Fig. 4. RAC1 interacts with IMPDH2 and recruits GMPS.

a Cells were transduced with the indicated constructs and subjected to immunoprecipitation with the antibodies indicated on the top. The immunoprecipitated materials were probed in immunoblotting with the antibodies indicated on the left. Shown are representative images of at least two independent experiments. b Cells were fixed with ice-cold methanol and separated into cell bodies (CB) or cell protrusions (CP) fractions, followed by immunoblotting with the indicated antibodies. Shown are representative images of two independent experiments. c Proximity ligation assay was performed on sparsely plated cells with the antibodies indicated on the top. Shown are representative images of two independent experiments. d Schematic representation of IMPDH2 deletion mutants. Shown are Bateman domain consisting of two cystathionine-β-synthase sequences (CBS). e Indicated GST-tagged recombinant IMPDH2 mutants were incubated with recombinant 6xHis-tagged RAC1 (shown on the top), followed by immunoprecipitation with anti-GST tag antibodies and immunoblotting with anti-RAC1 antibodies. Shown are representative images of two independent experiments. f Recombinant full-length IMPDH2 and RAC1 proteins were cross-linked followed by mass spectroscopy as described in Methods. Shown are intra-proteins cross-linked peptides identified with highest confidence. Highlighted in red are cross-linked lysines. Shown are representative gel images of at least two independent experiments.