J Exp Med. 2002 Jun 17;195(12):1647-52.

Protein kinase C beta controls nuclear factor kappaB activation in B cells through selective regulation of the IkappaB kinase alpha.

Saijo K, Mecklenbräuker I, Santana A, Leitger M, Schmedt C, Tarakhovsky A.

Source

Laboratory of Lymphocyte Signaling, Rockefeller University, New York, NY 10021, USA. saijok@mail.rockefeller.edu

Abstract

Activation of the nuclear factor (NF)-kappaB transcription complex by signals derived from the surface expressed B cell antigen receptor controls B cell development, survival, and antigenic responses. Activation of NF-kappaB is critically dependent on serine phosphorylation of the IkappaB protein by the multi-component IkappaB kinase (IKK) containing two catalytic subunits (IKKalpha and IKKbeta) and one regulatory subunit (IKKgamma). Using mice deficient for protein kinase C beta (PKCbeta) we show an essential role of PKCbeta in the phosphorylation of IKKalpha and the subsequent activation of NF-kappaB in B cells. Defective IKKalpha phosphorylation correlates with impaired B cell antigen receptor-mediated induction of the pro-survival protein Bcl-xL. Lack of IKKalpha phosphorylation and defective NF-kappaB induction in the absence of PKCbeta explains the similarity in immunodeficiencies caused by PKCbeta or IKKalpha ablation in B cells. Furthermore, the well established functional cooperation between the protein tyrosine kinase Bruton's tyrosine kinase (Btk), which regulates the activity of NF-kappaB and PKCbeta, suggests PKCbeta as a likely serine/threonine kinase component of the Btk-dependent NF-kappaB activating signal transduction chain downstream of the BCR.

PMID:: 12070292; [PubMed - indexed for MEDLINE]
PMCID:: PMC2193563

Free PMC Article

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Figure 4.

Impaired activation and NF-κB signaling in PKCβ^−/− B cells after BCR cross-linking. Absence of IKKα activation and shortened IKKβ activation in PKCβ^−/− B cells. Splenic B cells of PKCβ^−/− and PKCβ^+/+ mice were stimulated with 20 μg/ml of anti-IgM antibody (A, top panel) or 10 μg/ml anti-CD40 (B, top panel) for the indicated time (min). Western blot analysis of cytoplasmic extracts were done simultaneously with antibodies specific for phospho-Ser180 and phospho-Ser181 of IKKα and IKKβ, respectively (top panel). The membrane was stripped and reprobed with antibodies recognizing nonphosphorylated IKKα and IKKβ. For quantification, band intensities of the phosphorylation site–specific blots were first normalized to the respective signal of unphosphorylated protein and then calculated as fold-change relative to unstimulated PKCβ^+/+, which was set to 1.0. Inefficient degradation and phosphorylation of IκBα after IgM cross-linking on PKCβ^−/− B cells (A, middle panel) and no alteration in IkBa phosphorylation and degradation after CD40 stimulation (B, middle panel). Splenic B cells were treated as in A. IκBα expression (top) and Ser32 phosphorylation (middle) were analyzed by Western blotting. Equal protein loading was controlled by anti-actin Western blotting (bottom). Quantification was done as in Fig. 2. Reduced DNA binding activity of NF-κB in PKCβ^−/− B cells after BCR cross-linking (A, bottom panel) and no alteration after CD40 stimulation (B, bottom panel). EMSA was performed with nuclear extracts of splenic B cells treated as in above. The specificity of NF-κB shift was confirmed by supershift with antibodies for p50, RelA, and c-Rel (Ab) and by using mutant oligonucleotides (m). For quantification, signal intensities were first normalized to the respective signal of an Oct-1 EMSA done with the same nuclear lysates and then calculated as fold-change relative to unstimulated PKCβ^+/+, which was set to 1.0.

Protein Kinase C ? Controls Nuclear Factor ?B Activation in B Cells Through Selective Regulation of the I?B Kinase ?

J Exp Med. 2002 June 17;195(12):1647-1652.