Skip to main content
. 2017 Aug 28;18(9):1865. doi: 10.3390/ijms18091865

Figure 1.

Figure 1

(A) The site of autophagosome formation. Upstream autophagy factors and autophagy substrates accumulate at the autophagosome formation site independently from one another. The most upstream autophagy factors are ATG9A in vesicles and the ULK1 complex. Autophagy substrates include p62/SQSTM1, ferritin, and damaged mitochondria. The autophagosome formation site localizes in close apposition to the endoplasmic reticulum (ER); (B) Isolation membrane formation. Isolation membranes emerge from within the ring-shaped structure called the omegasome. The omegasome is a phosphatidylinositol 3-phosphate -rich ER subdomain characterized by the existence of double-FYVE domain-containing protein 1 (DFCP1). ATG factors commonly used to visualize isolation membranes include ULK1, WIPI1/2, ATG5, and LC3. Most ATG factors but not LC3 homologs detach from the isolation membranes before completion of autophagosomes. These isolation membrane markers are localized on the outer membrane, whereas LC3 and its homologs and associating p62/SQSTM1 are localized on both outer and inner membranes. Isolation membranes are observed by electron microscopy as crescent-shaped structures that are often flanked by rough ER; (C) Completion of autophagosome formation. Several minutes after detachment of ATG factors, LC3-positive autophagosomes acquire syntaxin17 (STX17). STX17 localizes to ER, mitochondria, and complete autophagosomes; therefore, not all the STX17 signals represent autophagosomes. Cytoplasmic components enclosed in the characteristic double-membraned structures mark autophagosomes observed by electron microscopy (EM). The width of the cleft between the outer and inner membranes can vary due to experimental procedures such as fixation; (D) Fusion with lysosomes. Autophagosomes that have acquired STX17 fuse with lysosomes to degrade the contents of the autophagosomes. Upon fusion with lysosomes, the space between the outer and inner autophagosomal membranes becomes acidified, followed by collapse of the inner membrane; (E) Autolysosome. Fusion of autophagosomes and lysosomes generates autolysosomes that contain degraded cytoplasmic components. Autolysosomes can be identified by EM or colocalization of LC3 and lysosomal markers by fluorescence microscopy.