Principles of Virology. Jane Flint

viral and cellular membranes. For nonenveloped viruses, conformational changes generally lead to disruption of the cellular membrane and the delivery of a subviral complex to the cell interior. In both cases, these changes ultimately allow the viral genome to access a cellular compartment that enables replication. Although the mechanisms of entry for the various virus families appear vastly different, certain themes repeatedly emerge. Structural rearrangements in viral proteins that enable entry are often dependent on proteolytic cleavage events that occur either during assembly in the virus-producing cell or at the surface or within endosomal compartments during entry into the target cell. The acid pH found in endosomes is a common trigger for conformational rearrangements that enable entry. Such conformational changes almost always result in the exposure of hydrophobic protein sequences that can interact with and disrupt cellular membranes and allow access to the cell interior.

      The cell is often not an idle target but an active participant in viral entry. Engagement of cell surface receptors by virus particles can trigger signal transduction pathways that lead to cytoskeletal rearrangement and endocytosis. Virus particle transport within the cell can be within vesicles, whose transport mechanisms are quite well understood. Conversely, vesicle-independent transport of viral or subviral particles on the cytoskeletal network is less well characterized. Notably, entry of various components of virus particles, nucleic acids and proteins, into the interior of the cell can be detected by specialized sensors that alert the innate immune system and elicit antiviral responses (a topic covered in Volume II, Chapter 3).

      For some viruses, the final destination, and the site of genome replication, is the cell’s nucleus. The nuclear envelope raises an additional barrier to virus entry, with a plethora of proteins regulating access to the nuclear interior through the nuclear pores. Virus particles or subviral structures are too large to pass through the nuclear pore. Therefore, interactions with the specialized nuclear transport machinery are usually necessary for subviral structures to be escorted into the nuclear interior. This process is not well understood for many viruses.

      Many questions about specific steps in the entry pathways of many viruses remain, including the elucidation of entry pathways used in whole organisms, a technically challenging endeavor. Understanding how entry proceeds and how particles “disassemble” to release the viral genome at the site of replication will allow us not only to develop specific interventions for prevention of virus infections but also to manipulate virus particles for use as viral vectors.

REFERENCES

       Books

      Pohlmann S, Simmons G. 2013. Viral Entry into Host Cells. Landes Bioscience, Austin, TX.

       Reviews

      Cosset F-L, Lavillette D. 2011. Cell entry of enveloped viruses. Adv Genet 73:121–183.

      Fay N, Panté N. 2015. Old foes, new understandings: nuclear entry of small non-enveloped DNA viruses. Curr Opin Virol 12:59–65.

      Grove J, Marsh M. 2011. The cell biology of receptor-mediated virus entry. J Cell Biol 195:1071–1082.

      Harrison SC. 2015. Viral membrane fusion. Virology 479-480:498–507.

      Moyer CL, Nemerow GR. 2011. Viral weapons of membrane destruction: variable modes of membrane penetration by non-enveloped viruses. Curr Opin Virol 1:44–49.

       Papers of Special Interest

      Bullough PA, Hughson FM, Skehel JJ, Wiley DC. 1994. Structure of influenza haemagglutinin at the pH of membrane fusion. Nature 371:37–43.

      Wilson IA, Skehel JJ, Wiley DC. 1981. Structure of the haemagglutinin membrane glycoprotein of influenza virus at 3 A resolution. Nature 289: 366–373.

       These two seminal papers were the first to report the structure of a viral fusion protein, influenza HA. These studies established the foundation of models for the conformational changes occurring in viral proteins during fusion.

      Butan C, Filman DJ, Hogle JM. 2014. Cryo-electron microscopy reconstruction shows poliovirus 135S particles poised for membrane interaction and RNA release. J Virol 88:1758–1770.

       This study shows the structural changes occurring in the poliovirus capsid shell during entry.

      Carette JE, Raaben M, Wong AC, Herbert AS, Obernosterer G, Mulherkar N, Kuehne AI, Kranzusch PJ, Griffin AM, Ruthel G, Dal Cin P, Dye JM, Whelan SP, Chandran K, Brummelkamp TR. 2011. Ebola virus entry requires the cholesterol transporter Niemann-Pick C1. Nature 477:340–343.

       This paper first identified the receptor for ebolavirus.

      Conley MJ, McElwee M, Azmi L, Gabrielsen M, Byron O, Goodfellow IG, Bhella D. 2019. Calicivirus VP2 forms a portal-like assembly following receptor engagement. Nature 565:377–381.

       This paper shows the formation of a pore in the capsid shell of caliciviruses that would allow the delivery of the (+) strand RNA genome to the cytoplasm.

      Harutyunyan S, Kumar M, Sedivy A, Subirats X, Kowalski H, Köhler G, Blaas D. 2013. Viral uncoating is directional: exit of the genomic RNA in a common cold virus starts with the poly-(A) tail at the 3′-end. PloS Pathog 9:e1003270.

       This study challenged the idea that the viral RNA 5′ end is first to exit the capsid during entry of all picornaviruses.

      Kane M, Rebensburg SV, Takata MA, Zang TM, Tamashita M, Kvaratskhelia M, Bieniasz PD. 2018. Nuclear pore heterogeneity influences HIV-1 infection and the antiviral activity of MX2. Elife 7:e35738.

       This paper examines the intertwined role of nucleoporins in HIV-1 nuclear import.

      Liu J, Yu C, Gui JF, Pang DW, Zhang QY. 2018. Real-time dissecting the entry and intracellular dynamics of single reovirus particle. Front Microbiol 9:2797.

       This paper uses microscopy techniques to track the entry of individual reovirus particles into cells.

      Wec AZ, Nyakatura EK, Herbert AS, Howell KA, Holtsberg FW, Bakken RR, Mittler E, Christin JR, Shulenin S, Jangra RK, Bharrhan S,Kuehne AI, Bornholdt ZA, Flyak AI, Saphire EO, Crowe JE Jr, Aman MJ, Lai JR, Chandran K. 2016. A “Trojan horse” bispecific-antibody strategy for broad protection against ebolaviruses. Science 354:350–354.

       This study relies on the unique understanding of Ebolavirus entry to generate novel antibodies that inhibit entry by being delivered to the appropriate subcellular compartment.

STUDY QUESTIONS

      1 You are studying a new DNA virus. You have two cell lines: cell line α expresses the receptor (permissive) but is not susceptible (multiple blocks to the viral reproduction cycle) and cell line β is not permissive but is susceptible. Which of the following statements are correct and why?Transfection of the virus DNA into α will lead to production of infectious particles.Transfection of virus DNA into β will produce infectious particles.Inhibiting production of the receptor in α will allow virus reproduction.Production of the virus receptor in β will allow virus reproduction.

      2 You are working on a rhabdovirus that is cytopathic in dog cells but not rodent cells. What strategy would you use