FIG. 4.

TFEB induction controls CSE-induced autophagy-impairment. (A, B) Flow cytometry analysis verifying that CSE-induced coexpression of Ub and p62 (aggresome-bodies) is controlled by treatment with GEM (mean ± SEM, n = 3, p = 0.07). (C, D) Western blot analysis showing accumulation of ubiquitinated proteins (Ub), VCP, and TFEB in the soluble and insoluble protein fractions of Beas2b cells treated with 10% CSE and/or GEM (10 μM) for 6 h. β-Actin was used as the loading control. The data represent three replicates and show that GEM treatment significantly inhibits CSE-induced ubiquitinated protein and TFEB accumulation in the insoluble protein fractions (aggresome-bodies). (D) Similarly, FIS (25 μM)-mediated TFEB/autophagy induction also showed clearance of CSE-induced ubiquitinated proteins from soluble and insoluble protein fractions. Densitometry analysis of Western blots is shown in (E and F). Data represent n = 3, mean ± SEM, *p < 0.05, **p < 0.01, ***p < 0.001. Data indicate that GEM or FIS-mediated TFEB induction controls CSE-induced autophagy-impairment. VCP, valosin-containing protein. To see this illustration in color, the reader is referred to the web version of this article at www.liebertpub.com/ars