Autophagy regulates transforming growth factor β signaling and receptor trafficking

by Selleckchem | Aug 27 2023

Transforming growth factor beta (TGFβ) stimulates tumorigenesis by inducing epithelial to mesenchymal transition (EMT) and cell migration. TGFβ signaling is regulated by the endocytosis of cell surface receptors and their subcellular trafficking into the endo-lysosomal system. Here we investigated how autophagy, a cellular quality control network that delivers material to lysosomes, regulates TGFβ signaling pathways that induce EMT and cell migration. We impaired autophagy in non-small cell lung cancer cells using chloroquine, spautin-1, ULK-101, or small interfering RNA (siRNA) targeting autophagy-related gene (ATG)5 and ATG7 and observed that inhibiting autophagy results in a decrease in TGFβ1-dependent EMT transcription factor and cell marker expression, as well as attenuated stress fiber formation and cell migration. This correlated with decreased internalization of cell surface TGFβ receptors and their trafficking to early/late endosomal and lysosomal compartments. The effects of autophagy inhibition on TGFβ signaling were investigated by Smad2/Smad3 phosphorylation and cellular localization using western blotting, subcellular fractionation, and immunofluorescence microscopy. We observed that inhibiting autophagy decreased the amount and timeframe of Smad2/Smad3 signaling. Taken together, our results suggest that inhibiting autophagy attenuates pro-tumorigenic TGFβ signaling by regulating receptor trafficking, resulting in impaired Smad2/Smad3 phosphorylation and nuclear accumulation.

Comments:

The passage describes a study that investigates the role of autophagy in regulating the TGFβ signaling pathway, which is known to stimulate tumorigenesis by inducing epithelial to mesenchymal transition (EMT) and cell migration. The researchers impaired autophagy in non-small cell lung cancer cells using various methods such as chloroquine, spautin-1, ULK-101, and small interfering RNA (siRNA) targeting autophagy-related genes (ATG5 and ATG7). They then observed the effects of autophagy inhibition on TGFβ1-dependent EMT transcription factor and cell marker expression, stress fiber formation, and cell migration.

The results showed that inhibiting autophagy led to a decrease in the expression of EMT transcription factors and cell markers, as well as attenuated stress fiber formation and cell migration in response to TGFβ1. This was accompanied by a reduction in the internalization of cell surface TGFβ receptors and their trafficking to endosomal and lysosomal compartments within the cell.

The study further investigated the effects of autophagy inhibition on TGFβ signaling by examining the phosphorylation and cellular localization of Smad2 and Smad3, which are key signaling molecules downstream of TGFβ receptors. The researchers used techniques such as western blotting, subcellular fractionation, and immunofluorescence microscopy to analyze Smad2/Smad3 signaling. They found that inhibiting autophagy resulted in decreased levels and duration of Smad2/Smad3 signaling, indicating impaired activation of the TGFβ signaling pathway.

Overall, these findings suggest that autophagy plays a role in regulating pro-tumorigenic TGFβ signaling by influencing the internalization and trafficking of TGFβ receptors, ultimately impacting the phosphorylation and nuclear accumulation of Smad2 and Smad3.