Abstract
Immunohistochemical localization of a calmodulin-dependent protein phosphatase, calcineurin, was studied in the mouse testis in relation to previous observations showing that calmodulin is unusually rich in spermatogenic stages from mid-pachytene spermatocytes to elongating spermatids. The antibodies raised against calcineurin from scallop testis reacted with subunit B, but not subunit A, of calcineurin isoforms from mouse brain and testis. Indirect immunofluorescence using these antibodies on the mouse testis revealed positive reactions only in the nuclei of round or elongating spermatids: calcineurin started to accumulate in nuclei from the acrosomal cap phase, peaked at the initial stage of nuclear elongation, and decreased thereafter. There was almost no signal in the cytoplasm; spermatogenic cells at other stages, including spermatogonia, spermatocytes, mature sperm, and other somatic cells in the seminiferous tubules were totally negative. Immuno-electron microscopy gave the same result, on the basis of measuring the density of immunogold particles. These results suggest a role for calcineurin in remodeling of the nuclear chromatin in metamorphosing spermatids.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
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
Bode J, Willmitzer L, Opatz K (1977) On the competition between protamines and histones: studies directed towards the understanding of spermiogenesis. Eur J Biochem 72:393–403
Dooher GB, Bennett D (1973) Fine structural observations on the development of the sperm head in the mouse. Am J Anat 136:339–362
Ingebritsen TS, Cohen P (1983) The protein phosphatases involved in cellular regulation. 1. Classification and substrate specificities. Eur J Biochem 132:255–261
Kägi U, Chafouleas JG, Norman AW, Heizmann CW (1988) Developmental appearance of the Ca2+-binding proteins parvalbumin, calbindin D-28K, S-100 proteins and calmodulin during testicular development in the rat. Cell Tissue Res 252:359–365
Kakiuchi S, Yasuda S, Yamazaki R, Teshima Y, Kanda K, Kakiuchi R, Sobue K (1982) Quantitative determinations of calmodulin in the supernatant and particulate fractions of mammalian tissues. J Biochem 92:1041–1048
Klee CB, Crouch TH, Richman PG (1980) Calmodulin. Annu Rev Biochem 49:489–515
Mali P, Welsh MJ, Toppari J, Vihko KK, Parvinen M (1985) Cell interactions in the seminiferous epithelium with special reference to the cellular distribution of calmodulin. Med Biol 63:237–244
Moriya M, Katagiri Ch, Yagi K (1993) Immuno-electron-microscopic localization of calmodulin and calmodulin-binding proteins in the mouse germ cells during spermatogenesis and maturation. Cell Tissue Res 271:441–451
Mukai H, Chang CD, Tanaka H, Ito A, Kuno T, Tanaka Ch (1991) cDNA cloning of a novel testis-specific calcineurin B-like protein. Biochem Biophys Res Commun 179:1325–1330
Muramatsu T, Giri PR, Higuchi S, Kincaid RL (1992) Molecular cloning of a calmodulin-dependent phosphatase from murine testis: identification of a developmentally expressed nonneural isoenzyme. Proc Natl Acad Sci USA 89:529–533
Nishio H, Matsui H, Etoh S, Moia LJMP, Tokuda M, Itano T, Hatase O (1992a) Identification of testis specific calcineurin β subunit isoform by monoclonal antibody and detection of a specific six amino acid sequence. Biochem Biophys Res Commun 182:34–38
Nishio H, Matsui H, Moia LJMP, Taketa S, Miyamoto K, Tokuda M, Itano T, Nakahara S, Hatase O (1992b) The evidence for post-meiotic expression of a testis-specific isoform of a regulatory subunit of calcineurin using a monoclonal antibody. Biochem Biophys Res Commun 187:828–831
Oakberg EF (1956) A description of spermiogenesis in the mouse and its use in analysis of the cycle of the seminiferous epithelium and germ cell renewal. Am J Anat 99:391–413
Sano M, Ohshima AS, Kawamura N, Kitajima S, Mizutani A (1987) Immunohistochemical study of calmodulin in developing mouse testis. J Exp Zool 241:51–59
Slaughter GR, Meistrich ML, Means AR (1989) Expression of RNAs for calmodulin, actins, and tubulins in rat testis cells. Biol Reprod 40:395–405
Talian JC, Olmsted JB, Goldman RD (1983) A rapid procedure for preparing fluorescein-labeled specific antibodies from whole antiserum: its use in analyzing cytoskeletal architecture. J Cell Biol 97:1277–1282
Tash JS, Krinks M, Patel J, Means RL, Klee CB, Means AR (1988) Identification, characterization, and functional correlation of calmodulin-dependent protein phosphatase in sperm. J Cell Biol 106:1625–1633
Towbin H, Staehelin T, Gordon J (1979) Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci USA 76:4350–4354
Wolff DJ, Ross JM, Thompson PN, Brostrom MA, Brostrom CO (1981) Interaction of calmodulin with histones. Alteration of histone dephosphorylation. J Biol Chem 256:1846–1860
Yamamoto N (1985) Immunoelectron microscopic localization of calmodulin in guinea pig testis and spermatozoa. Acta Histochem Cytochem 18:199–211
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Moriya, M., Fujinaga, K., Yazawa, M. et al. Immunohistochemical localization of the calcium/calmodulin-dependent protein phosphatase, calcineurin, in the mouse testis: its unique accumulation in spermatid nuclei. Cell Tissue Res. 281, 273–281 (1995). https://doi.org/10.1007/BF00583396
Received:
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF00583396