Abstract
To understand the role of chromatin structure in the expression of the mouse protamine 1, protamine 2, and transition protein 2 genes during spermatogenesis, we have examined the genomic organization of this cluster of “haploid-specific” genes. As seen in the human genome, protamine 2, transition protein 2, and approximately 2.8 kb of a CpG island, hereafter called CpG island-dTP2, were clustered in a small region. Methylation analyses of this region have demonstrated that i) unlike most other tissue-specific genes, the protamine 1, protamine 2, and transition protein 2 genes were located in a large methylated domain in round spermatids, the cell type where they are transcribed, ii) the protamine 1 gene was only partially methylated in somatic cells and in testes from 7-day-old mice, and iii) the approximately 2 kb upstream and downstream of the CpG island-dTP2 were only partially methylated in somatic tissues. DNase I analysis revealed the presence of at least five strong DNase I hypersensitive sites over the CpG island-dTP2 in somatic tissues, but not in germ cells, and sequence analysis indicated that the CpG island-dTP2 is homologous to a CpG island located approximately 10.6 kb downstream of the human transition protein 2 gene. Although the nature of a CpG island-dTP2 and the function of a CpG island-dTP2-containing somatic tissue-specific DNase I hypersensitive sites in close proximity to the germ cell-specific gene cluster are unclear, the “open” chromatin structure of the CpG island-dTP2 may be responsible for the partial methylation pattern of the flanking sequences including the transition protein 2 gene in somatic tissues.
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Aizawa A, Yoneyama T, Kazahari K, Ono M (1995) DNase I-hypersensitive sites in the chromatin of rat growth hormone gene locus and enhancer activity of regions with these sites. Nucleic Acids Res 23, 2236–2244
Ammer H, Henschen A, Lee C (1986) Isolation and amino acid sequence analysis of human sperm protamines PI and P2. Occurrence of two forms of protamine P2. Biol Chem Hoppe-Seyler 367, 515–522
Antequera F, Macleod D, Bird AP (1989) Specific protection of methylated CpGs in mammalian nuclei. Cell 58, 509–517
Ariel M, McCarrey J, Cedar H (1991) Methylation patterns of testisspecific genes. Proc Natl Acad Sci USA 88, 2317–2321
Ariel M, Cedar H, McCarrey J (1994) Developmental changes in methylation of spermatogenesis-specific genes include reprogramming in the epidlidymis. Nature Genet 7, 59–63
Balhorn R (1989) Mammalian protamines: structure and molecular interactions. In Molecular Biology of Chromosome Function, KW Adolph (ed). (New York: Springer-Verlag), pp 366–395
Balhorn R, Weston S, Thomas C, Wyrobek AJ (1984) DNA packaging in mouse spermatids. Synthesis of protamine variants and four transition proteins. Exp Cell Res 150, 298–308
Bellvé AR, Cavicchia JC, Millette CF, O’Brien DA, Bhatnagar YM, Dym M (1977) Spermatogenic cells of the prepubertal mouse: isolation and morphological characterization. J Cell Biol 74, 68–85
Bird AP (1986) CpG-rich islands and the function of DNA methylation. Nature 321, 209–213
Bonifer C, Vidal M, Grosveld F, Sippel AE (1990) Tissue-specific and position independent expression of the complete gene domain for chicken lysozyme in transgenic mice. EMBO J 9, 2843–2848
Boyes J, Bird A (1991) DNA methylation inhibits transcription indirectly via a methyl-CpG binding protein. Cell 64, 1123–1134
Choi Y-C, Chae C-B (1991) DNA hypomethylation and germ cell-specific expression of testis-specific H2B histone gene. J Biol Chem 266, 20504–20511
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, 5538–5548
Choi Y-C, Gu W, Hecht NB, Feinberg AP, Chae C-B (1996) Molecular cloning of mouse somatic and testis-specific H2B histone genes containing a methylated CpG island. DNA Cell Biol 15, 495–504
Choudhary SK, Wykes SM, Kramer JA, Mohamed AN, Koppitch F, Nelson JE, Krawetz SA (1995) A haploid expressed gene cluster exists as a single chromatin domain in human sperm. J Biol Chem 270, 8755–8762
Coulondre C, Miller JH, Farabaugh PJ, Gilbert W (1978) Molecular basis of base substitution hotspots in Escherichia coli. Nature 274, 775–780
Gardiner-Garden M, Frommer M (1987) CpG islands in vertebrate genomes. J Mol Biol 196, 261–282
Grosveld F, Blom van Assendelft G, Greaves DR, Kollias G (1987) Position-independent, high-level expression of the human beta-globin gene in transgenic mice. Cell 51, 975–985
Iguchi-Ariga SMM, Schaffner W (1989) CpG methylation of the cAMP-responsive enhancer/promoter sequence TGACGTCA abolishes specific factor binding as well as transcriptional activation. Genes Dev 3, 612–619
