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. 2021 May 17;12:2876. doi: 10.1038/s41467-021-23085-0

Fig. 7. p62 and NBR1 interact with PPARγ in brown adipocytes.

Fig. 7

a Representative immunoblotting of p62 and NBR1 levels in cytoplasmic/nuclear fractions in primary brown adipocytes. Cells were treated with 0.5 µM isoproterenol (ISO) for indicated time. Densitometric quantification of nuclear NBR1 and p62 levels was also shown (n = 3 independent experiments). b, c Endogenous interaction of PPARγ with p62 and NBR1. p62 (b) or PPARγ (c) immunoprecipitates from nuclear lysates extracted from ISO and rosiglitazone-treated brown adipocytes were analyzed for the levels of specified proteins. Representative blots and densitometric quantification were shown (n = 3 independent experiments for both). d, e Recombinant FLAG-p62 (d) or FLAG-NBR1 (e) was incubated with GST and GST-PPARγ proteins separately and the interactions were analyzed by immunoblotting in glutathione-beads pull-down. Representative blots and densitometric quantification were shown (n = 4 independent experiments for d and n = 3 independent experiments for e). f HEK293T cells were transfected with cDNA vectors expressing WT/mutants of HA-p62 or HA-NBR1, and GST-PPARγ. The interacting proteins were pulled down using glutathione-beads against GST-PPARγ and analyzed by immunoblotting. Representative blots from three independent replicates with similar results were shown. Data are presented as mean ± SEM (a–e). *p < 0.05, **p < 0.01. Two-tailed Student’s T-test (a–e). Source data are provided as a Source Data file.