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Transcriptional, biochemical, and immunohistochemical analyses of CaMKKß/2 splice variants that co-localize with CaMKIV in spermatids.
Ohtsuka, Satomi; Miyai, Yumi; Mima, Hiroyuki; Magari, Masaki; Chiba, Yoichi; Suizu, Futoshi; Sakagami, Hiroyuki; Ueno, Masaki; Tokumitsu, Hiroshi.
Affiliation
  • Ohtsuka S; Applied Cell Biology, Graduate School of Interdisciplinary Science and Engineering in Health Systems, Okayama University, Okayama, 700-8530, Japan.
  • Miyai Y; Inflammation Pathology, Department of Pathology and Host Defense, Faculty of Medicine, Kagawa University, Kagawa, 761-0793, Japan.
  • Mima H; Applied Cell Biology, Graduate School of Interdisciplinary Science and Engineering in Health Systems, Okayama University, Okayama, 700-8530, Japan.
  • Magari M; Applied Cell Biology, Graduate School of Interdisciplinary Science and Engineering in Health Systems, Okayama University, Okayama, 700-8530, Japan.
  • Chiba Y; Inflammation Pathology, Department of Pathology and Host Defense, Faculty of Medicine, Kagawa University, Kagawa, 761-0793, Japan.
  • Suizu F; Oncology Pathology, Department of Pathology and Host Defense, Faculty of Medicine, Kagawa University, Kagawa, 761-0793, Japan.
  • Sakagami H; Department of Anatomy, Kitasato University School of Medicine, Kanagawa, 252-0374, Japan.
  • Ueno M; Inflammation Pathology, Department of Pathology and Host Defense, Faculty of Medicine, Kagawa University, Kagawa, 761-0793, Japan.
  • Tokumitsu H; Applied Cell Biology, Graduate School of Interdisciplinary Science and Engineering in Health Systems, Okayama University, Okayama, 700-8530, Japan. Electronic address: tokumit@okayama-u.ac.jp.
Cell Calcium ; 117: 102820, 2024 01.
Article de En | MEDLINE | ID: mdl-37979343
Ca2+/calmodulin-dependent protein kinase kinase (CaMKK) phosphorylates and activates downstream protein kinases, including CaMKI, CaMKIV, PKB/Akt, and AMPK; thus, regulates various Ca2+-dependent physiological and pathophysiological pathways. Further, CaMKKß/2 in mammalian species comprises multiple alternatively spliced variants; however, their functional differences or redundancy remain unclear. In this study, we aimed to characterize mouse CaMKKß/2 splice variants (CaMKKß-3 and ß-3x). RT-PCR analyses revealed that mouse CaMKKß-1, consisting of 17 exons, was predominantly expressed in the brain; whereas, mouse CaMKKß-3 and ß-3x, lacking exon 16 and exons 14/16, respectively, were primarily expressed in peripheral tissues. At the protein level, the CaMKKß-3 or ß-3x variants showed high expression levels in mouse cerebrum and testes. This was consistent with the localization of CaMKKß-3/-3x in spermatids in seminiferous tubules, but not the localization of CaMKKß-1. We also observed the co-localization of CaMKKß-3/-3x with a target kinase, CaMKIV, in elongating spermatids. Biochemical characterization further revealed that CaMKKß-3 exhibited Ca2+/CaM-induced kinase activity similar to CaMKKß-1. Conversely, we noted that CaMKKß-3x impaired Ca2+/CaM-binding ability, but exhibited significantly weak autonomous activity (approximately 500-fold lower than CaMKKß-1 or ß-3) due to the absence of C-terminal of the catalytic domain and a putative residue (Ile478) responsible for the kinase autoinhibition. Nevertheless, CaMKKß-3x showed the ability to phosphorylate downstream kinases, including CaMKIα, CaMKIV, and AMPKα in transfected cells comparable to CaMKKß-1 and ß-3. Collectively, CaMKKß-3/-3x were identified as functionally active and could be bona fide CaMKIV-kinases in testes involved in the activation of the CaMKIV cascade in spermatids, resulting in the regulation of spermiogenesis.
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Texte intégral: 1 Collection: 01-internacional Base de données: MEDLINE Sujet principal: Spermatides / Calcium-Calmodulin-Dependent Protein Kinase Kinase Limites: Animals Langue: En Journal: Cell Calcium Année: 2024 Type de document: Article Pays d'affiliation: Japon Pays de publication: Pays-Bas

Texte intégral: 1 Collection: 01-internacional Base de données: MEDLINE Sujet principal: Spermatides / Calcium-Calmodulin-Dependent Protein Kinase Kinase Limites: Animals Langue: En Journal: Cell Calcium Année: 2024 Type de document: Article Pays d'affiliation: Japon Pays de publication: Pays-Bas