Regulation of protein kinase CKII by direct interaction with the C-terminal region of p47(phox).
Protein kinase CKII is a Ser/Thr kinase which is involved in many proliferation-related processes in the cell. p47(phox) is a component of the leukocyte NADPH oxidase, which is an important element of host defense against microbial infection. In this study, we demonstrate that a truncated form of the p47(phox) lacking its N-terminal region (p47(phox)/SH3-C), but not a truncated form of the p47(phox) lacking its C-terminal region (p47(phox)/N-SH3), undergoes better phosphorylation by CKII in the presence of arachidonic acid. The yeast two-hybrid test and the glutathione S-transferase (GST) pull-down assay showed that p47(phox) interacts specifically with the regulatory beta subunit (CKIIbeta), but not with the catalytic alpha subunit (CKIIalpha) of the tetrameric CKII holoenzyme. The binding of p47(phox) to CKIIbeta requires the C-terminal region of p47(phox) and is completely abolished by addition of spermine, indicating that a highly basic region in the C-terminal region of p47(phox) contributes to binding to CKIIbeta. In addition, p47(phox) stimulates the catalytic activity of CKII holoenzyme; this stimulation also requires the C-terminal region of p47(phox). Coimmunoprecipitation experiments showed that CKII holoenzyme interacts with p47(phox) in human neutrophils. Taken together, the present data indicate that the C-terminal region of p47(phox) plays a significant role in the arachidonic acid-dependent phosphorylation of p47(phox) by CKII and that the same region of p47(phox) associates directly with CKIIbeta and can modulate the catalytic activity of CKII holoenzyme.
Kim
Y S
YS
Department of Biochemistry, College of Natural Sciences, Kyungpook National University, Taegu 702-701, Korea.
Lee
J H
JH
Park
J W
JW
Bae
Y S
YS
D000595:Amino Acid Sequence; D001483:Base Sequence; D047390:Casein Kinase II; D002384:Catalysis; D017931:DNA Primers; D019255:NADPH Oxidases; D010750:Phosphoproteins / Q000737:chemistry / Q000378:metabolism*; D010766:Phosphorylation; D011485:Protein Binding; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D013096:Spermine / Q000494:pharmacology
D017931:DNA Primers; D010750:Phosphoproteins; D013096:Spermine; D019255:NADPH Oxidases; C060505:neutrophil cytosolic factor 1; D047390:Casein Kinase II; D017346:Protein Serine-Threonine Kinases