Cyclic AMP‐dependent protein kinase (cAK) in human B cells: co‐localization of type I cAK (RIα2C2) with the antigen receptor during anti‐immunoglobulin‐induced B cell activation

Abstract

Cyclic AMP (cAMP) inhibits antigen-stimulated B cell proliferation through activation of cAMP-dependent protein kinases (cAK). We have examined the molecular composition and cellular localization of cAK in human B cells. We find that human B cells contain substantial amounts of mRNA for RIα, RIIα, Cα and Cβ, barely detectable levels of RIβ mRNA, and no detectable RIIβ or Cγ mRNA. At the protein level, using Western blotting and subunit-specific antibodies against the different R subunits, we find RIα and RIIα, but no RIβ or RIIβ. The presence of catalytic subunits was demonstrated using a nonselective anti-C antiserum. By photoaffinity labeling of R subunits with 8-azido-[³²P]cAMP, followed by immunoprecipitation with subunit-specific antibodies, we were also able to demonstrate low levels of RIβ. Immunofluorescence staining of RIα and RIIα demonstrates a rather homogeneous intracellular (but extranuclear) distribution of RIα, whereas the RIIα subunits of cAK are localized to distinct perinuclear structures, previously identified as centrosomes in other cell types. Upon anti-Ig-mediated capping of B cells, RIα subunits redistribute to the cap, co-localizing with the antigen-receptors, whereas the intracellular localization of RIIα subunits remains unchanged.