REVIEWS Rewiring cellular networks by members of the Flaviviridae family Christopher J. Neufeldt1*, Mirko Cortese1*, Eliana G. Acosta1* and Ralf Bartenschlager1,2 Abstract | Members of the Flaviviridae virus family comprise a large group of enveloped viruses with a single-strand RNA genome of positive polarity. Several genera belong to this family, including the Hepacivirus genus, of which hepatitis C virus (HCV) is the prototype member, and the Flavivirus genus, which contains both dengue virus and Zika virus. Viruses of these genera differ in many respects, such as the mode of transmission or the course of infection, which is either predominantly persistent in the case of HCV or acutely self-limiting in the case of flaviviruses. Although the fundamental replication strategy of Flaviviridae members is similar, during the past few years, important differences have been discovered, including the way in which these viruses exploit cellular resources to facilitate viral propagation. These differences might be responsible, at least in part, for the various biological properties of these viruses, thus offering the possibility to learn from comparisons. In this Review, we discuss the current understanding of how Flaviviridae viruses manipulate and usurp cellular pathways in infected cells. Specifically, we focus on comparing strategies employed by flaviviruses with those employed by hepaciviruses, and we discuss the importance of these interactions in the context of viral replication and antiviral therapies. Guillain–Barré syndrome As obligate intracellular parasites, viruses strictly depend haemorrhage, shock or serious neuro logi cal compli- An acute neurological disease on their ability to manipulate the machinery of host cations, such as encephalitis and meningitis. Dengue (usually reversible) resulting cells to propagate. Consequently, viruses have evolved virus (DENV), the aetiological agent of dengue fever from autoimmune destruction numerous strategies to manipulate infected cells by trig- and dengue haemorrhagic fever, or dengue shock syn- of the peripheral nervous system that is frequently gering a series of metabolic and structural changes that drome, is responsible for an estimated 60 million symp- triggered by infection. facilitate viral replication. tomatic infections annually, causing approximately The Flaviviridae family provides many fascinating 10,000 deaths per year4. Although a DENV vaccine has examples of virus-driven cellular reprogramming. This recently been licensed, its overall efficacy is limited, family is composed of four genera: Flavivirus (with 53 especially in immunologically naive individuals, and its species); Hepacivirus (with 14 species); Pegivirus (with administration is not recommended for young children 11 species) and Pestivirus (with 4 species)1. Several mem- or elderly people, both of whom have a higher risk of bers within the Hepacivirus and Flavivirus genera have a serious disease5. substantial impact on human health. Chronic infection Examples of neurotrophic flaviviruses include 1 Department of Infectious by hepatitis C virus (HCV), the prototypic hepacivirus, West Nile virus (WNV), tick-borne encephalitis virus Diseases, Molecular Virology, Heidelberg University, is the leading cause of liver disease worldwide, with (TBEV), Japanese encephalitis virus (JEV) and Zika 69120 Heidelberg, Germany. ~71 million individuals at risk of developing liver cir- virus (ZIKV). The latter has recently spread world- 2German Center for Infection rhosis and hepatocellular carcinoma2. Recently, highly wide, and infections have been linked to Guillain–Barré Research, Heidelberg Partner effective direct-acting antiviral drugs targeting essential syndrome in adults and multiple neurodevelopmen- Site, 69120 Heidelberg, microcephaly Germany. viral processes have become available for clinical use; tal defects, including in infants born to Correspondence to R.B. however, a prophylactic vaccine to control the HCV pan- mothers infected during the first trimester of pregnancy ralf.bartenschlager@ demic is still missing (reviewed in REF. 3). In contrast to (reviewed in REF. 6). Although ZIKV is rarely neuro- med.uni-heidelberg.de the predominantly persistent infection by HCV, human invasive in adults, it can infect human neural progenitor *These authors contributed infections with flaviviruses are acute and self-limiting cells (hNPC), likely resulting in the congenital disorders equally to this work. and are either asymptomatic or present as an undiffer- mentioned above (reviewed in REF. 7). At present, no doi:10.1038/nrmicro.2017.170 entiated febrile illness that can, in specific cases, lead to approved anti viral drugs are available for the treatment Published online 12 Feb 2018 more severe symptoms, such