The epidermal growth factor receptor is central to the growth, differentiation and the mobility of normal and cancer cells. Notably, EGFR plays an important role in non-small cell lung carcinoma development. Two known...The epidermal growth factor receptor is central to the growth, differentiation and the mobility of normal and cancer cells. Notably, EGFR plays an important role in non-small cell lung carcinoma development. Two known nucleotide variants in EGFR promoter at position -216 (G > T) and -191 (C > A) are known to influence promoter activity. The transcription factor Sp1 (Specificity protein 1) binds with higher affinity the T allele at position -216 which results in 30% increased transcriptional activity of EGFR. Sequencing of EGFR promoter region and exon 1 in 18 patients with pulmonary carcinoma revealed that -216 G/T variant was associated with 58.82% of NSCLC patients especially those with squamous carcinoma, with predominance of homozygote (T/T) variants. Strikingly, the -191C/A polymorphism was detected in 11.11% of patients having a pulmonary carcinoma with the predominance of homozygote (C/C) variants. The distribution analysis of the four haplotypes (G-C, G-A, T-C and T-A) and the diplotype (G-C/T-C) of -216G/T and -191C/A polymorphisms, revealed a clear predominance of T-C haplotype (216T-191C) in squamous cell carcinoma. The G-C and G-A haplotypes have a lesser distribution while the T-A haplotype is non-existent. The incidence of (G-C/T-C) diplotype is more important than the G-A haplotype in all the studied cases. Our results are strongly correlated with the data of Caucasian population.展开更多
AIM: To study the detail mechanism of interaction between PKC and GRK2 and the effect of GRK2 on activity of PKC.METHODS: The cDNA of pleckstrin homology (PH) domain located in GRK2 residue 548 to 660 was amplified by...AIM: To study the detail mechanism of interaction between PKC and GRK2 and the effect of GRK2 on activity of PKC.METHODS: The cDNA of pleckstrin homology (PH) domain located in GRK2 residue 548 to 660 was amplified by PCR with the mRNA of human GRK2 (β1-adrenergic receptor kinase) as template isolated from human fresh placenta,the expression vector pGEX-PH inserted with the aboved cDNA sequence for GRK2 PH domain protein and the expression vectors for GST (glutathion-s-transferase) -GRK2PH domain fusion protein, BTK (Bruton′s tyrosine kinase)PH domain and GST protein were constructed. The expression of GRK2 in culture mammalian cells (6 cell lines:PC-3,MDCK, SGC7901, Jurkat cell etc.) was determined by SDS-PAGE and Co-immunoprecipitation. The binding of GRK2PH domain, GST-GRK2 PH domain fusion protein and BTK PH domain to PKC in Vitro were detected by SDS-PAGE and Western blot, upon prolonged stimulation of epinephrine,the binding of GRK2 to PKC was also detected by western blot and Co-immunoprecipitation.RESULTS: The binding of GRK2 PH domain to PKC in Vitro was confirmed by western blot, as were the binding upon prolonged stimulation of epinephrine and the binding of BTK PH domain to PKC. In the present study, GRK2 PH domain was associated with PKC and down-regulated PKC activity,but Btk PH domain up-regulated PKC activity as compared with GRK2 PH domain.CONCLUSION: GRK2 can bind with PKC and down-regulated PKC activity.展开更多
文摘The epidermal growth factor receptor is central to the growth, differentiation and the mobility of normal and cancer cells. Notably, EGFR plays an important role in non-small cell lung carcinoma development. Two known nucleotide variants in EGFR promoter at position -216 (G > T) and -191 (C > A) are known to influence promoter activity. The transcription factor Sp1 (Specificity protein 1) binds with higher affinity the T allele at position -216 which results in 30% increased transcriptional activity of EGFR. Sequencing of EGFR promoter region and exon 1 in 18 patients with pulmonary carcinoma revealed that -216 G/T variant was associated with 58.82% of NSCLC patients especially those with squamous carcinoma, with predominance of homozygote (T/T) variants. Strikingly, the -191C/A polymorphism was detected in 11.11% of patients having a pulmonary carcinoma with the predominance of homozygote (C/C) variants. The distribution analysis of the four haplotypes (G-C, G-A, T-C and T-A) and the diplotype (G-C/T-C) of -216G/T and -191C/A polymorphisms, revealed a clear predominance of T-C haplotype (216T-191C) in squamous cell carcinoma. The G-C and G-A haplotypes have a lesser distribution while the T-A haplotype is non-existent. The incidence of (G-C/T-C) diplotype is more important than the G-A haplotype in all the studied cases. Our results are strongly correlated with the data of Caucasian population.
文摘AIM: To study the detail mechanism of interaction between PKC and GRK2 and the effect of GRK2 on activity of PKC.METHODS: The cDNA of pleckstrin homology (PH) domain located in GRK2 residue 548 to 660 was amplified by PCR with the mRNA of human GRK2 (β1-adrenergic receptor kinase) as template isolated from human fresh placenta,the expression vector pGEX-PH inserted with the aboved cDNA sequence for GRK2 PH domain protein and the expression vectors for GST (glutathion-s-transferase) -GRK2PH domain fusion protein, BTK (Bruton′s tyrosine kinase)PH domain and GST protein were constructed. The expression of GRK2 in culture mammalian cells (6 cell lines:PC-3,MDCK, SGC7901, Jurkat cell etc.) was determined by SDS-PAGE and Co-immunoprecipitation. The binding of GRK2PH domain, GST-GRK2 PH domain fusion protein and BTK PH domain to PKC in Vitro were detected by SDS-PAGE and Western blot, upon prolonged stimulation of epinephrine,the binding of GRK2 to PKC was also detected by western blot and Co-immunoprecipitation.RESULTS: The binding of GRK2 PH domain to PKC in Vitro was confirmed by western blot, as were the binding upon prolonged stimulation of epinephrine and the binding of BTK PH domain to PKC. In the present study, GRK2 PH domain was associated with PKC and down-regulated PKC activity,but Btk PH domain up-regulated PKC activity as compared with GRK2 PH domain.CONCLUSION: GRK2 can bind with PKC and down-regulated PKC activity.
