scholarly journals Reovirus FAST Protein Transmembrane Domains Function in a Modular, Primary Sequence-Independent Manner To Mediate Cell-Cell Membrane Fusion

2009 ◽  
Vol 83 (7) ◽  
pp. 2941-2950 ◽  
Author(s):  
Eileen K. Clancy ◽  
Roy Duncan
Keyword(s):  
Cell Membrane ◽  
Membrane Fusion ◽  
Fusion Proteins ◽  
Transmembrane Domain ◽  
Fusion Reaction ◽  
Membrane Topology ◽  
Fast Proteins ◽  
Signal Anchor ◽  
Reverse Signal ◽  
Cell Cell

ABSTRACT The FAST proteins are a unique family of virus-encoded cell-cell membrane fusion proteins. In the absence of a cleavable N-terminal signal peptide, a single-pass transmembrane domain (TMD) functions as a reverse signal-anchor to direct the FAST proteins into the plasma membrane in an Nexo/Ccyt topology. There is little information available on the role of the FAST protein TMD in the cell-cell membrane fusion reaction. We show that in the absence of conservation in the length or primary amino acid sequence, the p14 TMD can be functionally exchanged with the TMDs of the p10 and p15 FAST proteins. This is not the case for chimeric p14 proteins containing the TMDs of two different enveloped viral fusion proteins or a cellular membrane protein; such chimeric proteins were defective for both pore formation and syncytiogenesis. TMD structural features that are conserved within members of the FAST protein family presumably play direct roles in the fusion reaction. Molecular modeling suggests that the funnel-shaped architecture of the FAST protein TMDs may represent such a conserved structural and functional motif. Interestingly, although heterologous TMDs exert diverse influences on the trafficking of the p14 FAST protein, these TMDs are capable of functioning as reverse signal-anchor sequences to direct p14 into lipid rafts in the correct membrane topology. The FAST protein TMDs are therefore not primary determinants of type III protein topology, but they do play a direct, sequence-independent role in the membrane fusion reaction.

scholarly journals Functional Relevance of the Transmembrane Domain and Cytoplasmic Tail of the Pseudorabies Virus Glycoprotein H for Membrane Fusion

2018 ◽  
Vol 92 (12) ◽  
Author(s):  
Melina Vallbracht ◽  
Walter Fuchs ◽  
Barbara G. Klupp ◽  
Thomas C. Mettenleiter
Keyword(s):  
Membrane Fusion ◽  
Fusion Proteins ◽  
Essential Role ◽  
Pseudorabies Virus ◽  
Transmembrane Domain ◽  
Cytoplasmic Tail ◽  
Class Iii ◽  
Complex Needs ◽  
Hsv 1

ABSTRACTHerpesvirus membrane fusion depends on the core fusion machinery, comprised of glycoproteins B (gB) and gH/gL. Although gB structurally resembles autonomous class III fusion proteins, it strictly depends on gH/gL to drive membrane fusion. Whether the gH/gL complex needs to be membrane anchored to fulfill its function and which role the gH cytoplasmic (CD) and transmembrane domains (TMD) play in fusion is unclear. While the gH CD and TMD play an important role during infection, soluble gH/gL of herpes simplex virus 1 (HSV-1) seems to be sufficient to mediate cell-cell fusion in transient assays, arguing against an essential contribution of the CD and TMD. To shed more light on this apparent discrepancy, we investigated the role of the CD and TMD of the related alphaherpesvirus pseudorabies virus (PrV) gH. For this purpose, we expressed C-terminally truncated and soluble gH and replaced the TMD with a glycosylphosphatidylinositol (gpi) anchor. We also generated chimeras containing the TMD and/or CD of PrV gD or HSV-1 gH. Proteins were characterized in cell-based fusion assays and during virus infection. Although truncation of the CD resulted in decreased membrane fusion activity, the mutant proteins still supported replication of gH-negative PrV, indicating that the PrV gH CD is dispensable for viral replication. In contrast, PrV gH lacking the TMD, membrane-anchored via a lipid linker, or comprising the PrV gD TMD were nonfunctional, highlighting the essential role of the gH TMD for function. Interestingly, despite low sequence identity, the HSV-1 gH TMD could substitute for the PrV gH TMD, pointing to functional conservation.IMPORTANCEEnveloped viruses depend on membrane fusion for virus entry. While this process can be mediated by only one or two proteins, herpesviruses depend on the concerted action of at least three different glycoproteins. Although gB has features of bona fide fusion proteins, it depends on gH and its complex partner, gL, for fusion. Whether gH/gL prevents premature fusion or actively triggers gB-mediated fusion is unclear, and there are contradictory results on whether gH/gL function requires stable membrane anchorage or whether the ectodomains alone are sufficient. Our results show that in pseudorabies virus gH, the transmembrane anchor plays an essential role for gB-mediated fusion while the cytoplasmic tail is not strictly required.

