A hallmark of macroautophagy is the formation of autophagosomes, double-membrane vesicles

A hallmark of macroautophagy is the formation of autophagosomes, double-membrane vesicles that enwrap cellular components destined for lysosomal degradation. member of the retromer complex, which is responsible for retrograde transport of cargo from endosomes to the trans-Golgi network and is composed of the selective subunits Vps26, Vps29 and Vps35 and the sorting nexin subunits Vps5 and Vps17. Also, strains missing the components Vps5, Vps17 and Vps29 show reduced autophagy. Hence, it appears that the retromer is usually actively modulating autophagy in yeast possibly playing a role in membrane trafficking. Moving back to the human system, we could validate the obtaining in yeast that eukaryotic translation elongation factor 1 gamma (EEF1G) is usually a positive regulator of autophagy signaling upstream of MTOR. We also identified an interesting link between the proteasome and autophagy, the proteasome seeming to be one of the favorite substrates of autophagosomesan observation already made by others. Atrasentan hydrochloride We identified proteasomal proteins associated with autophagosomes irrespective of the stimuli used. During active autophagy, proteasomal protein abundance, as well as proteasome activity decreases in cells. However, the molecular mechanisms underlying autophagy-proteasome crosstalk are still not Atrasentan hydrochloride fully uncovered. Thus, it remains unknown if proteasomes are active inside autophagosomes and if autophagosomes can be regarded as scaffolds bringing together the proteasome with its substrates. As we identified basically all components of the 20S proteasome this might be a valid option. Regardless of the mechanism, it becomes clear that there is a balance between the two degradation systems, which can be shifted in favor of one or the other. We also observed this for yeast: rapamycin increases levels of autophagy in temperature sensitive or mutants, which express inactive proteasomes Atrasentan hydrochloride upon a temperature rise, compared with wild-type strains treated with rapamycin. The double-mutant displays synergistic effects. Taken together, by employing an unbiased organellar proteomics approach we studied the dynamics of autophagosomes in response to different cues. We highlighted stimulus differences and similarities shedding new light on macroautophagy as a targeted degradation process. Combining yeast and human Atrasentan hydrochloride cell culture systems, new conserved autophagy regulators could be identified and the crosstalk between the autophagosome on the one hand and the retromer and the proteasome on the other hand was studied. To Atrasentan hydrochloride our opinion, global Rabbit Polyclonal to EMR2 omics approaches in combination with data modeling hold the promise to further delineate macroautophagy and underlying spatiotemporal protein dynamics. Our study provides a resource of proteins implicated in autophagy either as substrates or components of the autophagosomal machinerythis data set gives a glimpse of the autophagosomal food-chain and will hopefully aid further studies in delineating the signals and mechanisms that govern spatiotemporal specificities of cargo selection during (stress-induced) macroautophagy. Notes Dengjel J, H?yer-Hansen M, Nielsen MO, Eisenberg T, Harder LM, Schandorff S, et al. Identification of autophagosome-associated proteins and regulators by quantitative proteomic analysis and genetic screens Mol Cell Proteomics 2012 11 M111.01403 doi: 10.1074/mcp.M111.014035. Footnotes Previously published online: www.landesbioscience.com/journals/autophagy/article/20286.