Fig. 5: Decreases in intracellular glutamate levels and glutamate tRNA charging during adipocyte differentiation.
From: Cross-talks between Metabolic and Translational Controls during Beige Adipocyte Differentiation
a Diagram showing how GLS, GLUL, GLUD1, GOT1, GOT2, GPT2, BCAT1, and BCAT2 mediate glutamate metabolism (top) and bar plots depicting their expression changes during beige adipocyte differentiation (bottom). Replicate N = 3 for each bar. b Glutamate to glutamine ratio during beige adipocyte differentiation with or without treatment of methionine sulfoximine (MSO), a glutamine synthetase (GLUL) inhibitor. Replicate Nnon-MSO = 3, NMSO = 2. c Changes in the ratios of charged versus uncharged tRNAs were measured in preadipocytes and beige adipocytes using a qPCR-based tRNA aminoacylation assay. Primers specific to tRNAGly(TCC), tRNAGln(CTG), and tRNAGlu(TTC) were used. Replicate N = 3. d Assessment of tRNAGlu(TTC) charging. Top panel shows northern blot of tRNAGlu(TTC) with deacylated sample (left) and lysates of 0 day (middle) or 8 days (right) of beige adipocyte differentiation. Bottom panel shows 5S and 5.8S rRNA stained with SYBR Gold. Representative images among three replicates are shown. e Schematic of the reporter assay for the doxycycline inducible Glu or Ala codon-rich EGFP expression (top) and flow cytometry results of 3T3-L1 cells showing changes in EGFP expression of the reporters whether doxycycline and BPTES treatment. (bottom). Replicate N = 3. f Bar plots displaying expression changes of genes involved in glutamate metabolism during white and brown adipocyte differentiation. Replicate N = 2. g A heat map showing single-nucleus expression of indicated genes in ASPCs or adipocytes. Each column of the heat map represents one nucleus, and columns are grouped into ASPCs (left) and adipocytes (right). h qRT-PCR results showing relative mRNA levels of adipogenic and thermogenic markers under differentiation with or without MSO treatment. Replicate NPparg2,Fabp4,Adipoq = 6, NUcp1,Cox8b = 2. Statistical tests: two-sided unpaired t-test for p-values of b, h; and two-sided paired t-test for p-values of c, e. *p < 0.05, **p < 0.01, ***p < 0.001, NS means ‘not significant’. Bar plots of a–c, e–g are shown as mean (SD). Source data are provided as a Source Data file.
