Extended Data Fig. 3: N-terminal disordered sequence of ARAF drives PM clustering.
From: Plasma membrane-associated ARAF condensates fuel RAS-related cancer drug resistance
a, SKBR3 cells co-transfected with indicated plasmids were subjected for imaging. Scale bar, 5 μm. b, Upper panel, schematic representation of engineered GFP fusion proteins. Lower panel, images of SKBR3 cells transfected with the indicated plasmids. Scale bar, 5 μm. c, SKBR3 cells transfected with indicated ARAF 255 mutants were subjected for imaging (left), percentages of cells with PM signals were quantified, n = 3 (right). Scale bars, 5 μm. d, Circular dichroism spectroscopy analysis of the biophysical properties of chemically synthesized the N-terminal region of ARAF and a series of its mutants. e, NIH3T3 cells expressing WT ARAF together with increasing amounts of ARAF P/F mutant were subjected for immunoprecipitation. f, NIH3T3 cells expressing indicated constructs were subjected to subcellular fractionation, PM fractions were resolved by SDD-AGE (top) and SDS-PAGE (bottom). Immunoblots are representative of three independent experiments in e, f. Error bars denote the mean ± SD (biological replicates), and statistical analyses were performed using unpaired two-tailed Student’s t-test in c.