scholarly journals New Biophysical Approaches Reveal the Dynamics and Mechanics of Type I Viral Fusion Machinery and Their Interplay with Membranes

Viruses ◽  
2020 ◽  
Vol 12 (4) ◽  
pp. 413 ◽  
Author(s):  
Mark A. Benhaim ◽  
Kelly K. Lee
Keyword(s):  
Membrane Fusion ◽  
Conformational Changes ◽  
Viral Entry ◽  
Fusion Proteins ◽  
Structural Changes ◽  
Fusion Reaction ◽  
Type I ◽  
Viral Fusion ◽  
Technological Advances ◽  
Viral Fusion Proteins

Protein-mediated membrane fusion is a highly regulated biological process essential for cellular and organismal functions and infection by enveloped viruses. During viral entry the membrane fusion reaction is catalyzed by specialized protein machinery on the viral surface. These viral fusion proteins undergo a series of dramatic structural changes during membrane fusion where they engage, remodel, and ultimately fuse with the host membrane. The structural and dynamic nature of these conformational changes and their impact on the membranes have long-eluded characterization. Recent advances in structural and biophysical methodologies have enabled researchers to directly observe viral fusion proteins as they carry out their functions during membrane fusion. Here we review the structure and function of type I viral fusion proteins and mechanisms of protein-mediated membrane fusion. We highlight how recent technological advances and new biophysical approaches are providing unprecedented new insight into the membrane fusion reaction.

scholarly journals Viral Membrane Fusion and the Transmembrane Domain

Viruses ◽  
2020 ◽  
Vol 12 (7) ◽  
pp. 693 ◽  
Author(s):  
Chelsea T. Barrett ◽  
Rebecca Ellis Dutch
Keyword(s):  
Membrane Fusion ◽  
Host Cell ◽  
Fusion Proteins ◽  
Transmembrane Domain ◽  
Critical Role ◽  
Viral Fusion ◽  
Transmembrane Region ◽  
Host Cell Membrane ◽  
The Past ◽  
Viral Fusion Proteins

Initiation of host cell infection by an enveloped virus requires a viral-to-host cell membrane fusion event. This event is mediated by at least one viral transmembrane glycoprotein, termed the fusion protein, which is a key therapeutic target. Viral fusion proteins have been studied for decades, and numerous critical insights into their function have been elucidated. However, the transmembrane region remains one of the most poorly understood facets of these proteins. In the past ten years, the field has made significant advances in understanding the role of the membrane-spanning region of viral fusion proteins. We summarize developments made in the past decade that have contributed to the understanding of the transmembrane region of viral fusion proteins, highlighting not only their critical role in the membrane fusion process, but further demonstrating their involvement in several aspects of the viral lifecycle.

scholarly journals Correction of the DNA Sequence of theregB Gene of Rhodobacter capsulatus with Implications for the Membrane Topology of the Sensor Kinase RegB

2000 ◽  
Vol 182 (3) ◽  
pp. 818-820 ◽  
Author(s):  
Weizhen Chen ◽  
Andreas Jäger ◽  
Gabriele Klug
Keyword(s):  
Fusion Proteins ◽  
Rhodobacter Capsulatus ◽  
Transmembrane Domain ◽  
Membrane Topology ◽  
Sensor Kinase ◽  
N Terminus ◽  
Topology Model ◽  
Membrane Spanning ◽  
Histidine Sensor Kinase ◽  
Membrane Spanning Domains

ABSTRACT We corrected the previously published sequence for theregB gene, which encodes a histidine sensor kinase inRhodobacter capsulatus. The deduced RegB amino acid sequence has an additional putative transmembrane domain at the N terminus. Analysis of RegB-PhoA and RegB-LacZ fusion proteins supports a topology model for RegB with six membrane-spanning domains.

