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Adhesion of cells to the extracellular matrix (ECM) via focal adhesions (FAs) is crucial for cell survival, migration, and differentiation. Although the regulation of FAs, including by integrins and the ECM, is important to cell behavior, how FAs are regulated is not well known. Autophagy is induced by both cell adhesion and cell detachment. Here, we showed that autophagosomes are located close to internalized collagen and paxillin, which is a well-known marker of FAs. Autophagy-deficient cells showed increased levels of internalized collagen compared with control cells. Moreover, paxillin exhibited a more peripheral distribution and the area of paxillin was increased, and adhesion-induced focal adhesion kinase signaling was impaired and adhesion was enhanced, in autophagy-deficient cells. These results suggest that autophagy suppressed cell adhesion by regulating internalized ECM and FAs.