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Cellular proteins associated with the interior and exterior of vesicular stomatitis virus virions.

Moerdyk-Schauwecker M, Hwang SI, Grdzelishvili VZ - PLoS ONE (2014)

Bottom Line: Most of these proteins have not been previously shown to be associated with VSV.Functional enrichment analysis indicated the most overrepresented categories were proteins associated with vesicles, vesicle-mediated transport and protein localization.Our study provides a valuable inventory of virion-associated host proteins for further investigation of their roles in the replication cycle, pathogenesis and immunoreactivity of VSV.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences, University of North Carolina at Charlotte, Charlotte, North Carolina, United States of America; Center for Biomedical Engineering and Science, University of North Carolina at Charlotte, Charlotte, North Carolina, United States of America.

ABSTRACT
Virus particles (virions) often contain not only virus-encoded but also host-encoded proteins. Some of these host proteins are enclosed within the virion structure, while others, in the case of enveloped viruses, are embedded in the host-derived membrane. While many of these host protein incorporations are likely accidental, some may play a role in virus infectivity, replication and/or immunoreactivity in the next host. Host protein incorporations may be especially important in therapeutic applications where large numbers of virus particles are administered. Vesicular stomatitis virus (VSV) is the prototypic rhabdovirus and a candidate vaccine, gene therapy and oncolytic vector. Using mass spectrometry, we previously examined cell type dependent host protein content of VSV virions using intact ("whole") virions purified from three cell lines originating from different species. Here we aimed to determine the localization of host proteins within the VSV virions by analyzing: i) whole VSV virions; and ii) whole VSV virions treated with Proteinase K to remove all proteins outside the viral envelope. A total of 257 proteins were identified, with 181 identified in whole virions and 183 identified in Proteinase K treated virions. Most of these proteins have not been previously shown to be associated with VSV. Functional enrichment analysis indicated the most overrepresented categories were proteins associated with vesicles, vesicle-mediated transport and protein localization. Using western blotting, the presence of several host proteins, including some not previously shown in association with VSV (such as Yes1, Prl1 and Ddx3y), was confirmed and their relative quantities in various virion fractions determined. Our study provides a valuable inventory of virion-associated host proteins for further investigation of their roles in the replication cycle, pathogenesis and immunoreactivity of VSV.

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Related in: MedlinePlus

Overview of sample preparation and experimental approach.Mass spectrometry based analysis of host protein content was conducted on (a) purified whole virions and (b) purified virions treated with proteinase K to remove surface proteins. G, glycoprotein; M, matrix protein; P, phosphoprotein; L, large polymerase protein; N, nucleocapsid protein; yellow shapes, host proteins.
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pone-0104688-g002: Overview of sample preparation and experimental approach.Mass spectrometry based analysis of host protein content was conducted on (a) purified whole virions and (b) purified virions treated with proteinase K to remove surface proteins. G, glycoprotein; M, matrix protein; P, phosphoprotein; L, large polymerase protein; N, nucleocapsid protein; yellow shapes, host proteins.

Mentions: A portion of these whole virions was used directly for proteomic analysis and additional confirmation assays, while the other portion was treated with ProK and re-purified (Fig. 2). ProK treatment cleaves all proteins on the exterior of the virions as well as the extracellular domain of all transmembrane proteins. However, it cannot penetrate the viral envelope, leaving all VSV proteins except for G intact, as well as all host proteins contained within the virion and the transmembrane and cytoplasmic domains of the host membrane proteins located in the viral envelope. This treatment also tends to alter the density of any remaining cell derived vesicles relative to the treated virions, facilitating their removal during the subsequent repurification step [60]. However, proteins on the interior of any residual cellular vesicles would still be detectable.


Cellular proteins associated with the interior and exterior of vesicular stomatitis virus virions.

Moerdyk-Schauwecker M, Hwang SI, Grdzelishvili VZ - PLoS ONE (2014)

Overview of sample preparation and experimental approach.Mass spectrometry based analysis of host protein content was conducted on (a) purified whole virions and (b) purified virions treated with proteinase K to remove surface proteins. G, glycoprotein; M, matrix protein; P, phosphoprotein; L, large polymerase protein; N, nucleocapsid protein; yellow shapes, host proteins.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4126742&req=5

pone-0104688-g002: Overview of sample preparation and experimental approach.Mass spectrometry based analysis of host protein content was conducted on (a) purified whole virions and (b) purified virions treated with proteinase K to remove surface proteins. G, glycoprotein; M, matrix protein; P, phosphoprotein; L, large polymerase protein; N, nucleocapsid protein; yellow shapes, host proteins.
Mentions: A portion of these whole virions was used directly for proteomic analysis and additional confirmation assays, while the other portion was treated with ProK and re-purified (Fig. 2). ProK treatment cleaves all proteins on the exterior of the virions as well as the extracellular domain of all transmembrane proteins. However, it cannot penetrate the viral envelope, leaving all VSV proteins except for G intact, as well as all host proteins contained within the virion and the transmembrane and cytoplasmic domains of the host membrane proteins located in the viral envelope. This treatment also tends to alter the density of any remaining cell derived vesicles relative to the treated virions, facilitating their removal during the subsequent repurification step [60]. However, proteins on the interior of any residual cellular vesicles would still be detectable.

Bottom Line: Most of these proteins have not been previously shown to be associated with VSV.Functional enrichment analysis indicated the most overrepresented categories were proteins associated with vesicles, vesicle-mediated transport and protein localization.Our study provides a valuable inventory of virion-associated host proteins for further investigation of their roles in the replication cycle, pathogenesis and immunoreactivity of VSV.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences, University of North Carolina at Charlotte, Charlotte, North Carolina, United States of America; Center for Biomedical Engineering and Science, University of North Carolina at Charlotte, Charlotte, North Carolina, United States of America.

ABSTRACT
Virus particles (virions) often contain not only virus-encoded but also host-encoded proteins. Some of these host proteins are enclosed within the virion structure, while others, in the case of enveloped viruses, are embedded in the host-derived membrane. While many of these host protein incorporations are likely accidental, some may play a role in virus infectivity, replication and/or immunoreactivity in the next host. Host protein incorporations may be especially important in therapeutic applications where large numbers of virus particles are administered. Vesicular stomatitis virus (VSV) is the prototypic rhabdovirus and a candidate vaccine, gene therapy and oncolytic vector. Using mass spectrometry, we previously examined cell type dependent host protein content of VSV virions using intact ("whole") virions purified from three cell lines originating from different species. Here we aimed to determine the localization of host proteins within the VSV virions by analyzing: i) whole VSV virions; and ii) whole VSV virions treated with Proteinase K to remove all proteins outside the viral envelope. A total of 257 proteins were identified, with 181 identified in whole virions and 183 identified in Proteinase K treated virions. Most of these proteins have not been previously shown to be associated with VSV. Functional enrichment analysis indicated the most overrepresented categories were proteins associated with vesicles, vesicle-mediated transport and protein localization. Using western blotting, the presence of several host proteins, including some not previously shown in association with VSV (such as Yes1, Prl1 and Ddx3y), was confirmed and their relative quantities in various virion fractions determined. Our study provides a valuable inventory of virion-associated host proteins for further investigation of their roles in the replication cycle, pathogenesis and immunoreactivity of VSV.

Show MeSH
Related in: MedlinePlus