Microautophagic Vacuole Invagination Requires Calmodulin in a Ca2+-independent Function

Uttenweiler, Andreas; Schwarz, Heinz; Mayer, Andreas

doi:10.1074/jbc.m506086200

Cited by 45 publications

(45 citation statements)

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“…Vtc-Cmd1p interaction by starvation. This correlates with our ealier observation that calmodulin appears not to fulfill a Ca 2ϩ -dependent regulatory function in microautophagy (Uttenweiler et al, 2005).…”

Section: Vtc Complex In Microautophagysupporting

confidence: 76%

“…Both Vtc2p, Vtc3p, and Vtc4p coimmunoprecipitated with Cmd1p ( Figure 7C) in the presence of the Ca 2ϩ chelator EGTA, but less in the presence of Ca 2ϩ . In line with this, microautophagic uptake in vitro was stimulated by EGTA and inhibited by Ca 2ϩ (Uttenweiler et al, 2005 and data not shown). We also tested if the interaction between the Vtc proteins and Cmd1p is regulated by starvation.…”

supporting

confidence: 61%

“…Structural characterization may also resolve the apparent paradoxon that the calmodulin antagonist W-7 inhibits in vitro microautophagy (Uttenweiler et al, 2005) but does not disrupt the Vtc-Cmd1p interaction. Cmd1p might bind Vtc proteins in a conformation that is not influenced by W-7, or W-7 may alter the apo-conformation to partially resemble the Ca 2ϩ -loaded state.…”

Section: Discussionmentioning

confidence: 99%

“…According to their ability to block the reaction at different kinetic stages, these inhibitors have been defined as early acting class A inhibitors (nystatin, GTP␥S, aristolochic acid) and late acting class B inhibitors (W-7, valinomycin/FCCP, K252a, and rapamycin). A putative W-7 target, calmodulin (Cmd1p), acts late during invagination in a calcium-independent way (Uttenweiler et al, 2005).…”

Section: Introductionmentioning

confidence: 99%

“…GFP-Pho8p does not concentrate in invaginations upon treatment with rapamycin ( Figure 5C). Thus, Vtc proteins are recruited to the vacuole and also to vacuolar invaginations upon nutrient limitation, supporting their role in microautophagy.Calmodulin (Cmd1p) has a Ca 2ϩ -independent role in microautophagic vacuole invagination (Uttenweiler et al, 2005). It is a 16-kDa protein that in yeast binds three calcium ions via EF hands and simultaneously undergoes a dramatic conformational change (Luan et al, 1987;Matsuura et al, 1991;Starovasnik et al, 1993;Yazawa et al, 1999).…”

mentioning

confidence: 99%

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The Vacuolar Transporter Chaperone (VTC) Complex Is Required for Microautophagy

Uttenweiler

Schwarz

Neumann

et al. 2007

MBoC

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107

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Microautophagy involves direct invagination and fission of the vacuolar/lysosomal membrane under nutrient limitation.This occurs by an autophagic tube, a specialized vacuolar membrane invagination that pinches off vesicles into the vacuolar lumen. In this study we have identified the VTC (vacuolar transporter chaperone) complex as required for microautophagy. The VTC complex is present on the ER and vacuoles and at the cell periphery. On induction of autophagy by nutrient limitation the VTC complex is recruited to and concentrated on vacuoles. The VTC complex is inhomogeneously distributed within the vacuolar membranes, showing an enrichment on autophagic tubes. Deletion of the VTC complex blocks microautophagic uptake into vacuoles. The mutants still form autophagic tubes but the production of microautophagic vesicles from their tips is impaired. In line with this, affinity-purified antibodies to the Vtc proteins inhibit microautophagic uptake in a reconstituted system in vitro. Our data suggest that the VTC complex is an important constituent of autophagic tubes and that it is required for scission of microautophagic vesicles from these tubes. INTRODUCTIONAutophagy occurs in all eukaryotic cells (Reggiori and Klionsky, 2002). In yeast it has mainly been characterized as an adaptation to nutrient stress such as nitrogen limitation: during autophagy large portions of cytosolic and membraneous material are delivered to the lysosome (in yeast called vacuole) for degradation and recycling (Takeshige et al., 1992). This phenomenon enables cells to survive long periods of starvation. However, in higher eukaryotes autophagy also plays an important role in developmental changes (Levine and Klionsky, 2004), regulation of lifespan (Bergamini et al., 2003;Longo and Finch, 2003;Melendez et al., 2003;Vellai et al., 2003), cancer (Qu et al., 2003;Yue et al., 2003;Gozuacik and Kimchi, 2004), and in neurodegenerative disorders like Huntington's, Parkinson's, or Alzheimer's disease (Yuan et al., 2003). It is also part of the innate immune system assisting in eliminating intracellular pathogens after infection (Gutierrez et al., 2004;Nakagawa et al., 2004;Ogawa et al., 2005).Macroautophagy in yeast is defined as the uptake of cytosolic contents by fusion of double-layered vesicles (autophagosomes) with vacuoles (Baba et al., 1994). Autophagosomes originate from a preautophagosomal structure (PAS) in the vicinity of the vacuole and, during their formation, enwrap portions of cytosol. Fusion of the outer autophagosomal layer with the vacuolar membrane liberates autophagic bodies (single-layered intravacuolar vesicles) into the vacuolar lumen for degradation (Takeshige et al., 1992). Relevant actors of macroautophagy (Atg proteins; Klionsky et al., 2003) have been identified, mainly by genetic screens (Tsukada and Ohsumi, 1993;Thumm et al., 1994;Harding et al., 1995Harding et al., , 1996Titorenko et al., 1995) and have been studied intensively over the last decade.Little is known about microautophagy, a process consisting of a di...

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Section: Vtc Complex In Microautophagysupporting

confidence: 76%

supporting

confidence: 61%

Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

mentioning

confidence: 99%

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