Identification of genes involved in urogenital development

• Main Author: Van Hateren, Nick
• Published: University of Oxford 2002
• Subjects: 599; Genetics
• Online Access: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.249217

The mouse urogenital system comprises the gonads, mesonephroi, kidneys and adrenal glands. The indifferent gonad, or genital ridge, arises on the ventro-medial aspect of the mesonephros at approximately 10 days post coitum (dpc), and enlarges due to the influx of germ cells and migration of somatic cells from the coelomic epithelium and mesonephros. Male and female gonads are morphologically indistinguishable until 12.5 dpc when the expression of the testis-determining gene Sry in the gonadal supporting cell lineage results in the differentiation of Sertoli cells. This process initiates a cascade of gene expression leading to testis development and masculinisation of the embryo. In the absence of Sry the default ovarian pathway ensues. This project aims to identify genes exhibiting a spatio-temporal expression pattern consistent with a role in the development of the murine reproductive organs, with the primary focus on genes involved in Sertoli cell development. The main resource exploited was a normalised mouse urogenital ridge (NMUR) cDNA library constructed from gonad-mesonephros pairs at 11.5 dpc and 12.5 dpc. This library was screened by two methods: the first screen used sample sequencing of 472 NMUR cDNA clones to identify clones displaying homology to known developmentally important genes. The second screen employed DNA microarray technology to identify genes displaying differential expression in male and female gonads. The expression of candidate clones from both screens was examined by wholemount in situ hybridisation to identify clones displaying male-specific expression. These clones were studied in more detail to determine their cell-type specificity and timing of onset of expression. Two genes were found to display male-specific expression in Sertoli cells. One gene was the serine protease inhibitor, protease nexin 1 (Pn1); the other was a novel gene. The preliminary characterisation of this novel gene, termed Maestro (Mro), is presented. Pn1 and Mro are both expressed in a male-specific fashion prior to overt sexual differentiation. This timing, together with their expression in Sertoli cells, makes these genes candidates for regulation by the known sex-determining transcription factors (SRY, SOX9 etc.). In order to study the regulation of Pn1 expression during gonad development, a LacZ reporter construct containing a genomic region immediately upstream of the Pn1 gene was used to generate transgenic mice. This fragment did not drive Sertoli-cell specific expression of the reporter gene; therefore, the minimal gonadal enhancer for Pn1 could not be identified.