Phosphorylation of retinoblastoma-related protein by the cyclin D/cyclin-dependent kinase complex is activated at the G1/S-phase transition in tobacco.
In mammals, D-type cyclin-associated kinases mainly regulate the G1/S transition by phosphorylating the retinoblastoma (Rb) protein. We previously demonstrated that in tobacco, cyclin D (Nicta; CycD3;3) is complexed with the PSTAIRE-containing cyclin-dependent kinase (CDKA) from tobacco. Here, we show that Nicta; CycD3;3-associated kinases phosphorylate both the tobacco Rb-related protein (NtRb1) and histone H1. Although NtRb1 kinase activity was detected only during the middle G1- to early S-phase, histone H1 kinase activity was observed as two peaks in G1- to S-phase and G2/M- to M-phase. Importantly, we show that the proportion of cells in the G1-phase was reduced in transgenic Bright Yellow-2 cells overexpressing Nicta; CycD3;3-GFP. Mutational analyses revealed that phosphorylation of Thr-191 in Nicta; CycD3;3 possibly is required for both full kinase activity and localization predominantly to the nucleus. These data suggest that Nicta; CycD3;3 acts as a rate-limiting regulator in the G1/S transition by forming active complexes with CDKA or its related kinases to phosphorylate Rb-related protein and potentially plays a novel role during G2/M and mitosis.
Nakagami
Hirofumi
H
Graduate School of Biological Sciences, Nara Institute of Science and Technology, Takayama 8916-5, Ikoma, Nara 630-0101, Japan.
Kawamura
Kazue
K
Sugisaka
Keiko
K
Sekine
Masami
M
Shinmyo
Atsuhiko
A
D000595:Amino Acid Sequence; D000818:Animals; D018797:Cell Cycle Proteins / Q000378:metabolism; D002460:Cell Line; D002478:Cells, Cultured; D056741:Cyclin D; D018844:Cyclin-Dependent Kinases / Q000235:genetics / Q000378:metabolism*; D016213:Cyclins / Q000378:metabolism; D003593:Cytoplasm / Q000235:genetics / Q000378:metabolism; D016193:G1 Phase / Q000235:genetics / Q000502:physiology*; D016195:G2 Phase / Q000235:genetics / Q000502:physiology; D018506:Gene Expression Regulation, Plant; D005982:Glutathione Transferase / Q000235:genetics / Q000378:metabolism; D006657:Histones / Q000378:metabolism; D007313:Insecta / Q000166:cytology; D008938:Mitosis / Q000235:genetics / Q000502:physiology; D008969:Molecular Sequence Data; D009154:Mutation; D010766:Phosphorylation; D010940:Plant Proteins / Q000378:metabolism; D030821:Plants, Genetically Modified; D011233:Precipitin Tests; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D016160:Retinoblastoma Protein / Q000235:genetics / Q000378:metabolism*; D016196:S Phase / Q000235:genetics / Q000502:physiology*; D017386:Sequence Homology, Amino Acid; D013912:Threonine / Q000235:genetics / Q000378:metabolism; D014026:Nicotiana / Q000166:cytology / Q000235:genetics / Q000378:metabolism*
D018797:Cell Cycle Proteins; D056741:Cyclin D; D016213:Cyclins; D006657:Histones; D010940:Plant Proteins; C107938:RRB1 protein, Zea mays; D011993:Recombinant Fusion Proteins; D016160:Retinoblastoma Protein; D013912:Threonine; D005982:Glutathione Transferase; D018844:Cyclin-Dependent Kinases