scholarly journals Fusogenic Reoviruses and Their Fusion-Associated Small Transmembrane (FAST) Proteins

2019 ◽  
Vol 6 (1) ◽  
pp. 341-363 ◽  
Author(s):  
Roy Duncan
Keyword(s):  
Membrane Fusion ◽  
Fusion Proteins ◽  
Syncytium Formation ◽  
Nonstructural Proteins ◽  
Enveloped Virus ◽  
Fast Proteins ◽  
Nonenveloped Virus ◽  
Functional Features ◽  
Membrane Fusion Proteins ◽  
Viral Membrane Fusion

With no limiting membrane surrounding virions, nonenveloped viruses have no need for membrane fusion to gain access to intracellular replication compartments. Consequently, nonenveloped viruses do not encode membrane fusion proteins. The only exception to this dogma is the fusogenic reoviruses that encode fusion-associated small transmembrane (FAST) proteins that induce syncytium formation. FAST proteins are the smallest viral membrane fusion proteins and, unlike their enveloped virus counterparts, are nonstructural proteins that evolved specifically to induce cell-to-cell, not virus-cell, membrane fusion. This distinct evolutionary imperative is reflected in structural and functional features that distinguish this singular family of viral fusogens from all other protein fusogens. These rudimentary fusogens comprise specific combinations of different membrane effector motifs assembled into small, modular membrane fusogens. FAST proteins offer a minimalist model to better understand the ubiquitous process of protein-mediated membrane fusion and to reveal novel mechanisms of nonenveloped virus dissemination that contribute to virulence.

scholarly journals Important Role for the Transmembrane Domain of Severe Acute Respiratory Syndrome Coronavirus Spike Protein during Entry

2006 ◽  
Vol 80 (3) ◽  
pp. 1302-1310 ◽  
Author(s):  
Rene Broer ◽  
Bertrand Boson ◽  
Willy Spaan ◽  
François-Loïc Cosset ◽  
Jeroen Corver
Keyword(s):  
Severe Acute Respiratory Syndrome ◽  
Membrane Fusion ◽  
Cell Fusion ◽  
Transmembrane Domain ◽  
Cytoplasmic Tail ◽  
Transmembrane Domains ◽  
Spike Protein ◽  
Fusion Activity ◽  
Wild Type ◽  
Cell Cell

ABSTRACT The spike protein (S) of severe acute respiratory syndrome coronavirus (SARS-CoV) is responsible for receptor binding and membrane fusion. It contains a highly conserved transmembrane domain that consists of three parts: an N-terminal tryptophan-rich domain, a central domain, and a cysteine-rich C-terminal domain. The cytoplasmic tail of S has previously been shown to be required for assembly. Here, the roles of the transmembrane and cytoplasmic domains of S in the infectivity and membrane fusion activity of SARS-CoV have been studied. SARS-CoV S-pseudotyped retrovirus (SARSpp) was used to measure S-mediated infectivity. In addition, the cell-cell fusion activity of S was monitored by a Renilla luciferase-based cell-cell fusion assay. Svsv-cyt, an S chimera with a cytoplasmic tail derived from vesicular stomatitis virus G protein (VSV-G), and Smhv-tmdcyt, an S chimera with the cytoplasmic and transmembrane domains of mouse hepatitis virus, displayed wild-type-like activity in both assays. Svsv-tmdcyt, a chimera with the cytoplasmic and transmembrane domains of VSV-G, was impaired in the SARSpp and cell-cell fusion assays, showing 3 to 25% activity compared to the wild type, depending on the assay and the cells used. Examination of the oligomeric state of the chimeric S proteins in SARSpp revealed that Svsv-tmdcyt trimers were less stable than wild-type S trimers, possibly explaining the lowered fusogenicity and infectivity.

scholarly journals Palmitoylation, Membrane-Proximal Basic Residues, and Transmembrane Glycine Residues in the Reovirus p10 Protein Are Essential for Syncytium Formation

2003 ◽  
Vol 77 (18) ◽  
pp. 9769-9779 ◽  
Author(s):  
Maya Shmulevitz ◽  
Jayme Salsman ◽  
Roy Duncan
Keyword(s):  
Fusion Protein ◽  
Membrane Fusion ◽  
Transmembrane Domain ◽  
Nonstructural Protein ◽  
Fusion Reaction ◽  
Specific Interaction ◽  
Syncytium Formation ◽  
Surface Expression ◽  
Fusion Activity ◽  
Basic Region

