Inhibition of autophagy with bafilomycin and chloroquine …
2017年4月1日 · We found that both bafilomycin and chloroquine could significantly increase the autophagosome marker LC3-II and inhibit key parameters of mitochondrial function, and increase mtDNA damage. Furthermore, we observed significant alterations in TCA cycle intermediates, particularly those downstream of citrate synthase and those linked to ...
Robust LC3B lipidation analysis by precisely adjusting ... - Nature
2022年1月7日 · Here we scrutinize this approach and show that bafilomycin A1 (BafA) but not chloroquine is suitable for flux quantification due to the stimulating effect of chloroquine on non-canonical LC3B ...
Control of infection by LC3-associated phagocytosis, CASM, and
2022年10月26日 · This can be achieved by expression of the SopF virulence factor from Salmonella, which blocks binding of the WD domain of ATG16L1 to V-ATPase, and drugs that promote (saliphenylhalamide) or inhibit (bafilomycin) assembly of the V o-V 1 complex, and using diphenyleneiodonium (DPI) to inhibit NADPH oxidase (20, 26).
An improved method for high-throughput quantification of
2020年7月22日 · Chloroquine and Bafilomycin A1 treatment led to an accumulation of perinuclear autophagosomes (Fig. 2B). The pixel area of eGFP-LC3B, which correlates with autophagy levels, was quantified using...
Intracellular cholesterol transport inhibition Impairs autophagy flux ...
2022年11月25日 · Colocalization of lysosome marker LAMP1 to GFP-LC3 was tested to confirm whether MβCD could rescue the impaired autophagic flux by enhancing autophagosome–lysosome fusion in the presence of U18666A or bafilomycin A1. Interestingly, GFP-LC3 was found to be extensively colocalized with LAMP1 in the control cells (Additional file 1: Fig S2A, B ...
Application and interpretation of current autophagy inhibitors and ...
2013年3月25日 · Data from the relatively recent literature also show that bafilomycin A1 and rapamycin potentiate ethanol-increased LC3 lipidation, whereas wortmannin and a BECN1-specific shRNA inhibit ethanol...
LC3- and p62-based biochemical methods for the analysis of …
2015年3月15日 · The amount of LC3-II significantly increased upon bafilomycin A 1 treatment, suggesting that lysosomal degradation of LC3-II is inhibited. The difference in LC3-II levels between lane 1 and lane 2 indicates the amount of LC3-II that is constitutively delivered to the lysosome for degradation under normal growth conditions (i.e., basal ...
Differential regulation of autophagy during metabolic stress in ...
2019年12月26日 · Co-treatment with bafilomycin A 1 and torin1 increased LC3-II levels relative to bafilomycin A 1 alone (Figure 1F,G), indicating an increase in autophagic flux with torin1 treatment. Consistent with the immunofluorescence data, the immunoblotting results also support that MTOR inhibition activates autophagy in astrocytes.
Monitoring Autophagy Flux and Activity: Principles and …
2020年8月26日 · If autophagy is induced under starvation for a certain period (usually more than 2 h), in the presence of Bafilomycin A 1 or E64d plus Pepstatin A, LC3-II accumulates markedly compared with control (in the absence of inhibitors), due to inhibition of LC3-II …
Dissecting the dynamic turnover of GFP-LC3 in the autolysosome
Unsaturating concentrations of chloroquine or bafilomycin A1 increased the level of free GFP fragments while saturating concentrations did not. Data from the present study demonstrate that GFP-LC3 is degraded in a step-wise fashion in the autolysosome, in which the LC3 portion of the fusion protein appears to be more rapidly degraded than GFP.