Deacidification of the Endolysosomal System by the Vesicular Proton Pump V-ATPase Inhibitor Bafilomycin A1 Affects EGF Receptor Endocytosis Differently in Endometrial MSC and HeLa Cells

TL;DR

This study compares how tumor cells and endometrial MSCs handle EGF receptor endocytosis when the acidity of endosomes is reduced.

Contribution

The paper reveals distinct endocytic behaviors in non-tumor endometrial MSCs versus tumor cells under V-ATPase inhibition.

Findings

Bafilomycin A1 slows EGFR movement in HeLa and A549 cells but not in endometrial MSCs.

Endometrial MSCs retain activated EGFR in pH-independent endosomes longer than tumor cells.

V-ATPase inhibition alters EGF-receptor co-localization with endocytic markers in cell-type-specific ways.

Abstract

It is well-known that EGF binding to EGFR stimulates signal transduction and endocytosis, with the latter leading to lysosomal degradation of EGFR. However, the majority of data on the regulation of endocytosis have been obtained in tumor-derived cells. Here, we perform a comprehensive analysis of the role of endolysosome acidification in the regulation of endocytic pathway in tumor cells and in endometrial MSCs as a model of proliferating, undifferentiated, non-immortalized cells. Using QD-labeled EGF, the dynamics of co-localization of EGF-receptor complexes with endocytic markers in the control and upon inhibition of V-ATPase by Bafilomycin A1 (BafA1) were studied using confocal microscopy. Image analysis showed that in HeLa and A549 cells, BafA1 significantly slowed down EGFR entry into and exit from EEA1-positive early endosomes without disrupting passage through Rab7, CD63, and Lamp1 compartments, but rather shifting it to later times. In enMSCs, only a portion of EGF-containing endosomes entered the degradation pathway, and lysosomal delivery was significantly delayed. Unlike HeLa, in enMSC early endosomes, BafA1 increased the association of EGF-QDs with EEA1, suggesting a lower pH level, which is suboptimal for EEA1-dependent fusions. It is concluded that, unlike HeLa, enMSCs form a population of pH-independent endosomes containing activated EGFR for a long time.

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