Detection of a novel cell cycle-regulated kinase activity that associates with the amino terminus of the retinoblastoma protein in G2/M phases.
Recent genetic and functional evidence suggests that the amino terminus of the retinoblastoma (Rb) protein plays an important role in Rb-mediated growth suppression. To explore the mechanism(s) by which this portion of Rb may regulate cell growth, we have sought to characterize cellular proteins that associate with the Rb amino terminus using an in vitro protein-binding assay. Here we report that at least one such protein is a cell cycle-regulated Rb/histone H1 kinase (RbK) whose enzymatic and/or Rb association activity is most prevalent in G2/M phases of cells. In contrast to previously characterized cyclin-dependent and Rb-associated kinases, such as cdk1 (cdc2) and cdk2, G2/M RbK 1) is not depleted by incubation with p13suc-beads, 2) is not detected with antisera against several Rb-associated cyclins-cdks, and 3) associated with Rb via the Rb amino terminus, a region that is dispensable for interaction with other Rb-associated kinases. RbK is clearly distinct from previously characterized mitotic cdks since cyclin A-cdc2, cyclin A-cdk2, cyclin B-cdc2, and cyclin B-cdk2 did not associate with the Rb amino terminus. Coprecipitation experiments with Rb antisera confirmed the association of Rb with a RbK-like kinase in metaphase-arrested cells in vivo. Interestingly, G2/M RbK did not appreciably associate with an analogous portion of p107, a Rb-related protein. Taken together, these data indicate that the Rb amino terminus specifically associates with a novel cell cycle-regulated kinase in late cell cycle stages.
Sterner
J M
JM
Department of Molecular Cancer Biology, Duke University Medical Center, Durham, North Carolina 27710, USA.
Murata
Y
Y
Kim
H G
HG
Kennett
S B
SB
Templeton
D J
DJ
Horowitz
J M
JM
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D016195:G2 Phase; D006657:Histones / Q000378:metabolism; D008938:Mitosis; D008969:Molecular Sequence Data; D017346:Protein Serine-Threonine Kinases / Q000032:analysis*; D011817:Rabbits; D016160:Retinoblastoma Protein / Q000378:metabolism*
D006657:Histones; D016160:Retinoblastoma Protein; D017346:Protein Serine-Threonine Kinases