Flaviviruses dysregulate HSP90 to inhibit JAK/STAT signaling; a mechanism contributing to ZIKV-induced microencephaly.

Identification: Roby, Justin


Flaviviruses dysregulate HSP90 to inhibit JAK/STAT signaling; a mechanism contributing to ZIKV-induced microencephaly
Roby, Justin A1; Esser-Nobis, Katharina1; Dewey, Elyse C1; Fairgrieve, Marian RJ1; Aarreberg, Lauren D1; Hemann, Emily A1; Schwerk, Johannes1; Lu, Yun Hsuan1; Stokes, Caleb A1; Soveg, Frank W1; Keller, Brian C2; Shapiro, Alexander3; Forero, Adriana1; Stencel-Baerenwald, Jennifer E1; Young, Jessica E4; Savan, Ram1; and Gale, Michael Jr.1
1Center for Innate Immunity and Immune Disease, Department of Immunology, University of Washington School of Medicine, Seattle, WA, USA; 2Division of Pulmonary, Allergy, Critical Care & Sleep Medicine, The Ohio State University College of Medicine, Columbus OH, USA; 3Shorewood High School, Shoreline, WA, USA; 4Institute for Stem Cell and Regenerative Medicine, Department of Pathology, University of Washington School of Medicine, Seattle, WA, USA
Pathogenic flaviviruses (including West Nile virus (WNV) and Zika virus (ZIKV)) antagonize host cell Janus kinase/signal transducer and activator of transcription (JAK/STAT) signaling in response to interferons (IFNs). Circumvention of the host antiviral response by flaviviruses is of critical importance to human health as this inhibition facilitates pathogenesis and promotes emerging epidemics; likely contributing to disease outcomes such as ZIKV-induced microencephaly in infants born to infected mothers. Diverse mechanisms (even amongst closely related viruses) have been proposed to drive this inhibition, predominantly centering on activity of the viral nonstructural protein 5 (NS5). It seems unlikely that such an advantageous phenotype would emerge independently at many points in flavivirus evolution. Rather, we hypothesized that there exists an underlying JAK/STAT inhibitory mechanism common to flaviviruses. To explore the precise nature of JAK/STAT inhibition by flaviviruses, we conducted a thorough mapping of the entire signaling cascade in response to multiple unrelated cytokines in WNV and ZIKV infected cells. In initial experiments with infected A549 epithelial cells, THP-1-derived macrophage-like cells, and stable Huh7 replicon cells we demonstrated that STAT phosphorylation (pY-STAT) was inhibited in response to IFN-β, IFN-γ, IFN-λ3, interleukin (IL-6), IL-4, and IL-10 which signal through STAT1, 2, 3, 4, 5, and 6-dependent processes. In addition, infection was associated with the loss of JAK proteins via proteasomal degradation. This broad inhibition is concurrent with NS5 interaction with heat shock protein 90 (HSP90) within infected cells, as demonstrated by cross-linked immunoprecipitation. HSP90 interaction with NS5 and viral RNA replication complexes was associated with a loss of abundance of several host kinase clients of this chaperone (including the JAK family proteins), indicating perturbation of normal chaperone activity during infection. The importance of this general signaling blockade was illustrated by ZIKV inhibition of JAK/STAT signaling in response to cytokines critical for appropriate brain development. Experiments with immortalized human fetal cortical NSCs (ReNcell CX) demonstrated that ZIKV-infected cells are less responsive to LIF, CNTF, and IL-6 as measured by pY-STAT3 accumulation. Allowing NSCs to differentiate for several days revealed that these cytokines were able to promote the early differentiation of neurons (as measured by βIII-tubulin and Neurofilament-L staining) and astrocytes (GFAP staining). Importantly, ZIKV infection altered this cytokine-induced NSC differentiation, dysregulating cell fate. Thus, ZIKV infection of the fetal brain can impose restrictions to NSC maturation and neural development through regulation of the JAK/STAT pathway and may serve as a plausible mechanism driving the progression to microencephaly.


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