The vast majority of mRNAs in meiotic and haploid spermatogenic cells in mice exhibit high levels of translationally repressed free messenger ribonucleoprotein particles (mRNPs). Some mRNAs, such as protamine mRNAs, are stored as free-mRNPs in early haploid cells and recruited onto polysomes at specific times in late haploid cells. The proportions of other mRNAs in repressed free-mRNPs in meiotic and early haploid cells range from virtually all to none. Thus, translation of nearly every individual mRNA in meiotic and haploid spermatogenic cells is regulated by mRNA- specific controls specifying the stage of translational repression and/or the proportion of free-mRNPs. Very little is known about the mRNP proteins in spermatogenic cells, even though these proteins are likely to determine the translational competence of spermatogenic mRNAs. The overall objective of this proposal is to elucidate the structure and translational competence of free-mRNP and polysomal mRNP particles in meiotic and haploid cells in mice. The experiments are designed to characterize the abundant mRNP proteins, specifically those mRNP proteins that are complexed with large numbers of different mRNAs. These mRNP proteins are predicted to consist of two well- characterized proteins, the poly(A) binding proteins and Y- box proteins, and a set of unidentified proteins. There is growing evidence that Y-box proteins function as non- specific translational repressors by packaging many mRNAs in a wide variety of eukaryotic cells into free-mRNPs. It follows that the structure and translational properties of free-mRNPs may be determined primarily by Y-box proteins. The basic premise of this proposal is that elucidation of the general structure of free-mRNPs will be crucial for understanding translational control of individual mRNAs. A major objective of this proposal is to characterize the levels of various mRNP proteins in the cytosolic, polysomal and free-mRNP fractions in several stages of spermatogenic cells that exhibit high levels of translationally repressed free-mRNPs. The translational competence of native free- mRNPs and polysomal mRNPs will be compared in the reticulocyte and wheat germ cell-free translation system. This proposal is intended to introduce talented undergraduates to research in molecular biology.