Kadonaga J, Jones K, Tjian R (1986) Promoter-specific activation of RNA polymerase II transcription by Spl. Trends Biochem Sci 11, 20–23
Keime S, Heitland K, Kumm S, Schlosser M, Hroch N, Holtz W, Engel W (1992) Characterization of four genes encoding basic proteins of the porcine spermatid nucleus and close linkage of three of them. Biol Chem Hoppe-Seyler 373, 261–270
Kistler WS, Geroch ME, Williams-Ashman HG (1973) Specific basic proteins from mammalian testes: isolation and properties of small basic proteins from rat testes and epididymal spermatozoa. J Biol Chem 248, 4532–4543
Kistler WS, Noyes C, Hsu R, Heinrickson RL (1975) The amino acid sequence of a testis-specific basic protein that is associated with spermatogenesis. J Biol Chem 250, 1847–1853
Kleene KC, Distel RJ, Hecht NB (1985) Nucleotide sequence of a cDNA clone encoding mouse protamine 1. Biochemistry 24, 719–722
Kramer JA, Krawetz SA (1996) Nuclear matrix interactions within the sperm genome. J Biol Chem 271, 11619–11622
Krawetz SA, Herfort MH, Hamerton JL, Pon RT, Dixon GH (1989) Chromosomal localization and structure of the human PI protamine gene. Genomics 5, 639–645
Lichtenstein M, Keini G, Cedar H, Bergman Y (1994) B cell-specific demethylation: a novel role for the intronic k chain enhancer sequence. Cell 76, 913–923
Macleod D, Charlton J, Mullins J, Bird AP (1994) Spl sites in the mouse aprt gene promoter are required to prevent methylation of the CpG island. Genes Dev 8, 2282–2292
McKay DJ, Renaux BX, Dixon GH (1986) Human sperm protamines. Amino-acid sequences of two forms of protamine P2. Eur J Biochem 156, 5–8
Meehan RR, Lewis JD, Bird AP (1992) Characterization of MeCP2, a vertebrate DNA binding protein with affinity for methylated DNA. Nucleic Acids Res 20, 5085–5092
Meistrich ML (1989) Histone and basic nuclear protein transitions in mammalian spermatogenesis. In Histories and Other Basic Nuclear Proteins: Structure, Organization and Regulation of the Genes, LS Hnilica, GS Stein, JL Stein (eds) (Boca Raton, Fla: CRC Press), pp 165–182
Nelson JE, Krawetz SA (1993) Linkage of human spermatid-specific basic nuclear protein genes. J Biol Chem 268, 2932–2936
Nelson JE, Krawetz SA (1994) Characterization of a human locus in transition. J Biol Chem 269, 31067–31073
Queralt R, Oliva R (1995) Demonstration of trans-acting factors binding to the promoter region of the testis-specific rat protamine P1 gene. Biochem Biophys Res Commun 208, 802–812
Reeves RH, Gearhart JD, Hecht NB, Yelick P, Johnson P, O’Brien SJ (1989) Mapping of PRM1 to human Chromosome 16 and tight linkage of Prm-1 and Prm-2 on mouse chromosome 16. J Hered 80, 442–446
Schlüter G, Engel W (1995) The rat Prm3 gene is an intronless member of the protamine gene cluster and is expressed in haploid male germ cells. Cytogenet Cell Genet 71, 352–355
Schlüter G, Celik A, Obata R, Schlicker M, Hofferbert S, Schlung A, Adham IM, Engel W (1996) Sequence analysis of the conserved protamine gene cluster shows that it contains a fourth expressed gene. Mol Reprod Dev 43, 1–6
Stewart TA, Hecht NB, Hollingshead PG, Johnson PA, Leong JC, Pitts SL (1988) Haploid-specific transcription of protamine-myc and protamine-T-antigen fusion genes in transgenic mice. Mol Cell Biol 8, 1748–1755
Tamura T, Makino Y, Mikoshiba K, Muramatsu M (1992) Demonstration of a testis-specific trans-acting factor Tet-1 in vitro that binds to the promoter of the mouse protamine 1 gene. J Biol Chem 267, 4327–4332
Trasler JM, Hake LE, Johnson PA, Alcivar AA, Millette CF, Hecht NB (1990) DNA methylation and demethylation events during meiotic prophase in the mouse testis. Mol Cell Biol 10, 1828–1834
Tsukiyama T, Becker PB, Wu C (1994) ATP-dependent nucleosome disruption at a heat-shock promoter mediated by binding of GAGA transcription factor. Nature 367, 525–532
Yelick PC, Balhorn R, Johnson PA, Corzett M, Mazrimas JA, Kleene KC, Hecht NB (1987) Mouse protamine 2 is synthesized as a precursor whereas mouse protamine 1 is not. Mol Cell Biol 7, 2173–2179
Zambrowicz BP, Palmiter RD (1994) Testis-specific and ubiquitous proteins bind to functionally important regions of the mouse protamine-1 promoter. Biol Reprod 50, 65–72
Zambrowicz BP, Harendza CJ, Zimmermann JW, Brinster RL, Palmiter RD (1993) Analysis of the mouse protamine 1 promoter in transgenic mice. Proc Natl Acad Sci USA 90, 5071–5075
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Choi, YC., Aizawa, A. & Hecht, N.B. Genomic analysis of the mouse protamine 1, protamine 2, and transition protein 2 gene cluster reveals hypermethylation in expressing cells. Mammalian Genome 8, 317–323 (1997). https://doi.org/10.1007/s003359900431
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DOI: https://doi.org/10.1007/s003359900431