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. 2014 Nov;82(11):4666–4680. doi: 10.1128/IAI.02034-14

FIG 4.

FIG 4

(A, B) Modulation of autophagy was analyzed by RT-PCR in HeLa Tet-On Advanced (HTAC) cells transfected with mapA cloned in pTRE2Hyg (HTAC-pTRE33Hyg cells). p62 gene expression was studied by RT-PCR at 2 h and 16 h after doxycycline induction of HTAC-pTRE33Hyg cells. Relative expression was based on HTAC-pTRE2Hyg expression (y axis) at 16 h, which was assigned a value of 1. The same samples were analyzed for mapA expression by HTAC-pTRE33Hyg cells at 0 h, 2 h, and 16 h after doxycycline induction. (C) Detection of Omp33-36 and LC3B accumulation in HTAC-pTRE33Hyg cells by immunoblotting for the detection of Omp33-36 and LC3B-I/LC3B-II by polyclonal antibodies in HTAC-pTRE33Hyg cells under the control of a doxycycline-inducible promoter. *, densitometry of LC3B-II 0 and 24 h after doxycycline induction of HTAC-pTRE33Hyg cells. (D, E) Presence of the features of apoptosis and probable phagophore structures (isolate membrane to form an autophagosome) determined by TEM of HTAC-pTRE33Hyg cells expressing mapA after 24 h of doxycycline induction. (F, G) Absence of characteristics of the apoptosis and autophagy mechanisms determined by TEM of HTAC cells transfected with pTRE2Hyg (empty plasmid) after 72 h of doxycycline induction (negative control).