Browsing by Author "Cheshenko, Natalia"

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    Herpes simplex type 2 virus deleted in glycoprotein D protects against vaginal, skin and neural disease
    (2015) Petro, Christopher; González Muñoz, Pablo Alberto; Cheshenko, Natalia; Jandl, Thomas; Khajoueinejad, Nazanin; Benard, Angele; Sengupta, Mayami; Herold, Betsy C.; Jacobs, William R.
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    Herpes Simplex Virus Type 2 Glycoprotein H Interacts with Integrin αvβ3 To Facilitate Viral Entry and Calcium Signaling in Human Genital Tract Epithelial Cells
    (2014) Cheshenko, Natalia; Trepanier, Janie B.; Gonzalez, Pablo A.; Eugenin, Eliseo A.; Jacobs, William R., Jr.; Herold, Betsy C.
    Herpes simplex virus (HSV) entry requires multiple interactions at the cell surface and activation of a complex calcium signaling cascade. Previous studies demonstrated that integrins participate in this process, but their precise role has not been determined. These studies were designed to test the hypothesis that integrin alpha v beta 3 signaling promotes the release of intracellular calcium (Ca2+) stores and contributes to viral entry and cell-to-cell spread. Transfection of cells with small interfering RNA (siRNA) targeting integrin alpha v beta 3, but not other integrin subunits, or treatment with cilengitide, an Arg-Gly-Asp (RGD) mimetic, impaired HSV-induced Ca2+ release, viral entry, plaque formation, and cell-to-cell spread of HSV-1 and HSV-2 in human cervical and primary genital tract epithelial cells. Coimmunoprecipitation studies and proximity ligation assays indicated that integrin alpha v beta 3 interacts with glycoprotein H (gH). An HSV-2 gH-null virus was engineered to further assess the role of gH in the virus-induced signaling cascade. The gH-2-null virus bound to cells and activated Akt to induce a small Ca2+ response at the plasma membrane, but it failed to trigger the release of cytoplasmic Ca2+ stores and was impaired for entry and cell-to-cell spread. Silencing of integrin alpha v beta 3 and deletion of gH prevented phosphorylation of focal adhesion kinase (FAK) and the transport of viral capsids to the nuclear pore. Together, these findings demonstrate that integrin signaling is activated downstream of virus-induced Akt signaling and facilitates viral entry through interactions with gH by activating the release of intracellular Ca2+ and FAK phosphorylation. These findings suggest a new target for HSV treatment and suppression.
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    HSV activates Akt to trigger calcium release and promote viral entry: novel candidate target for treatment and suppression
    (2013) Cheshenko, Natalia; Trepanier, Janie B.; Stefanidou, Martha; Buckley, Niall; Gonzalez, Pablo; Jacobs, William; Herold, Betsy C.
    HSV triggers intracellular calcium release to promote viral entry. We hypothesized that Akt signaling induces the calcium responses and contributes to HSV entry. Exposure of human cervical and primary genital tract epithelial, neuronal, or keratinocyte cells to HSV serotype 2 resulted in rapid phosphorylation of Akt. Silencing of Akt with small interfering RNA prevented the calcium responses, blocked viral entry, and inhibited plaque formation by 90% compared to control siRNA. Susceptibility to infection was partially restored if Akt was reintroduced into silenced cells with an Akt-expressing plasmid. HSV-2 variants deleted in glycoproteins B or D failed to induce Akt phosphorylation, and coimmunoprecipitation studies indicated that Akt interacts with glycoprotein B. Cell-surface expression of Akt was rapidly induced in response to HSV exposure. Miltefosine (50 M), a licensed drug that blocks Akt phosphorylation, inhibited HSV-induced calcium release, viral entry, and plaque formation following infection with acyclovir-sensitive and resistant clinical isolates. Miltefosine blocked amplification of HSV from explanted ganglia to epithelial cells; viral yields were significantly less in miltefosine compared to control-treated cocultures (P<0.01). Together, these findings identify a novel role for Akt in viral entry, link Akt and calcium signaling, and suggest a new target for HSV treatment and suppression.Cheshenko, N., Trepanier, J. B., Stefanidou, M., Buckley, N., Gonzalez, P., Jacobs, W., and Herold, B. C. HSV activates Akt to trigger calcium release and promote viral entry: novel candidate target for treatment and suppression.