One-sentence summary: This review addresses the structural rearrangements of dengue virus proteins and their functions during virus entry into the host cells, exploring (a) the cellular elements involved in virus binding to mammalian and mosquito cells, (b) the internalization routes that ultimately lead to virus entry into the cell and (c) the mechanisms by which viral genome gain access to the cytoplasm, including original insights from our recent work that supports the hypothesis that the capsid protein has a role in this process. Editor: Urs Greber ABSTRACTDengue is the most prevalent arthropod-borne viral disease, caused by dengue virus, a member of the Flaviviridae family. Its worldwide incidence is now a major health problem, with 2.5 billion people living in risk areas. In this review, we integrate the structural rearrangements of each viral protein and their functions in all the steps of virus entry into the host cells. We describe in detail the putative receptors and attachment factors in mammalian and mosquito cells, and the recognition of viral immunocomplexes via Fcγ receptor in immune cells. We also discuss that virus internalization might occur through distinct entry pathways, including clathrin-mediated or non-classical clathrin-independent endocytosis, depending on the host cell and virus serotype or strain. The implications of viral maturation in virus entry are also explored. Finally, we discuss the mechanisms of viral genome access to the cytoplasm. This includes the role of low pH-induced conformational changes in the envelope protein that mediate membrane fusion, and original insights raised by our recent work that supports the hypothesis that capsid protein would also be an active player in this process, acting on viral genome translocation into the cytoplasm.

Dengue virus (DENV), a mosquito-borne flavivirus, is a public health problem in many tropical countries. IL-22 and IL-17A are key cytokines in several infectious and inflammatory diseases. We have assessed the contribution of IL-22 and IL-17A in the pathogenesis of experimental dengue infection using a mouse-adapted DENV serotype 2 strain (P23085) that causes a disease that resembles severe dengue in humans. We show that IL-22 and IL-17A are produced upon DENV-2 infection in immune-competent mice. Eur. J. Immunol. 2013Immunol. . 43: 1529Immunol. -1544 Keywords: Dengue virus · IL-17A · IL-22 · Infection · Inflammation Additional supporting information may be found in the online version of this article at the publisher's web-site IntroductionDengue fever (DF) and its severe forms, dengue haemorrhagic fever (DHF) and dengue shock syndrome, are mosquito-borne diseases caused by one of four serotypes of Dengue virus (DENV 1-4). There are an estimated 50-100 million cases of DF annually mostly in tropical and subtropical regions of the world, and 20 000 deaths are estimated to occur each year [1,2]. DHF is defined by the WHO as fever with haemorrhagic manifestations, thrombocytopenia, haemoconcentration, or other signs of plasma leakage [1,3]. Treatment of DF and severe forms of dengue infection is largely supportive [2,3]. Human studies have demonstrated that secondary infection by a heterologous serotype is the single greatest risk factor for DHF/dengue shock syndrome [4,5]. However, manifestations of severe disease in primary infection are also frequently reported [6,7]. The immunopathogenesis of DENV infection involves the effects of cytokines on both infected and bystander immune cells [8][9][10]. High levels of pro-inflammatory cytokines, including TNF-α, IL-6, IL-8, CCL2/MCP-1 and IFN-γ, have been reported in patients with severe dengue disease [3,9,11]. However, it is not clearly understood how this massive cytokine production is induced and eventually controlled.IL-22 is a member of the IL-10 cytokine family and is believed to play important roles in inflammation and tissue homeostasis [12,13]. IL-22 receptor complex (IL-22R) is expressed in nonhaematopoietic cells in the skin, kidney, liver, lungs and gut, allowing for IL-22-mediated regulation of local responses after infection or inflammation [14,15]. IL-22 can be produced not only by Th17 cells but also by NK cells, NKT cells, γδT cells, or lymphoid tissueinducer-like cells [15][16][17]. The Th17-cell population coexpresses IL-17A, 15] . Both IL-17 and IL-22 induce an innate immune response in epithelial cells, but their functional spectra are generally distinct. Whereas IL-17 induces an inflammatory tissue response, IL-22 is believed to be mainly protective and/or regenerative [12,13,15].In viral infections, IL-22 seems to a play a marginal protective role in primary respiratory infection by Influenza A, not contributing to viral clearance, whereas IL-17 and its receptor IL-17RA contribute to acute lung injury caused by the flu [18,19]. IL-22 appears t...

Dengue virus (DENV) infection is associated to exacerbated inflammatory response and structural and functional alterations in the vascular endothelium. However, the mechanisms underlying DENV-induced endothelial cell activation and their role in the inflammatory response were not investigated so far. We demonstrated that human brain microvascular endothelial cells (HBMECs) are susceptible to DENV infection, which induces the expression of the cytoplasmic pattern recognition receptor (PRR) RIG-I. Infection of HBMECs promoted an increase in the production of type I IFN and proinflammatory cytokines, which were abolished after RIG-I silencing. DENV-infected HBMECs also presented a higher ICAM-1 expression dependent on RIG-I activation as well. On the other hand, ablation of RIG-I did not interfere with virus replication. Our data suggest that RIG-I activation by DENV may participate in the disease pathogenesis through the modulation of cytokine release and expression of adhesion molecules, probably contributing to leukocyte recruitment and amplification of the inflammatory response.

Supercharged proteins are a recently identified class of proteins that have the ability to efficiently deliver functional macromolecules into mammalian cells. They were first developed as bioengineering products, but were later found in the human proteome. In this work, we show that this class of proteins with unusually high net positive charge is frequently found among viral structural proteins, more specifically among capsid proteins. In particular, the capsid proteins of viruses from the Flaviviridae family have all a very high net charge to molecular weight ratio (> +1.07/kDa), thus qualifying as supercharged proteins. This ubiquity raises the hypothesis that supercharged viral capsid proteins may have biological roles that arise from an intrinsic ability to penetrate cells. Dengue virus capsid protein was selected for a detailed experimental analysis. We showed that this protein is able to deliver functional nucleic acids into mammalian cells. The same result was obtained with two isolated domains of this protein, one of them being able to translocate lipid bilayers independently of endocytic routes. Nucleic acids such as siRNA and plasmids were delivered fully functional into cells. The results raise the possibility that the ability to penetrate cells is part of the native biological functions of some viral capsid proteins.

No abstract