Monovalent single-chain Fv fragments and bivalent miniantibodies bound to vesicular stomatitis virus protect against lethal infection

Mouse serum interferons induced by polyI:C, vesicular stomatitis virus (VSV), reovirus, and Mengo virus were assayed in monolayers of mouse L-929 cells by the plaque-reduction method using both VSV and Mengo as challenge viruses. Titers obtained with Mengo virus as challenge were all lower than with VSV. With the interferons induced by VSV, reovirus, and polyI:C, the reductions were of the order of two- to three-fold. With Mengo virus-induced interferon the reduction was much greater (about 17-fold). This offers an explanation for the observation that, unit for unit (measured by the plaque reduction of VSV), Mengo virus-induced interferon is only about [Formula: see text] as effective as polyI:C-induced interferon in protecting mice against lethal infection with Mengo virus. The data are consistent with the hypothesis that an interferon antagonist is produced in the serum of mice infected with Mengo virus. This antagonist, which is not produced in mice inoculated with polyI:C, or reovirus, effectively blocks the antiviral action of interferon during Mengo virus infections, both in vivo and in vitro.

Download Full-text

The Methyltransferase Region of Vesicular Stomatitis Virus L Polymerase Is a Target Site for Functional Intramolecular Insertion

Viruses ◽

10.3390/v11110989 ◽

2019 ◽

Vol 11 (11) ◽

pp. 989

Author(s):

Emmanuel Heilmann ◽

Janine Kimpel ◽

Stephan Geley ◽

Andreas Naschberger ◽

Carles Urbiola ◽

...

Keyword(s):

Vesicular Stomatitis Virus ◽

Fluorescent Protein ◽

Fluorescent Proteins ◽

Insertion Site ◽

Polymerase Activity ◽

Temperature Sensitive ◽

Single Chain ◽

Intracellular Location ◽

L Protein ◽

Vesicular Stomatitis

The L-protein of vesicular stomatitis virus (VSV) is a single-chain multi-domain RNA-dependent RNA polymerase. Previously reported attempts of intramolecular insertions of fluorescent proteins into the L-protein resulted in temperature-sensitive and highly attenuated polymerase activity. Here, we describe a novel insertion site that was selected based on in silico prediction. Of five preselected locations, insertion of the fluorescent protein mCherry in the VSV polymerase between amino acids 1620 and 1621 preserved polymerase function even after extended passaging and showed only mild attenuation compared to wildtype VSV polymerase. High magnification fluorescence imaging revealed a corpuscular cytosolic pattern for the L-protein. To confirm that the insertion site tolerates inclusion of proteins others than mCherry, we cloned mWasabi into the same position in L, generating a VSV-LmWasabi, which was also functional. We also generated a functional dual-color-dual-insertion VSV construct with intramolecularly labeled P and L-proteins. Together, our data present an approach to tag VSV polymerase intramolecularly without perturbing enzymatic activity. This L fusion protein might enable future tracing studies to monitor intracellular location of the VSV transcription and replication machinery in real-time life-imaging studies.

Download Full-text

Non-Lethal Infection of Aminergic Reticular Core Neurons: Age-Dependent Spread of ts Mutant Vesicular Stomatitis Virus from the Nose

Journal of Neuropathology & Experimental Neurology ◽

10.1097/00005072-198809000-00001 ◽

1988 ◽

Vol 47 (5) ◽

pp. 497-506 ◽

Cited By ~ 28

Author(s):

Barbro Lundh ◽

Arthur Löve ◽

Krister Kristensson ◽

Erling Norrby

Keyword(s):

Vesicular Stomatitis Virus ◽

Lethal Infection ◽

Age Dependent ◽

Ts Mutant ◽

Vesicular Stomatitis

Download Full-text

Recombinant vesicular stomatitis virus-based dengue-2 vaccine candidate induces humoral response and protects mice against lethal infection

Human Vaccines & Immunotherapeutics ◽

10.1080/21645515.2016.1183857 ◽

2016 ◽

Vol 12 (9) ◽

pp. 2327-2333 ◽

Cited By ~ 5

Author(s):

Flavio Lauretti ◽

Anasuya Chattopadhyay ◽

Rafael Freitas de Oliveira França ◽

Luiza Castro-Jorge ◽

John Rose ◽

...

Keyword(s):

Vesicular Stomatitis Virus ◽

Vaccine Candidate ◽

Humoral Response ◽

Lethal Infection ◽

Vesicular Stomatitis

Download Full-text

Selection of single-chain antibodies that specifically interact with vesicular stomatitis virus (VSV) nucleocapsid and inhibit viral RNA synthesis

Journal of Virological Methods ◽

10.1016/j.jviromet.2005.06.021 ◽

2006 ◽

Vol 131 (1) ◽

pp. 16-20 ◽

Cited By ~ 4

Author(s):

Jean-Claude Cortay ◽

Denis Gerlier ◽

Frédéric Iseni

Keyword(s):

Vesicular Stomatitis Virus ◽

Rna Synthesis ◽

Viral Rna ◽

Single Chain ◽

Single Chain Antibodies ◽

Vesicular Stomatitis ◽

Selection Of

Download Full-text

Rapid Adaptation of a Recombinant Vesicular Stomatitis Virus to a Targeted Cell Line

Journal of Virology ◽

10.1128/jvi.00142-06 ◽

2006 ◽

Vol 80 (17) ◽

pp. 8603-8612 ◽

Cited By ~ 22

Author(s):

Yanhua Gao ◽

Patricia Whitaker-Dowling ◽

Simon C. Watkins ◽

Judith A. Griffin ◽

Ira Bergman

Keyword(s):

Vesicular Stomatitis Virus ◽

Burst Size ◽

Gene Segment ◽

Serial Passage ◽

Viral Envelope ◽

Single Chain ◽

Single Chain Antibody ◽

Viral Yield ◽

Vesicular Stomatitis

ABSTRACT Vesicular stomatitis virus (VSV) is being developed for cancer therapy. We created a recombinant replicating VSV (rrVSV) that preferentially infected Her2/neu-expressing breast cancer cells. This rrVSV did not express the native VSV-G glycoprotein (gp). Instead, it expressed a chimeric Sindbis gp which included a single-chain antibody (SCA) directed to the human Her2/neu receptor. The virus infected mouse mammary carcinoma cells (D2F2/E2) expressing Her2/neu 23-fold better than the parent cells (D2F2). However, viral growth in cultured D2F2/E2 cells was curtailed after several cycles, and viral yield was very poor at 2 × 104 infectious doses (ID)/ml. We performed in vitro serial passage in D2F2/E2 cells to evolve a virus with improved growth that could be used for preclinical therapy trials in mice. Fifteen passes generated an adapted virus that progressed through multiple cycles in cultured D2F2/E2 cells until all cells were infected and had a viral yield of 1 × 108 ID/ml. Sequencing of the entire viral genomes found only 2 mutations in the adapted virus. Both mutations occurred in the gp gene segment coding for the SCA. An additional N-glycosylation site was created by one of the mutations. The adapted virus showed higher density of gp on the viral envelope, improved infectivity, much greater stability, higher burst size, and decreased induction of cellular interferon. The specificity for cells expressing the Her2/neu receptor was unchanged. These studies demonstrate that serial passage can be used to rapidly evolve a VSV genome encoding an improved chimeric glycoprotein.

Download Full-text