Evidence for unique calmodulin-dependent nuclear factor-kappaB regulation in WEHI-231 B cells.
Immature B cells express constitutive nuclear factor-kappaB (NF-kappaB) activity and inhibition of this activity is associated with the induction of apoptotic cell death. Previous studies have implicated a calcium-dependent proteolysis of the NF-kappaB inhibitory protein IkappaBalpha as critical in the maintenance of constitutive NF-kappaB activity in these cells. We tested whether modulation of diverse calcium-dependent processes affects the maintenance of constitutive NF-kappaB activity in the WEHI-231 immature B cell line. Calmodulin inhibitors, but not calcineurin inhibition, blocked both IkappaBalpha turnover and the maintenance of constitutive NF-kappaB activity. Inhibition of NF-kappaB DNA binding activity by the calmodulin antagonist W13 also resulted in a loss of the expression of the NF-kappaB target gene, IkappaBalpha. However, prolonged inhibition of NF-kappaB activity for up to 8 h did not lead to apoptotic induction in the WEHI-231 cells. Moreover, removal of calmodulin inhibitors resulted in the reappearance of constitutive NF-kappaB activity and the renewed expression of the IkappaBalpha gene. Thus, calmodulin activity is a requirement for the continual turnover of IkappaBalpha and the maintenance of constitutive NF-kappaB function in WEHI-231 cells. In addition, our findings suggest that inhibition of NF-kappaB activity does not lead to the immediate onset of apoptosis, indicating that prolonged inhibition of NF-kappaB-dependent gene expression is required to cause apoptosis of WEHI-231 B cells.
Shumway
Stuart D
SD
Program in Cellular and Molecular Biology, Department of Pharmacology, University of Wisconsin, Madison, Wisconsin 53706, USA.
Berchtold
Craig M
CM
Gould
Michael N
MN
Miyamoto
Shigeki
S
D000818:Animals; D017209:Apoptosis; D001402:B-Lymphocytes / Q000187:drug effects / Q000378:metabolism*; D019703:Calcineurin / Q000378:metabolism; D002147:Calmodulin / Q000037:antagonists & inhibitors / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D005260:Female; D015870:Gene Expression / Q000187:drug effects; D020672:I-kappa B Proteins; D051379:Mice; D008810:Mice, Inbred C57BL; D000072000:NF-KappaB Inhibitor alpha; D016328:NF-kappa B / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D013449:Sulfonamides / Q000494:pharmacology; D014407:Tumor Cells, Cultured
D002147:Calmodulin; D004268:DNA-Binding Proteins; D020672:I-kappa B Proteins; D016328:NF-kappa B; C000606205:Nfkbia protein, mouse; D013449:Sulfonamides; D000072000:NF-KappaB Inhibitor alpha; C033772:N-(4-aminobutyl)-5-chloro-2-naphthalenesulfonamide; D019703:Calcineurin