scholarly journals The yeast Batten disease orthologue Btn1 controls endosome–Golgi retrograde transport via SNARE assembly

2011 ◽  
Vol 195 (2) ◽  
pp. 203-215 ◽  
Author(s):  
Rachel Kama ◽  
Vydehi Kanneganti ◽  
Christian Ungermann ◽  
Jeffrey E. Gerst
Keyword(s):  
Disease Gene ◽  
Transmembrane Domain ◽  
Batten Disease ◽  
Retrograde Transport ◽  
Attachment Protein ◽  
Protein Receptor ◽  
Late Endosomes ◽  
Target Membrane ◽  
Opposing Effects ◽  
Protein Attachment

The human Batten disease gene CLN3 and yeast orthologue BTN1 encode proteins of unclear function. We show that the loss of BTN1 phenocopies that of BTN2, which encodes a retromer accessory protein involved in the retrieval of specific cargo from late endosomes (LEs) to the Golgi. However, Btn1 localizes to Golgi and regulates soluble N-ethyl-maleimide sensitive fusion protein attachment protein receptor (SNARE) function to control retrograde transport. Specifically, BTN1 overexpression and deletion have opposing effects on phosphorylation of the Sed5 target membrane SNARE, on Golgi SNARE assembly, and on Golgi integrity. Although Btn1 does not interact physically with SNAREs, it regulates Sed5 phosphorylation by modulating Yck3, a palmitoylated endosomal kinase. This may involve modification of the Yck3 lipid anchor, as substitution with a transmembrane domain suppresses the deletion of BTN1 and restores trafficking. Correspondingly, deletion of YCK3 mimics that of BTN1 or BTN2 with respect to LE–Golgi retrieval. Thus, Btn1 controls retrograde sorting by regulating SNARE phosphorylation and assembly, a process that may be adversely affected in Batten Disease patients.

scholarly journals Multiple ER–Golgi SNARE transmembrane domains are dispensable for trafficking but required for SNARE recycling

2016 ◽  
Vol 27 (17) ◽  
pp. 2633-2641 ◽  
Author(s):  
Li Chen ◽  
Martin S. Y. Lau ◽  
David K. Banfield
Keyword(s):  
Transmembrane Domain ◽  
Retrograde Transport ◽  
Steady State Distribution ◽  
State Distribution ◽  
Attachment Protein ◽  
Protein Receptor ◽  
Sensitive Factor ◽  
Strict Requirement ◽  
Membrane Tether ◽  
Essential Prerequisite

The formation of soluble N-ethylmaleimide–sensitive factor attachment protein receptor (SNARE) complexes between opposing membranes is an essential prerequisite for fusion between vesicles and their target compartments. The composition and length of a SNARE’s transmembrane domain (TMD) is also an indicator for their steady-state distribution in cells. The evolutionary conservation of the SNARE TMD, together with the strict requirement of this feature for membrane fusion in biochemical studies, implies that the TMD represents an essential protein module. Paradoxically, we find that for several essential ER- and Golgi-localized SNAREs, a TMD is unnecessary. Moreover, in the absence of a covalent membrane tether, such SNAREs can still support ER–Golgi vesicle transport and recapitulate established genetic interactions. Transport anomalies appear to be restricted to retrograde trafficking, but these defects are overcome by the attachment of a C-terminal lipid anchor to the SNARE. We conclude that the TMD functions principally to support the recycling of Qb-, Qc-, and R-SNAREs and, in so doing, retrograde transport.

scholarly journals Snarepins Are Functionally Resistant to Disruption by Nsf and αSNAP

2000 ◽  
Vol 149 (5) ◽  
pp. 1063-1072 ◽  
Author(s):  
Thomas Weber ◽  
Francesco Parlati ◽  
James A. McNew ◽  
Robert J. Johnston ◽  
Benedikt Westermann ◽  
...  
Keyword(s):  
Lipid Bilayers ◽  
Snare Proteins ◽  
Attachment Protein ◽  
Receptor Proteins ◽  
Protein Receptor ◽  
The Face ◽  
Target Membrane ◽  
The Moment ◽  
Protein Attachment ◽  
Lines Of Evidence

SNARE (SNAP [soluble NSF {N-ethylmaleimide–sensitive fusion protein} attachment protein] receptor) proteins are required for many fusion processes, and recent studies of isolated SNARE proteins reveal that they are inherently capable of fusing lipid bilayers. Cis-SNARE complexes (formed when vesicle SNAREs [v-SNAREs] and target membrane SNAREs [t-SNAREs] combine in the same membrane) are disrupted by the action of the abundant cytoplasmic ATPase NSF, which is necessary to maintain a supply of uncombined v- and t-SNAREs for fusion in cells. Fusion is mediated by these same SNARE proteins, forming trans-SNARE complexes between membranes. This raises an important question: why doesn't NSF disrupt these SNARE complexes as well, preventing fusion from occurring at all? Here, we report several lines of evidence that demonstrate that SNAREpins (trans-SNARE complexes) are in fact functionally resistant to NSF, and they become so at the moment they form and commit to fusion. This elegant design allows fusion to proceed locally in the face of an overall environment that massively favors SNARE disruption.

