•G protein receptor kinases (GRKs) regulate histamine H2R desensitization in human leukemic cells

FERNÁNDEZ N, MONCZOR F, TUBIO MR, BALDI A, SHAYO C, DAVIO C

Delphos, Grecia

Congreso; 35th Annual Congress of European Histamine Research Society. Delphos, Grecia; 2006

EHRS

Activation of H2R triggers adenylyl cyclase stimulation leading to cAMP accumulation and PKA activation. We have previously reported that GRKs participate in the regulation of H2R. Thus, in COS-7 cells cotransfected with H2R and GRKs we demonstrated that H2R rapid desensitization involves receptor phosphorylation by GRK2 and GRK3 (Mol. Pharmacol., 60:1049, 2001). Further in U937 promonocytic cell line endogenously expressing H2R as well as GRK2, GRK3 and GRK6, we showed the role of GRK2 in H2R desensitization (Mol. Pharmacol., 62:1506, 2002). In the present study we sought to establish the participation of GRK3 and GRK6 in H2R desensitization in U937 cells. For this purpose clones expressing low levels of GRK3 and GRK6 were generated by stable transfection with cDNA antisense for these kinases. Cyclic AMP was assayed following dose response curves as well as kinetic and desensitization experiments performed in the presence of a selective H2R agonist. Anti GRK3 clones showed a maximal response that resulted 25% to 47% lower than that of U937 cells. Further time course studies showed that these clones evoked reduced cAMP levels as compared to U937 cells and suffered a more rapid desensitization. Binding assays revealed that this behavior did not result from a reduced H2R number of sites in GRK3 clones. The response of anti-GRK6 clones was similar to that of U937 cells supporting that GRK6 is not involved in H2R desensitization. In view of the unexpected behavior shown by anti-GRK3 clones, GRK2 levels were determined by Western Blot. In anti-GRK6 clones GRK2 levels resulted similar to those of U937 cells but in anti-GRK3 clones they were increased by 50% and 70%. These results suggest that increased GKR2 levels may account for the H2R response in anti-GRK3 clones and further support the role of GRK2 in H2R desensitization. Present findings support that in U937 cells GRK6 is not involved in H2R desensitization and that GRK3 modulates GRK2 levels.