Investigation of the Vesicular Stomatitis Virus Matrix Protein: Uncoating and Assembly

Mire, Chad E.

doi:10.21007/etd.cghs.2008.0213

Cited by 1 publication

(2 citation statements)

References 151 publications

(351 reference statements)

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“…The glycoprotein, G, on the outer membrane of the virus binds the host cell through an unknown receptor (Coil and Miller 2004), is endocytosed by clathrin coated vesicles and transported through the endocytic pathway . Evidence from our lab (Mire 2008), as well as others, suggests the fusion event occurs within the acidic environment of the early endosomes while another group argues transport to late endosomes is required (Le Blanc, Luyet et al 2005). Upon acidification of the endosome, a reversible conformational change in the G protein is triggered.…”

Section: Involvement Of G Protein In Vsv Infectionmentioning

confidence: 63%

“…When the pH drops in the early endosome, VSV G undergoes a conformational change which mediates fusion. At this time, G also mediates the acidification of the virion interior which is suggested to affect the interaction between the membrane bound and the RNP bound M, leading to the release of the RNP into the host cell cytosol for replication (Mire 2008). The encapsidated RNA genome then is used as a template for the RNA-dependent RNA polymerase mediated transcription.…”

Section: Class III Viral Fusion Proteins Fusion Glycoproteins From Tmentioning

confidence: 99%

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Contribution of the Membrane-Proximal Region of the Vesicular Stomatitis Virus Glycoprotein to Host Cell Entry and Membrane Fusion

Matheny¹

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DedicationThis dissertation is dedicated to my late father Edward Matheny, my mother Dana Matheny, and brother Sean Matheny for their unyielding support and belief in me throughout the years.iv Acknowledgements I would like to thank my advisor Michael A. Whitt for the opportunity to work in his lab and the training I received. Also, thanks to my committee members Allen Portner, PhD., Charles J. Russell, Ph.D., Susan E. Senogles, Ph.D., and James Patrick Ryan, Ph.D. for providing excellent insights, ideas, and guidance during my research.My gratitude is also extended to present and past members of the Whitt laboratory. Erika Dillard, Chad Mire Ph.D., Himangi Jayakar Ph.D., Jeetendra Eswararka Ph.D., Makiko Wantannabe Ph.D., and Carolyn Matthews for their support and help. Additionally, I would like to thank other members of the Molecular Sciences department for offering input, guidance, and support.v Abstract Vesicular stomatitis virus (VSV) is an enveloped, nonsegmented, negative-sense, RNA virus belonging to the Rhabdoviridae family. VSV is considered the prototypic Rhabdovirus due to its simple genetic organization, broad host cell tropism, and ability to be easily grown in cell culture. Therefore, VSV has been used as the prototype to study viral entry, transcription, replication, and assembly. Viral entry, a critical step in the lifecycle of the virus, is mediated by the outer surface protein, G and will be the focus of this dissertation.We hypothesize that the highly conserved residues in the membrane-proximal region of VSV G protein are critical to membrane fusion through participation in low pHinduced conformational changes and stability of theses structures. This hypothesis was tested initially through the use of site directed mutagenesis and then by studying rapidly acquired second site mutations. Residues that are conserved completely or conserved for their biochemical properties were either deleted or mutated to alanines. The mutated G proteins were examined through the use of transient transfection assays for their surface expression levels, fusion capacity, and ability to undergo pH dependent conformational changes. Mutation of a conserved HPH motif (H423/P424/H425) resulted in a dramatic decrease in surface expression. ΔH423/P424/H425 was completely fusion defective as assessed by syncytium formation assays that measured cell-cell fusion. Likewise, the cell-cell fusion activity for ΔH425 was affected in that the pH threshold required to trigger the fusion event was decreased. These findings support previous reports suggesting a requirement for histidine protonation in order for the pH dependent conformational changes needed for membrane fusion to occur. Recombinant viruses encoding the mutated G proteins were recovered and all of the viruses expressing G proteins with reduced cell-cell fusion capacity, as compared to wild type, grew to lower titers, with the exception of one mutant: D435A/D436A/E437A. Mutation of the DDE motif resulted in a G protein with a reduced capacity to fuse in transiently t...

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Section: Involvement Of G Protein In Vsv Infectionmentioning

confidence: 63%

Section: Class III Viral Fusion Proteins Fusion Glycoproteins From Tmentioning

confidence: 99%

Contribution of the Membrane-Proximal Region of the Vesicular Stomatitis Virus Glycoprotein to Host Cell Entry and Membrane Fusion

Matheny¹

View full text Add to dashboard Cite

show abstract

Investigation of the Vesicular Stomatitis Virus Matrix Protein: Uncoating and Assembly

Cited by 1 publication

References 151 publications

Contribution of the Membrane-Proximal Region of the Vesicular Stomatitis Virus Glycoprotein to Host Cell Entry and Membrane Fusion

Contribution of the Membrane-Proximal Region of the Vesicular Stomatitis Virus Glycoprotein to Host Cell Entry and Membrane Fusion

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