Extended Data Fig. 5: RLR signaling is regulated by a lactate and glucose-independent mechanism despite increased glucose tolerance after Ubxn9 deletion.
From: UBXN9 governs GLUT4-mediated spatial confinement of RIG-I-like receptors and signaling
a, b, Immunoblots of LDHA and β-actin in Ldha+/+ and Ldha−/− C2C12 myocytes (a) and extracellular lactate levels after 12 h of culture (n = 5 biological replicates, two independent experiments (b). c, d, Cellular Ifnb1 transcripts in (c) and secreted IFN-β protein levels (d) from C2C12 cells (n = 2 biological replicates/group) stimulated with 3p-hpRNA. e, Immunoblot loading controls for the T-tubule and muscle homogenate fractions from Ubxn9+/+ and Ubxn9−/− mice (n = 3 mice/group for example purposes) shown in Fig. 3c. f, Plasma lactate concentrations (n = 6 mice/group) before and after infection with EMCV (100 PFU/mouse). g, Schematic for intraperitoneal (i.p.) glucose tolerance test (IPGTT) with Ubxn9+/+ and Ubxn9−/− littermates. h, Glycemia (left) and area under curve (AUC) (right) during IPGTT (n = 13 Ubxn9+/+ and 12 Ubxn9−/− mice pooled from 3 independent experiments). i, Intracellular lactate levels in C2C12 cells (n = 4 biological replicates/group) before and after EMCV (MOI: 0.5). j, IFN-β protein release from Ldha+/+ and Ldha−/− C2C12 cells (n = 2 biological replicates/group for untreated; n = 4 biological replicates/group for 3p-hpRNA) that were transfected with an empty vector or GLUT4 expression plasmid (pGLUT4) before 3p-hpRNA treatment for 12 h. k, 2-DG uptake in DMSO or Fasentin (25 μM) pretreated WT C2C12 cells (n = 3 biological replicates/group). l, Ifnb1 transcripts in C2C12 cells (n = 4 biological replicates/group) treated simultaneously with Fasentin (25 μM) and EMCV (MOI: 0.5) for 6 h. Bar: mean ± SEM. P values determined by unpaired two-tailed Student’s t-test (b, h, i-k) and two-way ANOVA with Šidák’s multiple comparisons test (f, h, l).
