Development of a male contraceptive could in principle be achieved through perturbation of the reproductiveendocrine axis, disruption of spermatogenesis in the testis, inhibition of sperm maturation in the epididymis,or alteration of sperm structure or function during ejaculation. Within the testis one could imagine blockingthe initiation of spermatogenesis, the progression through meiosis, or the terminal differentiation ofspermatids. Genetic models for the feasibility of some of these scenarios currently exist. For this proposal wehave chosen to expand the basic knowledge of spermatogonial stem cells under the belief that in the future itmight be feasible to develop contraceptive agents that reversibly block the initiation of spermatogenesis.Continued spermatogenesis in mammals requires maintaining an exquisite balance between the selfrenewaland differentiation of a small population of spermatogonial stem cells (SSCs). Until recently almostnothing was known about the maintenance of SSC self-renewal. Rapid advances in this exciting area haverevealed that SSCs respond to at least two independent extrinsic signaling pathways emanating from Sertolicells. Several intrinsic signals help maintain SSC homeostasis. An unexpected discovery has been therealization that adult SSCs can readily convert to a pluripotent state that appears very much like that of anembryonic stem cell, offering the promise that SSCs may be useful in regenerative medicine. During theprevious funding period we showed that the promyelocytic leukemia zinc finger protein, Plzf, is expressed inSSCs and required for their self-renewal. Loss of Plzf function shifts the balance between stem cell selfrenewaland differentiation, toward differentiation at the cost of self-renewal. Plzf is a transcriptionalrepresser that has been shown to associate with members of the N-Cor2/SMRT co-repressor complex, andwith Bmi1, a Polycomb group protein and component of the Polycomb repressive complex 1. Both of thesecomplexes confer epigenetic modifications to chromatin. Our hypothesis is that Plzf recruits members of theNcor-2/SMRT co-repressor complex, and perhaps components of the Polycomb group repressive complex 1,to pluripotency and/or differentiation genes in SSCs. Loss of Plzf creates an unstable epigenetic state thatcompromises self-renewal. A full understanding of how Plzf functions to maintain SSC self-renewal willrequire identification of the transcriptional targets of Plzf, investigation of the molecular mechanism by whichPlzf represses transcription in SSCs, and characterization of the various signaling pathways in the SSCniche that converge to regulate Plzf and the other intrinsic factors required for SSC self-renewal. The threeSpecific Aims in this proposal attempt to address each of these issues.
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