Skip to main content
. Author manuscript; available in PMC: 2020 Nov 1.
Published in final edited form as: Nat Metab. 2020 May 21;2(5):397–412. doi: 10.1038/s42255-020-0205-x

Figure 5. Functional role of LINC00473.

Figure 5.

a. RT-PCR of LINC00473 in SClav treated with control or LINC00473-directed siRNA oligos 48h prior to stimulation with NE. Plotted are individual values, means and SEM at each time point of n=3 independent experiments. Statistical differences relative to control (siScrambled) were calculate using ANOVA corrected for multiple comparisons using the Holm-Sidak test. b. Oxygen consumption in primary adipocytes at day 12 of differentiation, treated with scrambled or LINC00473 targeted siRNAs 72 h prior to assay. Indicated are the times of addition of norepinephrine (NE), Oligomycin (OLIG), FCCP, and rotenone/antimycin (ANT) at concentrations indicated in Methods. Values are means and SEM of three experiments (n=3). c. Mitochondrial respiratory parameters were calculated using three paired time points per condition, per experiment, for an n=9 as follows: NE-induced=NE minus Basal; ATP-linked=NE minus OLIG; Proton leak=ANT minus OLIG; Respiratory reserve capacity=FCCP minus Basal; Maximal Respiratory Capacity=FCCP minus ANT. Plotted are the individual values, means and SEM. Statistical significance between Control and siLINC00473 were calculated using paired, 2-tailed student t-tests. d,e. Expression levels of the major isoform of LINC00473 (d) and of 5 splice variants (e) in in adipocytes expressing either empty vector or overexpressing LINC00473 through the CRISPR-SAM guide 6 (Gd6), stimulated with 10 μM Fsk for 6 hours. Shown are the individual values, means and SEM of data from n=6 independent cell cultures. Statistical significance of differences was calculated using ANOVA corrected for multiple comparisons using the Sidak test. f,g. Oxygen consumption rates (f), and respiratory parameters (g) in adipocytes expressing either empty vector or Gd6. Indicated are the times of addition of Forskolin (Fsk), Oligomycin (OLIG), and FCCP. Plotted in (f) are the means and SEM of six experiments performed in triplicate using two cell populations (n=6). Plotted in (g) are the means and SEM of the respiratory parameters calculated for each experiment (n=6). Statistical significance between Empty Vector and Gd6 and siLINC00473 were calculated using paired, 2-tailed student t-tests. h,i. Free fatty acids (FFA) (mmol/L) in media with and without Fsk stimulation in adipocytes expressing either empty vector or Gd6, (h) and in Gd6 cells transfected with scrambled (siScrambled) or LINC00473-directed (siLINC00473) pools of siRNA oligonucleotides (i). Ploted are the individual values, means and SEM of n=3 (FSK stimulation), and n=5 (without stimulation) independent cell cultures. j. Mitochondria DNA levels in cells treated as in panels h,i. Plotted are the individual values, means and SEM of n=8 (Empty Vector) and n=4 (Gd6, siScrambled and siLINC00473) independent cell cultures.