Spermatogenesis involves sequential changes of precursor cells to haploid spermatozoa through regulated mitotic, meiotic, and spermiogenic cell differentiation events. During meiotic process, somatic histones are replaced by spermatogenic cell-specific histones. The major objective of this application is to investigate regulation of somatic and testis-specific histone genes and also the role of testis-specific histones in assembly of new chromatin structure and meiosis during spermatogenesis. The opposing mechanisms of spermatogenic cell-specific transcription and repression of S-phase specific transcription elements in a testis-specific histone TH2B gene will be investigated by determining the DNA sequence elements required for the cell- specific transcription and the binding sites of nuclear factors on the regulatory elements. The DNA elements will be identified by sequential deletions of the 5' or 3' flanking sequence of the TH2B gene fused with a reporter gene, chloramphenicol acetyl transferase (CAT). Expression of CAT under the control of TH2B regulatory sequences will be examined by introduction of the fused genes into rat spermatogenic cells. The TH2B gene has the DNA element(s) which allows the gene expressed in a cell-cycle specific manner but the elements are repressed in the cells. The mechanism of repression will be investigated. The role of testis-specific histones in assembly of new chromatin structure and meiosis will be investigated by introducing rat H2B and TH2B genes into the yeast which do not produce H2B histones. The amino acid sequences of TH2B which alter the binding of H2B histones to DNA will be determined. Rat H1 and TH1 histone genes and also combination of H1 + H2B and TH1 + TH2B genes will be introduced into yeast cells to investigate effects of the histones on chromatin structure and meiosis. We will sequence the H2A gene on the 3' side of TH2B gene to determine if the histone gene encodes the testis-specific H2A histones.

Agency
National Institute of Health (NIH)
Institute
Eunice Kennedy Shriver National Institute of Child Health & Human Development (NICHD)
Type
Research Project (R01)
Project #
5R01HD020136-06
Application #
3318025
Study Section
Reproductive Biology Study Section (REB)
Project Start
1985-04-01
Project End
1991-03-31
Budget Start
1990-04-01
Budget End
1991-03-31
Support Year
6
Fiscal Year
1990
Total Cost
Indirect Cost
Name
University of North Carolina Chapel Hill
Department
Type
Schools of Medicine
DUNS #
078861598
City
Chapel Hill
State
NC
Country
United States
Zip Code
27599
Choi, Y C; Chae, C B (1998) The genomic and sequence analysis of rat histone H2B genes. DNA Seq 9:353-8
Choi, Y C; Gu, W; Hecht, N B et al. (1996) Molecular cloning of mouse somatic and testis-specific H2B histone genes containing a methylated CpG island. DNA Cell Biol 15:495-504
Choi, Y C; Han, B D; Chae, C B (1994) A B2-L1 composite between rat histone H2A and H3 genes. Mamm Genome 5:52-4
Choi, Y C; Chae, C B (1993) Demethylation of somatic and testis-specific histone H2A and H2B genes in F9 embryonal carcinoma cells. Mol Cell Biol 13:5538-48
Lim, K; Chae, C B (1992) Presence of a repressor protein for testis-specific H2B (TH2B) histone gene in early stages of spermatogenesis. J Biol Chem 267:15271-3
Huh, N E; Hwang, I W; Lim, K et al. (1991) Presence of a bi-directional S phase-specific transcription regulatory element in the promoter shared by testis-specific TH2A and TH2B histone genes. Nucleic Acids Res 19:93-8
Choi, Y C; Chae, C B (1991) DNA hypomethylation and germ cell-specific expression of testis-specific H2B histone gene. J Biol Chem 266:20504-11
Hwang, I W; Lim, K; Chae, C B (1990) Characterization of the S-phase-specific transcription regulatory elements in a DNA replication-independent testis-specific H2B (TH2B) histone gene. Mol Cell Biol 10:585-92
Lim, K; Chae, C B (1989) A simple assay for DNA transfection by incubation of the cells in culture dishes with substrates for beta-galactosidase. Biotechniques 7:576-9
Hwang, I; Chae, C B (1989) S-phase-specific transcription regulatory elements are present in a replication-independent testis-specific H2B histone gene. Mol Cell Biol 9:1005-13

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