MAMMALIAN SPERMATOGENESIS--HISTONES/DNA REORGANIZATION

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Mammalian spermatogenesis is characterized by the progressive condensation of chromatin as protamines replace histones on the DNA. A novel mouse H2b gene which is transcribed and translated specifically in post-meiotic round spermatids codes for a protein which contains an extremely hydrophobic carboxyl-terminus, suggesting that it may interact with the nuclear membrane. The experiments in this proposal are designed to examine: 1) the signals that are important in restricting the transcription and translation of this gene to a particular spermatogenic population, i.e., round spermatids; and, 2) the possible role this protein has in chromatin reorganization. Analysis of the spermatid-specific H2b gene may identify sequence elements that are critical in restricting its expression. Unlike most other histone mRNAs, this transcript is polyadenylated. As spermiogenesis occurs over a two week period in the mouse, the possible role of polyadenylation in stabilizing these transcripts will be examined by generating transformed somatic cell lines with constructs containing the gene with and without its polyadenylation signal sequence. An antiserum raised against the unique carboxyl-terminal region will identify this protein. In particular, its localization within the haploid nucleus and its fate as spermiogenesis proceeds will be examined. The function of the protein in chromatin organization will be assessed by its production in transformed cell lines and its overproduction in transgenic mice. As nuclear abnormalities frequently are observed in sperm of genetically sterile mice, the lack of expression of specific genes may hinder normal chromatin condensation and subsequent sperm development. Reorganization of the chromatin may permit localization of specific DNA sequences during spermiogenesis. This compartmentalization may be important for transcriptional regulation both late in spermatogenesis and in the early stages of embryonic development.