Activation of CDK-activating kinase is dependent on interaction with H-type cyclins in plants.
cDNAs encoding cyclin H homologs were isolated from poplar (Populus tremula X tremuloides) and rice (Oryza sativa) plants, and were designated Pt;cycH;1 and Os;cycH;1, respectively. The deduced amino-acid sequences showed 40-60% similarity to human cyclin H and Schizosaccharomyces pombe Mcs2, with higher similarity in the cyclin box region. While Pt;cycH;1 and Os;cycH;1 were expressed in all tissues examined, the transcripts accumulated abundantly in dividing cells. Expression of Os;cycH;1 was abundant in the S-phase in partially synchronized suspension cells, and was induced by submergence in internodes of deepwater rice. A yeast two-hybrid assay demonstrated that both Pt;CycH;1 and Os;CycH;1 were able to interact with rice R2 kinase, which is structurally and functionally similar to cyclin-dependent kinase (CDK)-activating kinase (CAK) of vertebrates. Moreover, an in vitro pull-down assay showed that Os;CycH;1 specifically bound to R2 but not to other rice CDKs. When R2 was expressed in budding yeast CAK mutant, the suppression activity in terms of temperature-sensitivity was enhanced by co-expression with Os;cycH;1. Furthermore, in vitro kinase assay indicated that the kinase activities of R2 on CDKs and the carboxy-terminal domain (CTD) of the largest subunit of RNA polymerase II were markedly elevated by binding to Os;CycH;1. Our results suggest that cyclin H is a regulatory subunit of CAK, which positively controls CDK- and CTD-kinase activities in plant cells.
Yamaguchi
M
M
Institute of Molecular and Cellular Biosciences, The University of Tokyo, Yayoi 1-1-1, Bunkyo-ku, Tokyo 113-0032, Japan.
Fabian
T
T
Sauter
M
M
Bhalerao
R P
RP
Schrader
J
J
Sandberg
G
G
Umeda
M
M
Uchimiya
H
H
D000595:Amino Acid Sequence; D000818:Animals; D042846:CDC2-CDC28 Kinases; D002453:Cell Cycle; D003001:Cloning, Molecular; D051357:Cyclin-Dependent Kinase 2; D018844:Cyclin-Dependent Kinases / Q000235:genetics / Q000378:metabolism; D016213:Cyclins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D004789:Enzyme Activation; D006801:Humans; D008969:Molecular Sequence Data; D012275:Oryza / Q000166:cytology / Q000235:genetics / Q000378:metabolism*; D010802:Phylogeny; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D012441:Saccharomyces cerevisiae; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D014197:Trees / Q000166:cytology / Q000235:genetics / Q000378:metabolism*; D000095043:Cyclin-Dependent Kinase-Activating Kinase
D016213:Cyclins; D011994:Recombinant Proteins; D017346:Protein Serine-Threonine Kinases; D042846:CDC2-CDC28 Kinases; C495894:CDK2 protein, human; D051357:Cyclin-Dependent Kinase 2; D018844:Cyclin-Dependent Kinases; D000095043:Cyclin-Dependent Kinase-Activating Kinase