ABSTRACT
Objective:To investigate the correlation of GNG4 with DNA damage repair and chemosensitivity of ovarian cancer cisplatin-resistant A2780/DDP cells.Methods:A2780/DDP cells were divided into 500 ng/ml cisplatin group (cDDP group), short hairpin RNA (shRNA)-GNG4 silencing GNG4 expression group (shRNA group), 500 ng/ml cisplatin and shRNA-GNG4 intervention group (shRNA+cDDP group), and non cisplatin and shRNA-GNG4 intervention group (blank control group). Western blot was used to detect the expressions of GNG4 and γH2AX proteins in each group; DNA damage in each group was detected by single cell gel electrophoresis. The focus formation of γH2AX gene at the injury site was detected by immunofluorescence. The ability of cell clone formation was detected by plate clone formation experiment.Results:Compared with the other three groups, the expression level of GNG4 protein in shRNA+cDDP group was the lowest, the expression level of γH2AX protein was the highest, and the differences were statistically significant (all P < 0.01). Single cell gel electrophoresis assay showed that the comet tail DNA% in blank control group, cDDP group, shRNA group and shRNA+cDDP group were (7.7±2.5)%, (12.3±3.6)%, (20.1±2.1)%, (38.6±2.8)%, respectively, and Olive trailing distance were 5.12±1.89, 8.23±2.97, 14.99±3.65, 22.43±3.17, respectively, the comet tail DNA% and Olive tail distance in shRNA+cDDP group were higher than those in the other three groups, and the differences were statistically significant (all P < 0.05). Immunofluorescence assay showed that the focus numbers of γH2AX in each cell of blank control group, cDDP group, shRNA group and shRNA+cDDP group were 4.2±0.7, 5.1±0.5, 26.8±3.3, 71.3±6.2, respectively, the shRNA+cDDP group was higher than the other three groups, and the differences were statistically significant (all P < 0.05). The clone formation rates of blank control group, cDDP group, shRNA group and shRNA+cDDP group were (78.27±5.01)%, (45.67±3.29)%, (26.20±5.76)%, (1.56±0.21)%, respectively, the shRNA+cDDP group was lower than the other three groups, and the differences were statistically significant (all P < 0.001). Conclusions:Down-regulation of GNG4 expression can increase the cisplatin sensitivity of ovarian cancer A2780/DDP cells, which may be achieved by inhibiting the DNA damage repair function induced by cisplatin.
ABSTRACT
Phosphatidylinositol 3-kinase (PI3K) is essential for both G protein-coupled receptor (GPCR)- and receptor tyrosine kinase (RTK)-mediated cancer cell migration. Here, we have shown that maximum migration is achieved by full activation of phosphatidylinositol 3,4,5-trisphosphate-dependent Rac exchanger 1 (P-Rex1) in the presence of Gbetagamma and PI3K signaling pathways. Lysophosphatidic acid (LPA)-induced migration was higher than that of epidermal growth factor (EGF)-induced migration; however, LPA-induced activation of Akt was lower than that stimulated by EGF. LPA-induced migration was partially blocked by either Gbetagamma or RTK inhibitor and completely blocked by both inhibitors. LPA-induced migration was synergistically increased in the presence of EGF and vice versa. In correlation with these results, sphingosine-1-phosphate (S1P)-induced migration was also synergistically induced in the presence of insulin-like growth factor-1 (IGF-1). Finally, silencing of P-Rex1 abolished the synergism in migration as well as in Rac activation. Moreover, synergistic activation of MMP-2 and cancer cell invasion was attenuated by silencing of P-Rex1. Given these results, we suggest that P-Rex1 requires both Gbetagamma and PI3K signaling pathways for synergistic activation of Rac, thereby inducing maximum cancer cell migration and invasion.
Subject(s)
Humans , Cell Line, Tumor , Cell Movement/drug effects , Enzyme Activation/drug effects , GTP-Binding Protein beta Subunits/metabolism , GTP-Binding Protein gamma Subunits/metabolism , Guanine Nucleotide Exchange Factors/metabolism , Lysophospholipids/pharmacology , Neoplasms/enzymology , Phosphatidylinositol 3-Kinases/metabolism , Proto-Oncogene Proteins c-akt/metabolism , Receptors, G-Protein-Coupled/metabolism , Signal TransductionABSTRACT
Objetivos: revisar o assunto proteína G e seus mecanismos de transdução celular, de forma abrangente e didática.Fonte de dados: foram revisados artigos específicos sobre o tema, disponíveis em periódicos eletrônicos e encontrados através das bases de dados LILACS, PubMed e SciELO.Síntese dos dados: a transdução de sinais é uma função fisiológica que intermedeia o estímulo externo e a resposta celular, sendo o passo de conversão intracelular do agonismo de várias substâncias. Os compostos proteicos envolvidos nessa atividade estão presentes em todos os sistemas do organismo; consequentemente, disfunções na sua estrutura culminam em estados patológicos diversos. A descrição da dinâmica da transdução, da estrutura e funções da proteína G e do seu papel em algumas doenças foram abordados nesta revisão.Conclusões: a revisão da literatura mostra que o tema proteína G não tem gerado muitos trabalhos experimentais.Entretanto, o estudo desse composto protéico evidencia sua grande importância na fisiologia, indicando que disfunções na sua estrutura resultam em vários estados patológicos.
Aims: To review, in a comprehensive and didactic way, the issue G protein and its mechanisms of cellular transduction.Source of data: Articles that address the specific issue, available online, and found through the databases LILACS, PubMed and SciELO, were reviewed.Summary of findings: Signal transduction is a physiological function that mediates the external stimulus and cellular response; it is the conversion step of agonism of several intracellular substances. The protein compounds involved in this activity are present in all body systems, thus dysfunction in its structure results in several pathological states.The description of the dynamics of transduction, structure and functions of G protein and its role in some diseases were addressed in this review.Conclusions: The literature review shows that the subject protein G has not generated many experimental studies.However, the study of this protein compound makes evident its great importance in physiology and indicates that dysfunctions in its structure result in various pathological conditions.