Phosphorylation of CD20 in cells from a hairy cell leukemia cell line. Evidence for involvement of calcium/calmodulin-dependent protein kinase II.
Hairy cell leukemia is an uncommon B cell lymphoproliferative disease of unknown etiology. We previously observed that CD20, a membrane protein involved in B cell activation, is hyperphosphorylated on hairy cells and that these cells have unusually high levels of intracellular free Ca2+. Therefore, we used a hairy cell line, HCLL-7876, to study the potential involvement of Ca(2+)-activated protein kinases in CD20 phosphorylation. Addition of the Ca2+ ionophore, ionomycin, increased CD20 phosphorylation both in activated B cells and in cells from the hairy cell line; addition of EGTA to either cell type decreased basal levels of CD20 phosphorylation. Ionomycin treatment of these cells resulted in increased kinase activity of cytosolic extracts toward syntide-2, a synthetic peptide substrate for calcium/calmodulin-dependent kinase II (CaM-KII), with kinetics similar to those of CD20 phosphorylation in the cell line. CD20 isolated from the cell line was a substrate for purified CaM-KII in vitro. Phosphopeptide maps of CD20 from untreated hairy cells or ionomycin-treated HCLL-7876 cells were similar to maps of CD20 that had been phosphorylated in vitro by CaM-KII. These results suggest that the unusually high levels of intracytoplasmic Ca2+ in hairy cells may enhance the phosphorylation of key surface proteins.
Genot
E M
EM
Unite INSERM 196, Institut Curie, Paris, France.
Meier
K E
KE
Licciardi
K A
KA
Ahn
N G
NG
Uittenbogaart
C H
CH
Wietzerbin
J
J
Clark
E A
EA
Valentine
M A
MA
D000595:Amino Acid Sequence; D015703:Antigens, CD / Q000378:metabolism*; D018951:Antigens, CD20; D000944:Antigens, Differentiation, B-Lymphocyte / Q000378:metabolism*; D002118:Calcium / Q000502:physiology; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D002460:Cell Line; D006801:Humans; D066298:In Vitro Techniques; D015759:Ionomycin / Q000494:pharmacology; D007943:Leukemia, Hairy Cell / Q000378:metabolism*; D008969:Molecular Sequence Data; D010449:Peptide Mapping; D010455:Peptides / Q000737:chemistry; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D011493:Protein Kinase C / Q000378:metabolism; D011494:Protein Kinases / Q000378:metabolism*
D015703:Antigens, CD; D018951:Antigens, CD20; D000944:Antigens, Differentiation, B-Lymphocyte; D010455:Peptides; D010750:Phosphoproteins; D015759:Ionomycin; D011494:Protein Kinases; D011493:Protein Kinase C; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D002118:Calcium