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 β subunit (CKIIβ), but not with the catalytic a subunit (CKIIα) of the tetrameric CKII holoenzyme. The binding of p47^phox to CKIIβ 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 CKIIβ. 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 CKIIβ and can modulate the catalytic activity of CKII holoenzyme.