Membrane traffic to and from lysosomes.

2005 ◽  
Vol 72 ◽  
pp. 77-86 ◽  
Author(s):  
J. Paul Luzio ◽  
Paul R. Pryor ◽  
Sally R. Gray ◽  
Matthew J. Gratian ◽  
Robert C. Piper ◽  
...  
Keyword(s):  
Endocytic Pathway ◽  
Receptor Protein ◽  
Snare Protein ◽  
Membrane Bilayer ◽  
Attachment Protein ◽  
Protein Receptor ◽  
Late Endosomes ◽  
Recorded Data ◽  
Protein Attachment ◽  
Fusion Hypothesis

In the late endocytic pathway, it has been proposed that endocytosed macromolecules are delivered to a proteolytic environment by 'kiss-and-run' events or direct fusion between late endosomes and lysosomes. To test whether the fusion hypothesis accounts for delivery to lysosomes in living cells, we have used confocal microscopy to examine content mixing between lysosomes loaded with rhodamine-dextran and endosomes subsequently loaded with Oregon-Green-dextran. Both kissing and explosive fusion events were recorded. Data from cell-free content-mixing assays have suggested that fusion is initiated by tethering, which leads to formation of a trans-SNARE (soluble N-ethylmaleimide-sensitive fusion protein attachment protein receptor) protein complex and then release of lumenal Ca2+, followed by membrane bilayer fusion. We have shown that the R-SNARE (arginine-containing SNARE) protein VAMP (vesicle-associated membrane protein) 7 is necessary for heterotypic fusion between late endosomes and lysosomes, whereas a different R-SNARE, VAMP 8 is required for homotypic fusion of late endosomes. After fusion of lysosomes with late endosomes, lysosomes are re-formed from the resultant hybrid organelles, a process requiring condensation of content and the removal/recycling of some membrane proteins.

scholarly journals Syntaxin 7 and VAMP-7 are SolubleN-Ethylmaleimide–sensitive Factor Attachment Protein Receptors Required for Late Endosome–Lysosome and Homotypic Lysosome Fusion in Alveolar Macrophages

2000 ◽  
Vol 11 (7) ◽  
pp. 2327-2333 ◽  
Author(s):  
Diane McVey Ward ◽  
Jonathan Pevsner ◽  
Matthew A. Scullion ◽  
Michael Vaughn ◽  
Jerry Kaplan
Keyword(s):  
Alveolar Macrophages ◽  
Transmembrane Domain ◽  
Late Endosome ◽  
Endocytic Pathway ◽  
Attachment Protein ◽  
Protein Receptor ◽  
Early Endosomes ◽  
Sensitive Factor ◽  
Protein Receptors

Endocytosis in alveolar macrophages can be reversibly inhibited, permitting the isolation of endocytic vesicles at defined stages of maturation. Using an in vitro fusion assay, we determined that each isolated endosome population was capable of homotypic fusion. All vesicle populations were also capable of heterotypic fusion in a temporally specific manner; early endosomes, isolated 4 min after internalization, could fuse with endosomes isolated 8 min after internalization but not with 12-min endosomes or lysosomes. Lysosomes fuse with 12-min endosomes but not with earlier endosomes. Using homogenous populations of endosomes, we have identified Syntaxin 7 as a soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) required for late endosome–lysosome and homotypic lysosome fusion in vitro. A bacterially expressed human Syntaxin 7 lacking the transmembrane domain inhibited homotypic late endosome and lysosome fusion as well as heterotypic late endosome–lysosome fusion. Affinity-purified antibodies directed against Syntaxin 7 also inhibited lysosome fusion in vitro but had no affect on homotypic early endosome fusion. Previous work suggested that human VAMP-7 (vesicle-associated membrane protein-7) was a SNARE required for late endosome–lysosome fusion. A bacterially expressed human VAMP-7 lacking the transmembrane domain inhibited both late endosome–lysosome fusion and homotypic lysosome fusion in vitro. These studies indicate that: 1) fusion along the endocytic pathway is a highly regulated process, and 2) two SNARE molecules, Syntaxin 7 and human VAMP-7, are involved in fusion of vesicles in the late endocytic pathway in alveolar macrophages.

scholarly journals Comparative analysis of tandem C2 domains from the mammalian synaptotagmin family

2004 ◽  
Vol 378 (2) ◽  
pp. 681-686 ◽  
Author(s):  
Colin RICKMAN ◽  
Molly CRAXTON ◽  
Shona OSBORNE ◽  
Bazbek DAVLETOV
Keyword(s):  
Membrane Trafficking ◽  
Sequence Similarity ◽  
Attachment Protein ◽  
C2 Domains ◽  
Phospholipid Binding ◽  
Calcium Dependent ◽  
Functional Studies ◽  
Protein Receptor ◽  
High Degree ◽  
Protein Attachment