as vascular leakage, severe of Flavivirus infections. NATURE REVIEWS | MICROBIOLOGY VOLUME 16 | MARCH 2018 | 125 ©2018 Mac millan Publishers Li mited, part of Spri nger Nature. All ri ghts reserved. REVIEWS In addition to the marked differences in tropism cap-dependent pathway in the case of flaviviruses or and pathogenesis, viruses within the Flavivirus and an internal ribosome entry site (IRES) in the case of HCV Hepacivirus genera, while sharing similarities in their (FIG. 1). Along the same line, the general principle of overall genome organization, differ in several respects, the replication cycles of flaviviruses and hepaciviruses such as their mechanism of translation: a canonical, is similar (BOX 1 and BOX 2, respectively), but multiple a Flavivirus ? C prM E NS1 NS2A NS3 NS4A NS4B NS5 Stop Cap AUG NS2B 2K 5ʹ NTR Structural proteins Non-structural proteins 3ʹ NTR NS5 NS3 Polymerase Helicase Capsid Methyltransferase Protease NS2B Cofactor prM E NS2A 2K NS4B Capsid NS1 NS4A Membrane RNA synthesis reorganization Envelope Replication glycoprotein assembly b Hepacivirus C E1 E2 NS2 NS3 NS4B NS5A NS5B AUG * p7 NS4A Stop 5ʹ NTR Structural proteins Non-structural proteins 3ʹ NTR IRES D2 D3 NS3 Helicase NS5B Protease NS5A Core NS4A D1 protein E1 E2 p7 NS2 NS4B Capsid Protease Membrane Phosphoprotein assembly reorganization RNA replication assembly Envelope Ion channel Cofactor glycoproteins Nature Reviews | Microbiology 126 | MARCH 2018 | VOLUME 16 www.nature.com/nrmicro ©2018 Mac millan Publishers Li mited, part of Spri nger Nature. All ri ghts reserved. REVIEWS Microcephaly differences have been discovered during the past few Architecture of replication organelles A neurological condition of years, including the dependency on a specific lipid kinase Replication of the genome of positive-strand RNA ((+) abnormal brain development in the case of HCV but not DENV or ZIKV. These dif- RNA) viruses occurs in close association with cellu- that causes substantially ferences and similarities reflect the strategies that are lar endomembranes within organelle-like structures smaller infant head circumferences relative to used by these viruses to manipulate host cells, offering defined as replication organelles (ROs). These ROs age-matched controls. the opportunity to learn about the roles of host cell fac- serve to increase the local concentration of cellular tors, pathways and the mechanisms of their manipulation and viral factors that are required for genome repli- Tropism by conducting comparative analyses. In this Review, we cation, coordinate the different steps of viral replica- Tissue specificity of a virus. summarize the current understanding of how members tion through their compartmentalization and to shield Tropism is determined primarily by the presence of of the Flaviviridae family take control of cellular compo- genomic RNA from cellular innate immune sensors. membrane receptors that can nents or processes in order to create environments that ROs can be grouped into two morphologically distinct be exploited by the virus to are favourable for viral replication. We will compare and classes designated as invaginated/spherule-type ROs or gain access to host cells. contrast DENV and ZIKV with HCV as representatives protrusion-type ROs. Invaginated/spherule-type ROs of the Flavivirus and Hepacivirus genera, respectively. are generated by invaginations of the donor membrane Understanding the details of how viruses exploit their into the organelle lumen. The protrusion-type ROs host cells opens new avenues for the development of are composed of clusters of single-membrane vesicles antiviral strategies, and comparing replication strategies (SMVs), double-membrane vesicles (DMVs), multi- within the Flaviviridae may help us to identify shared membrane vesicles (MMVs) and often multimembrane essential host-dependency factors that are suitable for tubules. Interestingly, although donor membranes can the development of broad-spectrum antivirals with be provided by different organelles, ROs from all (+) ­pan-flaviviral activity. RNA viruses can be classified into one of these two morphotypes, suggesting evolutionarily conserved ­mechanisms of their biogenesis8. In the case of flaviviruses, electron microscopy ana­ Figure 1 | Flavivirus and Hepacivirus genome organization and membrane topology ◀ lysis of cells infected with DENV or ZIKV revealed the of mature viral proteins. The ORF encoding the dengue virus (DENV) (part a) or hepatitis C virus (HCV) (part b) polyprotein and the predicted secondary structures of formation of clusters of vesicles ~90 nm in diameter, the 5ʹ and 3ʹ non-translating regions (NTR) are depicted on the top of each panel. a | The defined as vesicle packets (VPs), that are created by DENV genome