a. Splice variants of IRGM. G1-G5, GTPase motifs. The S47N mutation is indicated in blue. b. HeLa cells were transfected with IRGMb, IRGMd-WT (wild type IRGMd, unaltered) or IRGM-S47N mutant (SN) for 48 h, labeled with MTR and imaged by live microscopy. c. Fluorescence intensity analysis of green and red channels along a line drawn through two adjacent cells, one GFP-IRGMd positive and one GFP-IRGMd negative. d. Quantification of GFP+MTR+ cells. e,f. Analysis of GST-IRGMd binding to lipids by lipid protein binding dot blots (e) and cardiolipin bound agarose bead pull down assay (f; details in Methods). g. IRGMd, IRGMdS47 mutant, and IRGMb isoform concentration-dependent analysis of binding to lipids on strip blots (numbers identifying lipids correspond to the legend in panel e). GST control is in Suppl. Fig. S1d. Membranes in e-g were probed with anti-GST antibody.