Lipid interaction of the C terminus and association of the transmembrane segments facilitate atlastin-mediated homotypic endoplasmic reticulum fusion

Liu, Tina Y.; Bian, Xin; Sun, Sha; Hu, Xiaoyu; Klemm, Robin W.; Prinz, William A.; Rapoport, Tom A.; Hu, Junjie

doi:10.1073/pnas.1208385109

Cited by 104 publications

(158 citation statements)

References 49 publications

(68 reference statements)

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“…Fusion may require the cooperation of multiple molecules, because each dimer only forms transiently during GTP hydrolysis; as one dimer dissociates, the presence of other dimers forming nearby would be needed to keep the vesicles tethered. Our results and recent data reported by others (25) also show that tethering of membranes is insufficient to cause membrane fusion; whereas tethering occurs with just the cytoplasmic domains of ATL, fusion requires the native TM segments and is facilitated by an amphipathic helix in the cytosolic C-terminal tail (13,24).…”

Section: Discussionsupporting

confidence: 71%

“…With WT ATL molecules, the density at the site of tethering and fusion is further increased by their nucleotideindependent association through their TM segments (24). Fusion may require the cooperation of multiple molecules, because each dimer only forms transiently during GTP hydrolysis; as one dimer dissociates, the presence of other dimers forming nearby would be needed to keep the vesicles tethered.…”

Section: Discussionmentioning

confidence: 99%

“…WT ATL or ATL-Sec61β was reconstituted into preformed liposomes with Texas Red-DPPE or Oregon Green-DPPE (Invitrogen), as described previously (24). The vesicles were mixed 1:1 in a reaction buffer containing 5 mM MgCl 2 .…”

Section: Methodsmentioning

confidence: 99%

“…Drosophila ATL, ATL-Sec61β, or cyt-TM2 were reconstituted into preformed liposomes as described previously (24). CytATLHis was immobilized on liposomes containing Ni 2+ -NTA lipids.…”

Section: Methodsmentioning

confidence: 99%

“…Both mutants were indeed defective in membrane fusion, as measured by a lipid-mixing assay (SI Appendix, Fig. S4) (24); however, both were active in membrane tethering, as demonstrated by dynamic light scattering (Fig. 3B).…”

Section: Gtp Hydrolysismentioning

confidence: 99%

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Cis and trans interactions between atlastin molecules during membrane fusion

Liu

Bian

Romano

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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115

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Atlastin (ATL), a membrane-anchored GTPase that mediates homotypic fusion of endoplasmic reticulum (ER) membranes, is required for formation of the tubular network of the peripheral ER. How exactly ATL mediates membrane fusion is only poorly understood. Here we show that fusion is preceded by the transient tethering of ATL-containing vesicles caused by the dimerization of ATL molecules in opposing membranes. Tethering requires GTP hydrolysis, not just GTP binding, because the two ATL molecules are pulled together most strongly in the transition state of GTP hydrolysis. Most tethering events are futile, so that multiple rounds of GTP hydrolysis are required for successful fusion. Supported lipid bilayer experiments show that ATL molecules sitting on the same (cis) membrane can also undergo nucleotide-dependent dimerization. These results suggest that GTP hydrolysis is required to dissociate cis dimers, generating a pool of ATL monomers that can dimerize with molecules on a different (trans) membrane. In addition, tethering and fusion require the cooperation of multiple ATL molecules in each membrane. We propose a comprehensive model for ATL-mediated fusion that takes into account futile tethering and competition between cis and trans interactions. T he endoplasmic reticulum (ER) consists of tubules and sheets connected into a characteristic network by membrane fusion (1-3). In contrast to heterotypic fusion between viral and cellular membranes or between intracellular transport vesicles and target membranes, the fusing ER membranes are identical (i.e., homotypic fusion). The mechanism of heterotypic fusion has been studied extensively, leading to the concept that a conformational change of a viral fusion protein or the assembly of SNARE proteins pulls two opposing membranes together so that they can fuse (4-7). Recently, some insight has been obtained into homotypic ER fusion as well. This process is mediated by the atlastins (ATLs) in metazoans and by Sey1p/ROOT HAIR DEFECTIVE3 (RHD3)-related proteins in yeast and plants (8,9).The ATLs and Sey1p/RHD3 are membrane-bound GTPases that belong to the dynamin family (reviewed in refs. 10, 11). They contain cytosolic N-terminal GTPase (G) and helical bundle domains, followed by two closely spaced transmembrane (TM) segments and a cytosolic C-terminal tail (12)(13)(14). A role for the GTPases in ER fusion is suggested by the observation that their depletion or mutational inactivation leads to long, nonbranched tubules or fragmented ER (8, 9). Nonbranched ER tubules are also observed on expression of dominant-negative ATL mutants (8,12). In addition, the fusion of ER vesicles in Xenopus laevis egg extracts is prevented by ATL antibodies or a cytosolic fragment of ATL (8,15), and the fusion of the ER in mating yeast cells is delayed on SEY1 deletion (16). Most convincingly, proteoliposomes containing purified ATL, Sey1p, or RHD3 undergo GTP-dependent fusion in vitro (9,13,16,17). Defects in ATL-mediated ER fusion can cause hereditary spastic paraplegia, a neurodegenerativ...

