Figure 2
From: Comparative RNA sequencing reveals that HPV16 E6 abrogates the effect of E6*I on ROS metabolism
Expression profiles of HPV16 E6 and E6*I in stable cell lines. (A) Western blotting analysis showing p53, E6 and β-actin protein levels in U-2 OS parental cell line (U-CTL#par) and in stably transfected cells with pXJ40 empty vector (U-CTL #1 and #3) or pXJ40-E6All vector (U-E6All #1, #6 and #7) or pXJ40-E6*I vector (U-E6*I #1, #6 and #14). β-actin is used as loading control. Full-length blots are presented in Figure S4. (B) Western blotting analysis comparing protein levels of E6 and p53 between U-2 OS cellular clones (U-CTL#par, U-E6All #1, #6 and #7) and naturally HPV16-infected cells (W12_20861, W12_20862, W12_20863, SiHa and Ca Ski cell lines). β-actin is used as loading control. (C) RT-PCR analysis of alternatively spliced HPV16 E6 transcripts in U-E6All and U-E6*I stably transfected cells. As expected no E6 transcripts were observed in U-2 OS parental cell line (U-CTL#par) and in U-CTL #1 and #3 stably transfected cells. The detection of the vector in stably transfected cells was done by pXJ40 vector specific primers and RPLP0 was used as loading control. Full-length gels are presented in Figure S5. (D) RT-PCR analysis of alternatively spliced E6 transcripts in HPV16 + cell lines SCC090, SiHa, Ca Ski, in stable cell lines U-E6All #1, #6 and #7 and in HPV16 + HNSCC tumor samples. RPLP0 was used as loading control. Note that a band can be observed at 400 bp and could correspond to heteroduplexes between the different amplicons generated during PCR reaction. Full-length gels are presented in Figure S6. (E) RT-PCR showing alternatively spliced E6 transcripts in naturally HPV16-infected cells (W12_20861, W12_20862, W12_20863).
