Cloning and characterization of a novel Ca2+/calmodulin-dependent protein kinase I homologue in Xenopus laevis.
In order to investigate protein kinases expressed in the different developmental stages of Xenopus laevis, recently developed expression cloning was carried out. When two different expression libraries, Xenopus oocyte and Xenopus head (embryonic stage 28/30) cDNA libraries, were screened by kinase-specific monoclonal antibodies, cDNA clones for various known and novel protein serine/threonine kinases (Ser/Thr kinases) were isolated. In addition to well-characterized Ser/Thr kinases, one cDNA clone for a putative kinase was isolated from the Xenopus head library. The sequence of the open reading frame of the cDNA encoded a protein of 337 amino acid residues with a predicted molecular weight of 38,404. Since the deduced animo acid sequence of this protein was 75% identical to that of rat Ca(2+)/calmodulin-dependent protein kinase I (CaMKI), it was designated as CaMKIx. Although recombinant CaMKIx expressed in Escherichia coli showed no protein kinase activity against syntide-2, a synthetic peptide substrate, it was activated when phosphorylated by mouse Ca(2+)/calmodulin-dependent protein kinase kinase alpha (CaMKKalpha). Activated CaMKIx significantly phosphorylated various proteins including synapsin I, histones, and myelin basic protein. CaMKIx could not be detected in the early stages of embryogenesis, but was detected in late embryos of stages 37/38 and thereafter when examined by Western blotting using a specific antibody. This kinase was found to be highly expressed in adult brain and heart, and an upstream kinase that could activate CaMKIx was detected in these tissues. These results suggest that CaMKIx plays some critical role in the late stages of embryogenesis of Xenopus laevis.
Kinoshita
Shun
S
Department of Life Sciences, Faculty of Agriculture, Kagawa University, Ikenobe 2393, Miki-cho, Kagawa 761-0795.
Sueyoshi
Noriyuki
N
Shoju
Hiroyuki
H
Suetake
Isao
I
Nakamura
Masahiko
M
Tajima
Shoji
S
Kameshita
Isamu
I
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry; D001483:Base Sequence; D019921:Biotinylation; D015153:Blotting, Western; D054737:Calcium-Calmodulin-Dependent Protein Kinase Kinase; D054729:Calcium-Calmodulin-Dependent Protein Kinase Type 1; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics*; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000378:metabolism; D015723:Gene Library; D008969:Molecular Sequence Data; D016366:Open Reading Frames; D010455:Peptides / Q000737:chemistry; D010766:Phosphorylation; D010957:Plasmids / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000737:chemistry; D051381:Rats; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity; D013997:Time Factors; D029867:Xenopus Proteins / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics*; D014982:Xenopus laevis
D000911:Antibodies, Monoclonal; D018076:DNA, Complementary; D010455:Peptides; D011994:Recombinant Proteins; D029867:Xenopus Proteins; D017346:Protein Serine-Threonine Kinases; C496114:CaMKIx protein, Xenopus laevis; D054737:Calcium-Calmodulin-Dependent Protein Kinase Kinase; D054729:Calcium-Calmodulin-Dependent Protein Kinase Type 1; D017871:Calcium-Calmodulin-Dependent Protein Kinases; C517699:Camkk1 protein, mouse; C517701:Camkk1 protein, rat