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Section: Discussionsupporting

confidence: 71%

Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

“…Drosophila ATL, ATL-Sec61β, or cyt-TM2 were reconstituted into preformed liposomes as described previously (24). CytATLHis was immobilized on liposomes containing Ni 2+ -NTA lipids.…”

Section: Methodsmentioning

confidence: 99%

Section: Gtp Hydrolysismentioning

confidence: 99%

See 3 more Smart Citations

Cis and trans interactions between atlastin molecules during membrane fusion

Liu

Bian

Romano

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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115

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In VivoAnalysis of Membrane Fusion

Palfreyman

Jørgensen

2015

Encyclopedia of Life Sciences

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Membranes provide a barrier that allows chemical reactions to be isolated from the environment. The plasma membrane, for example, delineates self from nonself, and thus must have played an essential role in the evolution of life. Yet under numerous circumstances it is equally important that membranes be breached. Numerous forces oppose the spontaneous fusion of membranes; thus, specialized proteins have evolved to fuse membranes. The most well-understood fusion proteins are the viral fusion proteins and the SNARE proteins used in the secretory pathway. In addition, recent discoveries have lead to models for the fusion of organelles such as mitochondria and peroxisomes, as well as for cell-cell fusion. Despite the diverse structures of fusion proteins, it is possible that they function to drive membranes through a series of common lipid intermediates. Here we review the mechanisms of fusion for biological membranes, and highlight the similarities and differences in these processes.

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Invited review: Mechanisms of GTP hydrolysis and conformational transitions in the dynamin superfamily

2016

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Dynamin superfamily proteins are multidomain mechano‐chemical GTPases which are implicated in nucleotide‐dependent membrane remodeling events. A prominent feature of these proteins is their assembly‐ stimulated mechanism of GTP hydrolysis. The molecular basis for this reaction has been initially clarified for the dynamin‐related guanylate binding protein 1 (GBP1) and involves the transient dimerization of the GTPase domains in a parallel head‐to‐head fashion. A catalytic arginine finger from the phosphate binding (P‐) loop is repositioned toward the nucleotide of the same molecule to stabilize the transition state of GTP hydrolysis. Dynamin uses a related dimerization‐dependent mechanism, but instead of the catalytic arginine, a monovalent cation is involved in catalysis. Still another variation of the GTP hydrolysis mechanism has been revealed for the dynamin‐like Irga6 which bears a glycine at the corresponding position in the P‐loop. Here, we highlight conserved and divergent features of GTP hydrolysis in dynamin superfamily proteins and show how nucleotide binding and hydrolysis are converted into mechano‐chemical movements. We also describe models how the energy of GTP hydrolysis can be harnessed for diverse membrane remodeling events, such as membrane fission or fusion. © 2016 Wiley Periodicals, Inc. Biopolymers 105: 580–593, 2016.

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Lipid interaction of the C terminus and association of the transmembrane segments facilitate atlastin-mediated homotypic endoplasmic reticulum fusion

Cited by 104 publications

References 49 publications

Cis and trans interactions between atlastin molecules during membrane fusion

Cis and trans interactions between atlastin molecules during membrane fusion

In VivoAnalysis of Membrane Fusion

Invited review: Mechanisms of GTP hydrolysis and conformational transitions in the dynamin superfamily

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