Meiosis is a critical stage of gametic differentiation and, particularly in male mammals, chromosome pairing and recombination are important determinants off gametic success. In pachytene spermatocytes of the mouse, the autosomal chromosomes are engaged in recombination and transcription while the sex chromosomes form a transcriptionally and recombinationally inactive domain known as the sex body. This feature permits the cytological and molecular investigations of meiotic chromosome structure and function that are proposed here. The goal of this work is to assess structural organization of meiotic nuclei and to determine how meiotic chromatin structure is related to meiotic function. Ultimately knowledge about the events of meiosis will lead to better understanding of the role of this important process in gametogenesis. The hypothesis to be tested is that the sex chromosomes of meiotic prophase nuclei assume a chromatin conformation that is different from the chromatin configuration of the paired and recombining autosomes, and that the sex chromosomes do not participate in events leading to effective pairing and recombination.
The first aim i s to determine differences in meiotic chromosome structure between DNA sequences on the sex chromosomes and sequences on the autosomal chromosomes. At the cytological level the technique of in situ nick translation will be used to assess differences in DNase sensitivity between the sex and autosomal chromatin domains. At the level of gene sequences, the relative DNase sensitivity of specific sequences in the sex and autosomal chromatin domains will be determined. DNase sensitivity assays will also be used to determine the chromatin configuration of the same gene sequences in both autosomal and sex chromatin domains to determine if they are under chromosomal or gene-sequence control with respect to chromatin configuration. Genetic and transgenic mouse models will be used for this experimental analysis.
The second aim i s to determine differences in meiotic function between the sex pair and the autosomal pairs. At the cytological level, differences in pachytene DNA synthesis will be assessed. Techniques of immunolocalization will be used to determine the nuclear distribution of proteins involved in pairing and recombination to learn if these proteins are excluded from the non- recombining sex chromatin domain. The results of these studies will contribute to our basic knowledge about meiosis, which is the unique and defining event of gametogenesis and is also an important control point in gametogenic differentiation.
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