This subproject is one of many research subprojects utilizing theresources provided by a Center grant funded by NIH/NCRR. The subproject andinvestigator (PI) may have received primary funding from another NIH source,and thus could be represented in other CRISP entries. The institution listed isfor the Center, which is not necessarily the institution for the investigator.Male factor infertility is a significant concern throughout the world. The majority of the male infertility cases are idiopathic. In the male, primordial germ cells (PGCs) migrate, proliferate, and colonize the genital ridges to ultimately form testicular cords, where they establish contacts with the Sertoli cells. Key factors that regulate primordial germ cell migration and proliferation are not completely understood. The long-term goal of this project is to delineate the mechanisms of germ cell interactions with Sertoli cells in the testis. A mechanistic understanding of how germ cells develop and function is relevant to clinical conditions of male infertility that manifest as Sertoli cell-only syndrome for which there is currently no treatment. To begin to explore the developmental biology of the male germ cells and to understand the pathobiology of the human Sertoli cell-only syndrome, we have characterized atrichosis, the naturally occurring homozygous recessive mouse mutant. The atrichosis mutant testis histology closely resembles that of Sertoli cell-only syndrome patients and demonstrates tubules lined with only Sertoli cells and contains no germ cells.
Three Specific Aims are proposed to test the central hypothesis that a cell autonomous defect leads to complete absence of germ cells in the atrichosis mutant testis.
In Specific Aim 1, we will express a GFP transgene using a germ cell specific, Oct4 promoter in the atrichosis mutant background, and analyze the migration and proliferation of PGCs by confocal microscopy.
In Specific Aim 2, we will determine whether the germ cell loss is due to a PGC cell autonomous defect or due to a defective somatic cell niche compartment. In a co-culture in vitro assay, we will determine whether the Sertoli cells from atrichosis mutants establish functional contacts withwild type germ cells and initiate the formation of adherens junctions. Additionally, lacZ-tagged donor male germ cells will be transplanted into the mutant tubules and spermatogeneis monitred.
In Specific Aim 3, we will identify the candidate gene(s) at the atrichosis locus by positional cloning or a gene expression-based strategy using flow-sorted PGCs isolated from the GFP-positive atrichosis mutant embryos. Collectively, these studies will allow us to elucidate the mechanisms of development, function and fate of the germ cells and give new insights into how they influence somatic cells in the testis. These studies will identify the gene(s) responsible for the absence of germ cells in the atrichosis mutant mouse and provide a starting pointfor further loss-of-function and gain-of-function genetic approaches to understand germ cell migration and function. Finally, this work will establish atrichosis mutant as a genetically tractable new mouse model for human male infertility conditions associated with Sertoli cell-only tubules and germ cell aplasia, thus impacting clinical protocols of male fertility restoration.
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