Figure 3

PCBP1 is a negative regulator of CD44 v6. (A) Different doses of PCBP1 expression vector as indicated were transfected into HepG2 cells. Cells were harvested 24 hours later and total RNA were extracted for semi-quantitative RT-PCR and Real-time PCR analysis (B) to determine the relative amounts of CD44 v6. (C) HepG2 cells were transfected with a negative control siRNA (N.C) or PCBP1 specific siRNA oligos for 48 hours and then total RNA were extracted. The mRNA level of v6 was examined by semi-quantitative RT-PCR and Real-time PCR analysis (D). (E) PCBP1 inhibits the up-regulation of CD44 v6 induced by Ras activation. HepG2 cells were transfected with a constitutively activated mutant H-Ras V12 construct along with PCBP1 expression vector just as indicated. 24 hours later, total RNA were extracted for semi-quantitative PCR and Real-time PCR analysis (F). (G-H) PCBP1 inhibits the upregulation of CD44 v6 induced by HGF. HepG2 cells were transfected with control vector or PCBP1 expression vector, and then 24 hours later the cells were stimulated with 20 ng/ml HGF for 8 hours. Total RNA was extracted and RT-PCR (G) or Real-time PCR (H) was performed to detect the level of CD44 v6. All the PCR bands were scanned for densitometry analysis with the value obtained from control cells set as 1. The values were normalized with those of GAPDH. Results represented mean ± SD of 3 independent experiments. The statistical difference between the samples was demonstrated as * p ≤ 0.05 or ** p ≤ 0.001.