Intracellular membrane traffic is governed by a conserved set of proteins, including Syts (synaptotagmins). The mammalian Syt family includes 15 isoforms. Syts are membrane proteins that possess tandem C2 domains (C2AB) implicated in calcium-dependent phospholipid binding. We performed a pair-wise amino acid sequence comparison, together with functional studies of rat Syt C2ABs, to examine common and divergent properties within the mammalian family. Sequence analysis indicates three different C2AB classes, the members of which share a high degree of sequence similarity. All the other C2ABs are highly divergent in sequence. Nearly half of the Syt family does not exhibit calcium/phospholipid binding in comparison to Syt I, the major brain isoform. Syts do, however, possess a more conserved function, namely calcium-independent binding to target SNARE (soluble N-ethylmaleimide-sensitive fusion protein attachment protein receptor) heterodimers. All tested isoforms, except Syt XII and Syt XIII, bound the target SNARE heterodimer comprising syntaxin 1 and SNAP-25 (25 kDa synaptosome-associated protein). Our present study suggests that many Syt isoforms can function in membrane trafficking to interact with the target SNARE heterodimer on the pathway to calcium-triggered membrane fusion.

The molecular mechanisms of the mammalian exocyst complex in exocytosis

2006 ◽  
Vol 34 (5) ◽  
pp. 687-690 ◽  
Author(s):  
S. Wang ◽  
S.C. Hsu
Keyword(s):  
Plasma Membrane ◽  
Molecular Mechanisms ◽  
Early Stage ◽  
Secretory Vesicles ◽  
Attachment Protein ◽  
Trafficking Pathway ◽  
Exocyst Complex ◽  
Protein Receptor ◽  
Associated Proteins ◽  
Protein Attachment

Exocytosis is a highly ordered vesicle trafficking pathway that targets proteins to the plasma membrane for membrane addition or secretion. Research over the years has discovered many proteins that participate at various stages in the mammalian exocytotic pathway. At the early stage of exocytosis, co-atomer proteins and their respective adaptors and GTPases have been shown to play a role in the sorting and incorporation of proteins into secretory vesicles. At the final stage of exocytosis, SNAREs (soluble N-ethylmaleimide-sensitive fusion protein attachment protein receptor) and SNARE-associated proteins are believed to mediate the fusion of secretory vesicles at the plasma membrane. There are multiple events that may occur between the budding of secretory vesicles from the Golgi and the fusion of these vesicles at the plasma membrane. The most obvious and best-known event is the transport of secretory vesicles from Golgi to the vicinity of the plasma membrane via microtubules and their associated motors. At the vicinity of the plasma membrane, however, it is not clear how vesicles finally dock and fuse with the plasma membrane. Identification of proteins involved in these events should provide important insights into the mechanisms of this little known stage of the exocytotic pathway. Currently, a protein complex, known as the sec6/8 or the exocyst complex, has been implicated to play a role at this late stage of exocytosis.

Sec22b-dependent assembly of endoplasmic reticulum Q-SNARE proteins

2008 ◽  
Vol 410 (1) ◽  
pp. 93-100 ◽  
Author(s):  
Takehiro Aoki ◽  
Masaki Kojima ◽  
Katsuko Tani ◽  
Mitsuo Tagaya
Keyword(s):  
Endoplasmic Reticulum ◽  
Membrane Fusion ◽  
Transmembrane Domain ◽  
Cd Spectroscopy ◽  
Snare Proteins ◽  
Peripheral Membrane Proteins ◽  
Protein Receptor ◽  
Snare Motif ◽  
Α Helix ◽  
Protein Attachment

SNARE (soluble N-ethylmaleimide-sensitive fusion protein-attachment protein receptor) proteins involved in membrane fusion usually contain a conserved α-helix (SNARE motif) that is flanked by a C-terminal transmembrane domain. They can be classified into Q-SNARE and R-SNARE based on the structural property of their motifs. Assembly of four SNARE motifs (Qa, b, c and R) is supposed to trigger membrane fusion. We have previously shown that ER (endoplasmic reticulum)-localized syntaxin 18 (Qa) forms a complex with BNIP1 (Qb), p31/Use1 (Qc), Sec22b (R) and several peripheral membrane proteins. In the present study, we examined the interaction of syntaxin 18 with other SNAREs using pulldown assays and CD spectroscopy. We found that the association of syntaxin 18 with Sec22b induces an increase in α-helicity of their SNARE motifs, which results in the formation of high-affinity binding sites for BNIP1 and p31. This R-SNARE-dependent Q-SNARE assembly is quite different from the assembly mechanisms of SNAREs localized in organelles other than the ER. The implication of the mechanism of ER SNARE assembly is discussed in the context of the physiological roles of the syntaxin 18 complex.

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