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Newt (Cynops pyrrhogaster) testis proteomics during FSH/PRL stimulations

ABSTRACT: Newt testis is an appropriate model to isolate germ cells at a specific stage and study the effect of various factors on specific stage of spermatogenesis. Spermatogenesis is an essential process for sexual reproduction in development. It is triggered by the sequential mitotic divisions of spermatogonia, followed by their differentiation into spermatocytes. After two meiotic divisions, spermatocytes develop into spermatids, which possess half the normal complement of genetic material, and then into spermatozoa, mature male gametes in many sexually reproducing organisms. This complex process is controlled by cooperation with several hormones and testicular somatic cells, such as Follicle-stimulating hormone (FSH) and Prolactin (PRL). They are secreted by the pituitary gland and act on testicular somatic cells, mainly Sertoli cells, through the specific receptor. Sertoli cells have essential roles in the regulation of spermatogenesis. They not only represent the only cellular component of the blood–testis barrier but also produce and secrete local factors to germ cells. In this study, Primary Sertoli cells were established from Newt testis, and analyzed their proteome changes during the stimulation of FSH/PRL by 2D-DIGE (IC-Dyes).

ORGANISM(S): Cellular Organisms

SUBMITTER: Norie Araki

PROVIDER: PXD008342 | JPOST Repository | Sun Dec 01 00:00:00 GMT 2019

REPOSITORIES: jPOST

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Project description:The Rb/E2F tumor suppressor regulates gene expression to control the onset and timing of differentiation in many different cell types during development. This critical function makes Rb/E2F a frequent target for inactivation in tumors of diverse tissue origin. However, the mechanisms by which Rb/E2F governs tissue-specific gene regulation in vivo are poorly understood. We have determined the genome-wide binding profiles for components of the C. elegans Rb/E2F transcriptional regulatory pathway in the germ line, the intestine, and broadly in somatic tissues. Rb/E2F exhibits highly tissue-specific regulation, with unique sets of target genes that are activated or repressed depending on whether cells are in a progenitor (germ line) or terminally differentiated (somatic) state. In particular, LIN-35 binding is impaired in the germ line relative to the soma. Moreover, in the intestine, LIN-35 binds independently of EFL-1/DPL-1 at broad intergenic sites at which the heterochromatin protein HPL-2 also binds. Finally, recently defined âHOTâ sites, which are non-specifically bound by many unrelated factors, appear to be restricted primarily to somatic tissues, suggesting that germ line chromatin is in a state that globally restricts promiscuous association of regulatory factors. In sum, these tissue-specific binding profiles led to significant mechanistic insights into tissue-specific properties of Rb/E2F function in C. elegans that are likely relevant to other organisms. Staged C. elegans worms carrying different transgenes expressing GFP-transcription factors under different promoters were treated to ChIP-seq in biological replicate. A total of eleven different transgenic lines were analyzed. Three transcription factors (LIN-35, EFL-1 and DPL-1) were each tagged with GFP and expressed under the pie-1 promoter (germline), the ges-1 promoter (intestine), or their endogenous promoter (somatic expression), for a total of nine lines. In addition, LIN-35:GFP was expressed under the mex-5 promoter (germline), and HPL-2 was expressed under the ges-1 promoter, for an additional two lines. All strains with germline expression were analyzed as young adults, while the intestine or somatic expressed lines were analyzed as L1s. For each replicate of each transgenic line at the appropriate stage, anti-GFP was used to immunoprecipitate the tagged transcription factor, and the factor-enriched chromatin was subjected to Illumina sequencing. As a control, for every ChIP sample, the input (without immunoprecipitation) was also sequenced.

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Newt (Cynops pyrrhogaster) testis proteomics during FSH/PRL stimulations

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