| dc.description.abstracteng | Autophagy is a degradative pathway that is conserved from yeast to mammals. Marcoautophagy is characterized by the formation of a unique double-membrane layered vesicle, the so-called autophagosome. Proteins and organelles are engulfed by the autophagosome and transported to the degradative compartment, which is the vacuole in yeast and plants or the lysosome in mammals. At the degradative compartment, the outer membrane of the autophagosome fuses with the membrane. Then the mono-layered inner vesicle, the autophagic body, is released and its contents are degraded. Autophagy can be a selective process that targets specific proteins or organelles to the degradative compartment by using specific cargo receptors. During unselective autophagy proportions of the cytoplasm are enclosed in a non-specific manner and degraded. The autophagic process is important to maintain cell viability especially under stress conditions and to adapt to changing environments. Degraded components can finally be reused to maintain nutrient supply for the cell. Autophagy plays a role in various processes in the cell like aging, cell homeostasis and host defense. Malfunction of the autophagic process is associated with various diseases like cancer or neurodegeneration. Thus, autophagy has to be tightly controlled. One element of autophagy that needs to be tigthly controlled is the membrane traffic to the PAS, which is required for the formation of the autophagosome.
Key regulators of intracellular membrane traffic are the Ypt/Rab GTPases (Ypts in yeast and Rabs in mammals), which mediate directed and specified intracellular vesicle transport through the cell to their final destination. Ypt/Rab GTPases are localized to different compartments and contribute to membrane identity. To fulfill their duties in the cell, Ypt/Rab GTPases cycle between a GTP-bound form, which activates the protein and allows binding to its effectors, and a GDP-bound form. The transition of both forms requires GEFs (guanine-nucleotide exchange factors) and GAPs (GTPase-activating proteins). Ypt1 is a yeast Rab GTPase that mediates vesicle transport and also plays a role in autophagy. Ypt1 interacts with Atg proteins and contributes to phagophore formation.
Previous to this study, Gyp1 has been identified as a potential interaction partner of Atg8 by mass spectrometry. Atg8 plays a role in elongation and closure of the phagophore and targets cargos to the autophagosome during selective forms of autophagy. Gyp1 is a GAP and known to negatively regulate the Rab GTPase Ypt1 by stimulating its GTPase activity. Three different GAPs, Gyp1, Gyp5 and Gyp8 have been shown to negatively regulate the Rab GTPase Ypt1 by stimulating its GTPase activity in vivo.
This study shows that Gyp1 affects the Cvt-pathway, demonstrated by analysis of pApe1 maturation, proper proceeding of post-log mitophagy and might also play a role during Atg8-dependent recognition of other targets. Two functions of Gyp1 are proposed. First, Gyp1 seems to regulate the efficient dissociation of the Ypt1-Atg1 complex during initial steps of phagophore assembly. In the absence of Gyp1, this complex is enriched and the recruitment of downstream proteins is impaired, as shown for Atg14 in this study. This indicates that dissociation of Ypt1 and Atg1 is a prerequisite for recruitment of Atg proteins and proceeding of selective autophagy. For this process, the GAP activity of Gyp1 is needed. Gyp5 and Gyp8 might compensate the loss of Gyp1 as additional deletion of the two other Ypt1 GAPs leads to a more severe impairment of the Cvt-pathway. Single-deletion of both proteins causes no defect. Thus, Gyp1 might function as the main GAP during the Cvt-pathway.
The second proposed function of Gyp1 was detected in later steps of selective autophagy. Here, Gyp1 interacts in an AIM-dependent manner with Atg8. In the absence of Gyp1, the function of Atg8 is impaired, which was demonstrated in this study with the mitophagy receptor Atg32. Upon deletion of GYP1, Atg8 and Atg32 show a reduced binding, which indicates a role of Gyp1 during cargo recognition. The analysis of Gyp1 mutants revealed that this function is AIM-dependent but GAP activity independent.
Thus, this study proposes a dual role for Gyp1 during selective autophagy. First, it leads to the efficient dissociation of the Ypt1-Atg1 complex and later Gyp1 interacts with Atg8, which seems to be important for proper binding of Atg8 to selective cargo receptors. | de |