Stimulation of H1 receptor inhibits cell proliferation through activation of small G proteins in CHO-H1 cells

NOTCOVICH CINTIA; DIEZ FEDERICO; COPSEL SABRINA; KAZANIETZ MARCELO; DAVIO CARLOS; SHAYO CARINA

Estocolmo, Suecia

Congreso; XXXVI Annual Meeting of the European Histamine Research Society; 2008

EUROPEAN HISTAMINE RESEARCH SOCIETY

The main signaling pathway described for the histamine H1 receptor (H1R) includes activation of PLC via Gq. Some GPCRs coupled to Gq/11 activate small Rho GTPases, modulating processes such as proliferation, migration and apoptosis. As histamine (His) modulates cell proliferation through H1R, the aims of the present study were to find out whether a H1r and Rho GTPases association existed, to evaluate His effect on CHO-H1 cell proliferation and to determine the link between Rho GTPases and H1r-induced proliferation. The studies were performed in CHO cells stably transfected with H1R. Pull down assays precipitating the GTP-bound forms of the main Rho GTPases, showed that His and H1 agonists induced Rac1 and RhoA activation in a time and dose dependent manner, but not of Cdc42. The response was abolished by mepyramine (H1R antagonist) and by U73122 (PLC inhibitor) supporting that Rac 1 and RhoA activation was mediated by H1R coupled to PLC stimulation. Rho modulation in cells can be monitored by serum response factor activity with the SRE-luciferase reporter system. Stimulation of CHO-H1 with His led to a 3-fold increase in luciferase activity. The response was inhibited by the coexpression with C3 toxin, which inactivates Rho, strongly supporting that His stimulates downstream signals through RhoA activation. In addition, Rac dependent JNK activation was detected by western blot within 30 min of His stimulation, inhibited by â2-chimaerin (Rac-GAP). The stimulation of H1 also led to ERK1/2 activation which was not dependent on either Rho or Rac. [3H]-thymidine incorporation assays showed that His and the H1 agonist inhibited cell proliferation in a dose dependent fashion. Inhibition of RhoA and Rac by expressing either C3 toxin or â2-chimaerin, respectively revealed that Rac was partly involved in His inhibitory effect. Present findings support that Rho GTPases activation constitutes a new step in the H1R signaling pathway that leads to the inhibition of cell proliferation.