Figure 3
hnRNPK post-transcriptionally regulates p65BTK expression via IRES-dependent translation of exon 1b-containing mRNA. (a) Anti-hnRNPK antibodies immunoprecipitate a complex containing hnRNPK, eIF4G2 (top) and p65BTK mRNA (bottom) from HCT116p53KO lysates. (b) Fluorescence of HeLa cells transfected with a bicistronic vector encoding RFP under the control of CMV promoter and GFP not preceded by a regulatory region (first row) or under the control of p65BTK 5′UTR (second to fourth row) and left untreated (second row) or treated with Rapamycin 200 nM (third row) or Cymarin 100 nM (fourth row) for 36 h. DAPI was used to stain nuclei. (c) Time-dependent variation of p65BTK expression after treatment of colon cancer cells with 200 nM Rapamycin (left) and 200 nM Cymarin (right). Fold variation of p65BTK protein expression of each sample was calculated and normalized to actin by ImageJ program. (d) HeLa cells were transfected with the same bicistronic reporter as in (b) and luc-targeted siRNAs (second row) or hnRNPK-targeted siRNAs (third row). DAPI was used to stain nuclei.
