title: Expression analyses of splice variants of zebrafish cyclin-dependent kinase-like 5 and its substrate, amphiphysin 1.
abstract: Mammalian cyclin-dependent kinase-like 5 (CDKL5) is a Ser/Thr protein kinase mainly expressed in the central nervous system and believed to be involved in neuronal functions. However, the functions of CDKL5 in fishes have not been investigated. Therefore, in this study, we cloned and characterized zebrafish CDKL5 (zCDKL5) and its substrate, amphiphysin 1 (zAmph1). Two alternative splice variants of zCDKL5, zCDKL5-Long (zCDKL5-L) and zCDKL5-Short (zCDKL5-S), and three splice variants of zAmph1, zAmph1a, zAmph1b and zAmph1c, were cloned from a zebrafish cDNA library. Using zAmph1a point mutants, we identified Ser-285 and Ser-293 as phosphorylation sites of zAmph1a by CDKL5. Transiently expressed zCDKL5-L and zCDKL5-S colocalized with zAmph1a in the cytoplasm of 293T cells. RT-PCR analysis revealed that zCDKL5-L was first observed 12hours post-fertilization (hpf) and increased thereafter, while zCDKL5-S appeared just after fertilization. zAmph1a was detected in all embryogenic stages and zAmph1b appeared from 12hpf, but the expression of zAmph1c was not observed in our experiments. In adult fish, zCDKL5-L was mainly expressed in the brain, but zCDKL5-S showed ubiquitous expression. zAmph1a was observed most abundantly in the eyes, whereas zAmph1b was predominantly expressed in the brain. zAmph1c was scarcely detected. These results suggest that phosphorylation of Amph1 by CDKL5 may be a common feature throughout animal species.
authors.affiliation: Department of Life Sciences, Faculty of Agriculture, Kagawa University, Kagawa 761-0795, Japan.
authors.forename: Syouichi
authors.initials: S
authors.lastname: Katayama
authors.affiliation: Department of Life Sciences, Faculty of Agriculture, Kagawa University, Kagawa 761-0795, Japan; Biomedical Research Institute, National Institute of Advanced Industrial Science and Technology (AIST), Tsukuba, Ibaraki 305-8566, Japan.
authors.forename: Yukako
authors.initials: Y
authors.lastname: Senga
authors.affiliation: Department of Life Sciences, Faculty of Agriculture, Kagawa University, Kagawa 761-0795, Japan.
authors.forename: Ami
authors.initials: A
authors.lastname: Oi
authors.affiliation: Department of Life Sciences, Faculty of Agriculture, Kagawa University, Kagawa 761-0795, Japan.
authors.forename: Yosuke
authors.initials: Y
authors.lastname: Miki
authors.affiliation: Department of Life Sciences, Faculty of Agriculture, Kagawa University, Kagawa 761-0795, Japan.
authors.forename: Yasunori
authors.initials: Y
authors.lastname: Sugiyama
authors.affiliation: Department of Life Sciences, Faculty of Agriculture, Kagawa University, Kagawa 761-0795, Japan.
authors.forename: Noriyuki
authors.initials: N
authors.lastname: Sueyoshi
authors.affiliation: Department of Life Sciences, Faculty of Agriculture, Kagawa University, Kagawa 761-0795, Japan. Electronic address: kamesita@ag.kagawa-u.ac.jp.
authors.forename: Isamu
authors.initials: I
authors.lastname: Kameshita
mesh_terms: D017398:Alternative Splicing; D000595:Amino Acid Sequence; D000818:Animals; D001921:Brain / Q000378:metabolism; D003001:Cloning, Molecular; D003593:Cytoplasm / Q000378:metabolism; D004625:Embryo, Nonmammalian; D005786:Gene Expression Regulation; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins / Q000235:genetics* / Q000378:metabolism; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism; D015027:Zebrafish / Q000196:embryology / Q000235:genetics*; D029961:Zebrafish Proteins / Q000235:genetics* / Q000378:metabolism
chemical_list: D009419:Nerve Tissue Proteins; D029961:Zebrafish Proteins; C075637:amphiphysin; D017346:Protein Serine-Threonine Kinases
keywords: Amphiphysin 1; Cyclin-dependent kinase-like 5; Expression analysis; Protein kinase; Splice variants; Zebrafish