Cell biological studies with monoclonal and polyclonal antibodies against human casein kinase II subunit beta demonstrate participation of the kinase in mitogenic signaling.
Casein kinase II (CKII) is a highly conserved ubiquitous serine/threonine kinase composed of two catalytically active (alpha and/or alpha') and two regulatory (beta) subunits. It has been suspected that, among numerous other cellular functions, CKII might play a role in the control of mitogenic signaling. To test for such a role and its mechanism in intact cells, monoclonal antibodies (mAbs) were generated against CKII beta using a recombinant protein containing amino acids 20-200 of human CKII beta. The CKII beta-specific mAb with the highest reactivity, mAb IVG6 (classified as IgG1 with kappa light chains), was purified to homogeneity. It recognized a CKII beta epitope comprising the amino acids 140-156, a basic and highly conserved region. In addition, polyclonal antibodies (pAbs) were raised and made monospecific by affinity purification. pAbs-mediated quantitative immunofluorescence microscopy of human IMR-90 fibroblasts and/or Western blots of cell fractions revealed (i) CKII beta was present in exponentially growing cells at a 2-3-fold higher level than in quiescent cells, (ii) CKII beta was localized predominantly in the nucleus of cells (3-15-fold cytoplasmic level depending on cellular state and assay used), and (iii) the nuclear/cytoplasmic ratio of CKII beta was higher by a factor of 2 in exponentially growing cells. Consequently, mitogenic stimulation of quiescent cells by fetal calf serum doubled the nuclear/cytoplasmic ratio of CKII beta. The increase occurred within the 1st h of stimulation. The translocation of CKII beta into the nucleus was inhibited when mAb IVG6 was injected into the cytoplasm at the time of mitogenic stimulation. This microinjection also significantly inhibited the cell proliferation. The data imply that cytoplasmic CKII participates in the transmission of mitogenic signals by translocation into the nucleus.
Lorenz
P
P
Biochemical Cell Physiology, German Cancer Research Center, Heidelberg.
Pepperkok
R
R
Ansorge
W
W
Pyerin
W
W
D000911:Antibodies, Monoclonal / Q000276:immunology; D047390:Casein Kinase II; D002451:Cell Compartmentation; D002453:Cell Cycle; D002467:Cell Nucleus / Q000201:enzymology; D002478:Cells, Cultured; D003593:Cytoplasm / Q000201:enzymology; D005455:Fluorescent Antibody Technique; D006801:Humans; D007158:Immunologic Techniques; D066298:In Vitro Techniques; D008845:Microinjections; D008938:Mitosis; D017346:Protein Serine-Threonine Kinases / Q000502:physiology*
D000911:Antibodies, Monoclonal; D047390:Casein Kinase II; D017346:Protein Serine-Threonine Kinases