Figure 3:

HnRNP K translocated into SGs with clotrimazole treatment. (A) Cells were incubated in serum-free medium in presence of 20 μM clotrimazole for 30 min. After cell fixation with 4% PFA solution and methanol, cells were stained using various antibodies, as described in Materials and Methods (Gray/Green, hnRNP K; Blue, EIF3α). Scale bar, 10 μm. (B) Cytosol accumulation of hnRNP K. U-2OS cells were treated with clotrimazole for 2 h. Western blot analysis using anti-Lamin A/C and anti-β-Tubulin was performed and the successful subcellular fractionation for nuclear and cytoplasmic fractions was confirmed, respectively. Same concentration of proteins were used for the following Western blot analysis to measure the protein expression levels of hnRNP K in cytoplasmic vs nuclear fractions. ImageJ software was used to quantify the band intensities to determine the ratio of nuclear vs cytoplasmic hnRNP K (right).