Figure 1: The A-Rafshort isoform expression is induced after hnRNP H knockdown.
(A) a-rafshort, a-rafWT/short, and hnRNP H mRNA expression in HeLa cells was analyzed by RT-PCR after transfection with control or hnRNP H-specific siRNA (siRNA#1). Shown is a representative result from two independent experiments. (B) A-Rafshort, A-RafWT, and hnRNP H protein expression was analysed by immunoblotting after transfection with control or hnRNP H-specific siRNA (siRNA#1). As a positive control for A-Rafshort, a lysate containing transfected recombinant HA-tagged A-Rafshort was used. Shown are the representative results from two independent experiments. (C) Pre-adsorption of polyclonal A-Raf antibodies with peptides used for immunisation abrogated binding of A-Raf but not HA-specific antibody. Increasing concentrations of HA-tagged A-Rafshort was expressed in HeLa and analysed by immunoblotting using HA- or A-Raf N-terminus specific antibodies. The A-Raf N-terminus specific antibody but not the HA-antibody signal was blocked by adding the A-Raf N-terminal peptide as competitor, showing that the detection of the A-Rafshort isoform to be specific. Shown are the representative results from two independent experiments.