Fig. 3: RAS regulates localization of SLC3A2 and MTOR to endolysosomes. | Nature Communications

Fig. 3: RAS regulates localization of SLC3A2 and MTOR to endolysosomes.

From: Oncogenic RAS commandeers amino acid sensing machinery to aberrantly activate mTORC1 in multiple myeloma

Fig. 3

a BioID2-SLC3A2 interactomes in XG2 cells transduced with control shRNA (x-axis) or shKRAS (y-axis), proteins are colored that decreased by at least 0.5 log2fc (purple) or increased by at least 0.5 log2fc (blue). b Proximity ligation assay (PLA) (red) of SLC3A2 with MTOR in RPMI 8226 and SKMM1 cells. Cells counterstained with DAPI (blue) and wheat germ agglutinin (WGA; green); Scale bar is 10 μm. Representative images; n = 3. c SLC3A2-MTOR PLA score of cells transduced with indicated shRNAs. Details about box plots below. d Gene Ontology pathway enrichment of SLC3A2 interactors that decreased by at least 0.5 log2fc following RAS knockdown. Bonferroni corrected P value plotted on the x-axis. e PLA of SLC3A2 with LAMP1 (red) in RPMI 8226 and SKMM1 cells. Cells counterstained with DAPI (blue) and WGA (green); Scale bar is 10 μm. Representative images; n = 3. f SLC3A2-LAMP1 PLA score of cells transduced with control shRNA or shRNAs specific for KRAS, NRAS, SLC3A2 and MTOR. Details about box plots below. g Immunofluorescence images of MTOR (green), LAMP1 (red) and DAPI (blue) in RPMI 8226 and SKMM1 cells expressing shCTRL, shKRAS.2 or shNRAS.1. Scale bar is 10 μm. Representative images; n = 3. h PLA of MTOR with LAMP1 (red) in RPMI 8226 and SKMM1 cells. Cells counterstained with DAPI (blue) and WGA (green); Scale bar is 10 μm. Representative images; n = 3. i MTOR-LAMP1 PLA score of cells transduced with control shRNA or shRNAs specific for MTOR, KRAS, NRAS, SLC3A2 and RPTOR. Details about box plots below. For all box plots: Data are pooled independent experiments; the number of cells quantified per condition indicated in the source data file. *** denotes P value <0.0001, ** denotes P value = 0.0023, n.s. denotes not significant, as determined by one-way ANOVA with Dunnett’s post test. The number of independent experiments and the number of cells quantified for each condition can be found in the source data. Box plots represent median and 25–75% of data, whiskers incorporate 10–90% of data, outliers are displayed as dots. Source data are provided as a Source Data file.

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