ABSTRACT Avian reovirus and Nelson Bay reovirus are two unusual nonenveloped viruses that induce extensive cell-cell fusion via expression of a small nonstructural protein, termed p10. We investigated the importance of the transmembrane domain, a conserved membrane-proximal dicysteine motif, and an endodomain basic region in the membrane fusion activity of p10. We now show that the p10 dicysteine motif is palmitoylated and that loss of palmitoylation correlates with a loss of fusion activity. Mutational and functional analyses also revealed that a triglycine motif within the transmembrane domain and the membrane-proximal basic region were essential for p10-mediated membrane fusion. Mutations in any of these three motifs did not influence events upstream of syncytium formation, such as p10 membrane association, protein topology, or surface expression, suggesting that these motifs are more intimately associated with the membrane fusion reaction. These results suggest that the rudimentary p10 fusion protein has evolved a mechanism of inducing membrane merger that is highly dependent on the specific interaction of several different motifs with donor membranes. In addition, cross-linking, coimmunoprecipitation, and complementation assays provided no evidence for p10 homo- or heteromultimer formation, suggesting that p10 may be the first example of a membrane fusion protein that does not form stable, higher-order multimers.

scholarly journals Ovine Herpesvirus 2 Glycoproteins B, H, and L Are Sufficient for, and Viral Glycoprotein Ov8 Can Enhance, Cell-Cell Membrane Fusion

2017 ◽  
Vol 91 (6) ◽  
Author(s):  
Salim M. AlHajri ◽  
Cristina W. Cunha ◽  
Anthony V. Nicola ◽  
Hector C. Aguilar ◽  
Hong Li ◽  
...  
Keyword(s):  
Cell Culture ◽  
Cell Membrane ◽  
Membrane Fusion ◽  
Culture System ◽  
Malignant Catarrhal Fever ◽  
Cell Culture System ◽  
Viral Glycoprotein ◽  
Cellular Factors ◽  
Entry Mechanism ◽  
Cell Cell

ABSTRACT Ovine herpesvirus 2 (OvHV-2) is a gammaherpesvirus in the genus Macavirus that is carried asymptomatically by sheep. Infection of poorly adapted animals with OvHV-2 results in sheep-associated malignant catarrhal fever, a fatal disease characterized by lymphoproliferation and vasculitis. There is no treatment or vaccine for the disease and no cell culture system to propagate the virus. The lack of cell culture has hindered studies of OvHV-2 biology, including its entry mechanism. As an alternative method to study OvHV-2 glycoproteins responsible for membrane fusion as a part of the entry mechanism, we developed a virus-free cell-to-cell membrane fusion assay to identify the minimum required OvHV-2 glycoproteins to induce membrane fusion. OvHV-2 glycoproteins B, H, and L (gB, gH, and gL) were able to induce membrane fusion together but not when expressed individually. Additionally, open reading frame Ov8, unique to OvHV-2, was found to encode a transmembrane glycoprotein that can significantly enhance membrane fusion. Thus, OvHV-2 gB, gH, and gL are sufficient to induce membrane fusion, while glycoprotein Ov8 plays an enhancing role by an unknown mechanism. IMPORTANCE Herpesviruses enter cells via attachment of the virion to the cellular surface and fusion of the viral envelope with cellular membranes. Virus-cell membrane fusion is an important step for a successful viral infection. Elucidating the roles of viral glycoproteins responsible for membrane fusion is critical toward understanding viral entry. Entry of ovine herpesvirus 2 (OvHV-2), the causative agent of sheep associated-malignant catarrhal fever, which is one of the leading causes of death in bison and other ungulates, has not been well studied due to the lack of a cell culture system to propagate the virus. The identification of OvHV-2 glycoproteins that mediate membrane fusion may help identify viral and/or cellular factors involved in OvHV-2 cell tropism and will advance investigation of cellular factors necessary for virus-cell membrane fusion. We found that OvHV-2 glycoproteins B, H, and L are sufficient for, and viral glycoprotein Ov8 can significantly enhance, cell-cell membrane fusion.

Sign in / Sign up

Export Citation Format

Share Document