Epstein-Barr Virus BART9 miRNA Modulates LMP1 Levels and Affects Growth Rate of Nasal NK T Cell Lymphomas
Figure 4
Immunoblot and Q-RT-PCR analysis of LMP1 expression in SNK6 following inhibition of EBV-BART9 miRNA.
(A) SNK6 cells were transfected with anti-EBV-BART9 miRNA or Scramble control miRNA and cell lysates prepared 96 hours post-transfection. LMP1 protein expression was analyzed in immunoblots. When compared to cells transfected with control miRNA and normalized to β-actin loading control, quantification of immunoblots showed that BART9 inhibition reduced LMP1 protein levels by ∼50%. (B) SNK6 cells were transfected with control or anti-EBV-BART9 miRNA and samples collected every 24 hours in a time-course experiment. Cell lysates were prepared and immunoblot analysis carried out to determine LMP1 expression. Quantification of LMP1 levels using Image J as described above showed that LMP1 protein levels are reduced only at later time-point. (C) SNK6 cells were transfected with either anti-EBV-BART9 or control miRNA and cells collected 96 hours post-transfection. Total RNA was extracted and cDNA synthesized using iScript cDNA synthesis kit. Using LMP1 specific primers, Q-PCR was carried out and data analyzed using the ΔΔCt method. Data shown is the average ± SD from three independent experiments. (** represents p value of <0.005 in a paired t-test).