Expression of the recently discovered cyclin A1 gene, Ccna1, is restricted to the adult testis and is germ cell-specific. Ccna1 mRNA and protein are expressed in late meiotic prophase spermatocytes. Targeted mutagenesis of Ccna1 resulted in viable progeny but male sterility, while females were fully fertile. Spermatocytes in cyclin A1-deficient mice have apparently normal chromosome synapsis but cannot enter into the first meiotic division. The activation of MPF is deficient and the cells undergo apoptosis. This proposal will extend studies on the unique regulation and function of cyclin A1 in the male germ line and to explore the functional redundancy of the two mammalian A-type cyclins.
The specific aims are to: (1) Identify regulatory elements required for the proper in vivo expression of Ccna1. This will involve the expression of reporter constructs in transgenic mice, sequence analysis, and a yeast one-hybrid screen to identify the transcription factors responsible for both activating Ccna1 in meiotic prophase spermatocytes as well as repressing its expression earlier in the male germ cell lineage and in other tissues. (2) Assess the functional redundancy of the two mouse A-type cyclins biochemically by systematically analyzing differences in substrate specificity ands protein-protein interactions between cyclin A1/Cdk and cyclin A2/Cdk complexes in vitro and in vivo. (3) Further explore the functional redundancy between cyclin A1 and cyclin A2 at the molecular genetic level in vivo by asking if cyclin A2 can rescue the cyclin A1- deficient phenotype of meiotic arrest in spermatogenesis, in transgenic mice in vivo. And (4) Study the unique role and functions of cyclin A1 in spermatogenesis by examining the effects of an amino acid change identified in the human CCNA1 gene in an infertile man. The corresponding mutation will be generated in the mouse Ccna1 gene and used to produce protein for in vitro biochemical studies and to generate transgenic mice expressing the altered cyclin A1 protein in vivo. These studies will provide new and novel insight into the function of the mammalian A-type cyclins, information that cannot be obtained from simpler model systems such as yeast, which lack this class of cyclins. Higher organisms such as mammals also contain two A-type cyclins, the functional significance of which remains to be understood. Given the clear role of cyclin A in male meiotic progression, our; studies will also further our understanding of the genetic control of mammalian meiosis.
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