Small GTPase Rab1B is associated with ATG9A vesicles and regulates autophagosome formation

Abstract

ATG9 is a membrane protein that is essential for autophagy and is considered to be directly involved in the early steps of autophagosome formation. Yeast Atg9 is mainly localized to small vesicles (Atg9 vesicles), whereas mammalian ATG9A is reportedly localized to the -Golgi network, the endosomal compartment, and other unidentified membrane structures. To dissect the ATG9A-containing membranes, we examined the subcellular localization of ATG9A and performed immunoisolation of those membranes. ATG9A-green fluorescent protein in human culture cells was observed as numerous puncta that move rapidly throughout the cytoplasm. We isolated these cytoplasmic membranes and found that they were small vesicles that resemble the yeast Atg9 vesicle. One of the proteins obtained proteomic analyses of the mammalian ATG9A vesicle was Rab1, a small GTPase that is essential in endoplasmic reticulum-to-Golgi vesicle trafficking. Knockdown studies of Rab1B showed a suppression of autophagy. In these Rab1B-depleted cells, ATG9A accumulated in intermediate membrane structures at autophagosome formation sites. These results indicate that Rab1B is involved in regulating the proper development of autophagosomes.-Kakuta, S., Yamaguchi, J., Suzuki, C., Sasaki, M., Kazuno, S., Uchiyama, Y. Small GTPase Rab1B is associated with ATG9A vesicles and regulates autophagosome formation.

ER ‐to‐lysosome‐associated degradation of proteasome‐resistant ATZ polymers occurs via receptor‐mediated vesicular transport

“…Yet, nutrient deprivation‐induced ER‐phagy critically depends on FIP200 (Khaminets et al , ), a core component of the ULK kinase complex that is required for the generation of autophagosomes that eventually capture and deliver ER fragments to the lysosomal compartments (Hara et al , ; Hosokawa et al , ; Smith et al , ). Strikingly, our analyses reveal that the FAM134B‐dependent ATZ delivery to endolysosomes is not affected in cells lacking FIP200 (Fig D and I, and EV3B) or other components of the autophagosome biogenesis machinery such as ULK1 and ULK2 (Fig E and I, and EV3A and B), ATG13 (Fig F and I, and EV3A and B) or ATG9 (Fig G and I, and EV3A and B; Saitoh et al , ; Shang et al , ; McAlpine et al , ; Bento et al , ; Kaizuka & Mizushima, ; Hurley & Young, ; Kakuta et al , ). Thus, defective autophagosome biogenesis impairs several types of receptor‐mediated ER‐phagy (Khaminets et al , ; Grumati et al , ; Fregno & Molinari, ; Loi et al , ; Smith et al , ) and conventional macroautophagy (Fig EV3B; Hara et al , ; Hosokawa et al , ; Kakuta et al , ).…”

RAB2 regulates the formation of autophagosome and autolysosome in mammalian cells

“…Studies in yeast have shown that YPT1 is recruited and activated by its GEF, TRAPIII, at PAS, which is followed by YPT1-mediated ATG1 recruitment [35,71]. RAB1, the mammalian orthologue of YPT1, also regulates autophagy [28,[72][73][74][75][76][77]. In addition, RAB1, together with TRAPPIII and TBC1D14, maintains an ATG9 pool on the Golgi apparatus by acquiring the vesicles from recycling endosomes [28].…”

Calmodulin‐mediated events during the life cycle of the amoebozoan Dictyostelium discoideum

“…A Calmodulin Target Database scan revealed a putative, unusually short CaMBD in D. discoideum Atg9 (221IANRIMRK228) suggesting the ortholog may also bind to CaM but via a non‐canonical motif (S. Mathavarajah, unpublished data). This idea is strengthened by the result that CaM co‐immunoprecipitated during a proteomic screen of Atg9 interactors in human cells (Kakuta et al, ). These results link Ca 2+ fluxes to the progression of autophagy and suggest a potential role for CaM in regulating autophagosome formation.…”