatin proteins M33. Disruption of the murine M33 gene, a PcG (Polycomb Group) member, displayed posterior transformation of the vertebral columns and sternal ribs. In addition, failure of T-cell expansion and hypoplasia and sex-reversal of the gonads, have been observed. In the present study, we identified defects in the splenic and adrenal formation of M33-knockout (KO) mice on a C57BL/6 genetic background. The spleen in these animals was smaller than in the wild-type and was spotted red due to non-uniform distribution of blood cells. Histological examinat
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ion revealed disorganization of the vascular endothelium and its surrounding structures, and immunohistochemistry demonstrated disturbances in vascular formation and colonization of immature hematopoietic cells. These splenic phenotypes observed in the M33-KO mice were quite similar to those seen in Ad4BP/SF-1 (Nr5a1) knockouts. Moreover, the adrenal glands of M33-KO and Ad4BP/SF-1 heterozygous KO mice were smaller than those of the wild-type. Western-blot, immunohistochemistry and RT-PCR analyses of the M33 knockouts all indicated significantly low expression of Ad4BP/SF-1, indicating that M33 is an essential upstream regulator of Ad4BP/SF-1. In agreement with these observations, chromatin immunoprecipitation assays with adrenocortical Y-1 cells revealed direct binding of the M33-containing PcG to the Ad4BP/SF-1 gene locus. We also detected relationship of these two genes in gonadal development. On the other hand, sex reversal phenotype of M33-KO mice were rescued by Sox9 overexpression. These observation indicate the possibilities that the other target locus of M33 is the Sox9 locus. Less
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