Type II cyclic guanosine monophosphate-dependent protein kinase inhibits Rac1 activation in gastric cancer cells

Author:              

Publisher: Spandidos Publications

E-ISSN: 1792-1082|10|1|502-508

ISSN: 1792-1074

Source: Oncology Letters, Vol.10, Iss.1, 2015-01, pp. : 502-508

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Abstract

Enhanced motility of cancer cells is a critical step in promoting tumor metastasis, which remains the major cause of gastric cancerassociated mortality. The small GTPase Rac1 is a key signaling component in the regulation of cell migration. Previous studies have demonstrated that Rac1 activity may be regulated by protein kinase G (PKG); however, the underlying mechanism is not yet clear. The current study aimed to investigate the effect of type II cyclic guanosine monophosphate (cGMP)dependent protein kinase (PKG II) on Rac1 activity. The human gastric cancer cell line AGS was infected with adenoviral constructs encoding PKG II to increase the expression of this enzyme, and treated with a cGMP analog (8pCPTcGMP) to induce its activation. A Transwell assay was employed to measure cell migration, and the activity of Rac1 was assessed using a pulldown assay. Immunoprecipitation was used to isolate the Rac1 protein. Phosphorylation of phosphatidylinositol 4,5 bisphosphate 3 kinase (PI3K) and its downstream effecter protein kinase B (Akt) are associated with lysophosphatidic acid (LPA)induced motility/migration of cancer cells. Extracellular signal regulated kinase (ERK) is the major signaling molecule of the Mitogen activated protein kinase (MAPK) mediated signaling pathway. ERK and its upstream activator MAPK kinase (MEK) are also involved in LPAinduced motility/migration of cancer cells. Phosphorylation of PI3K/Akt, MEK/ERK and enriched Rac1 were detected by western blotting. The results revealed that blocking the activation of Rac1 by ectopically expressing an inactive Rac1 mutant (T17N) impeded LPAinduced cell migration. Increased PKG II activity inhibited LPAinduced migration and LPAinduced activation of Rac1; however, it had no effect on the phosphorylation of Rac1. PKG II also inhibited the activation of PI3K/Akt and MEK/ERK mediated signaling, which is important for LPAinduced Rac1 activation. These results suggest that PKG II affects LPAstimulated migration of AGS cells by blocking Rac1 activation, via inhibition of PI3K/Akt and MEK/ERK mediated signaling.