Do viruses require the cytoskeleton?
© Matthews et al.; licensee BioMed Central Ltd. 2013
Received: 9 May 2012
Accepted: 11 April 2013
Published: 18 April 2013
It is generally thought that viruses require the cytoskeleton during their replication cycle. However, recent experiments in our laboratory with rubella virus, a member of the family Togaviridae (genus rubivirus), revealed that replication proceeded in the presence of drugs that inhibit microtubules. This study was done to expand on this observation.
The replication of three diverse viruses, Sindbis virus (SINV; family Togaviridae family), vesicular stomatitis virus (VSV; family Rhabdoviridae), and Herpes simplex virus (family Herpesviridae), was quantified by the titer (plaque forming units/ml; pfu/ml) produced in cells treated with one of three anti-microtubule drugs (colchicine, noscapine, or paclitaxel) or the anti-actin filament drug, cytochalasin D. None of these drugs affected the replication these viruses. Specific steps in the SINV infection cycle were examined during drug treatment to determine if alterations in specific steps in the virus replication cycle in the absence of a functional cytoskeletal system could be detected, i.e. redistribution of viral proteins and replication complexes or increases/decreases in their abundance. These investigations revealed that the observable impacts were a colchicine-mediated fragmentation of the Golgi apparatus and concomitant intracellular redistribution of the virion structural proteins, along with a reduction in viral genome and sub-genome RNA levels, but not double-stranded RNA or protein levels.
The failure of poisons affecting the cytoskeleton to inhibit the replication of a diverse set of viruses strongly suggests that viruses do not require a functional cytoskeletal system for replication, either because they do not utilize it or are able to utilize alternate pathways when it is not available.
KeywordsVirus replication Cytoskeleton Microtubules Actin filaments
Viruses used in this study
Site of replication
Herpes simplex virus (HSV-1)
Sindbis virus (SINV)
Vesicular stomatitis virus (VSV)
Drugs used in this study
Mode of action
Plants of the Papaveraceae family
Inhibits microtubule dynamics
Inhibits mitosis by stabilizing microtubules
Depolymerizes actin filaments
In summary, following studies in our lab with rubella virus  which found that its replication was not inhibited by four anti-microtubule drugs, we decided to test the hypothesis that viruses can replicate in the presence of drugs which compromise the cytoskeletal system by broadening our study to include another positive-strand RNA virus, a negative-strand RNA virus, and a DNA virus. Our findings demonstrate that viruses can produce normal titers in the absence of a functional cytoskeletal system, (similar results were reported for SINV in another lab ) which challenges the currently accepted notion . In this regard, it was shown that poliovirus can complete its entire infection cycle in a cell-free system lacking a cytoskeleton system . To address the alternate hypotheses of whether viruses simply do not need the cytoskeletal system or use alternate pathways when it is not available, we investigated the replication cycle of SINV in the presence of the anti-cytoskeletal drugs in more detail. No obvious changes occurred to any of the stages of SINV infection in the presence of noscapine, paclitaxel, and cytochalasin D. However, the Golgi through which the SINV envelope glycoproteins mature during transport to the plasma membrane, was severely compromised by colchicine, concomitantly affecting the distribution of the SINV structural proteins. It will be of interest to study the effect of colchicine treatment on maturation and transport of these SINV proteins to see if an alternate pathway exists that the virus uses in this step of its replication cycle in the presence of this drug.
RM is attending vet school at the University of Georgia. JDM is a Postdoctoral Fellow in the Department of Pathology, Emory University School of Medicine.
The research was supported by a grant from NIH (AI21389) to TKF.
Thanks to Dr. Ilya Frolov for the SINV/NSP3-GFP construct, Dr. Charlie Rice for the nsP2 antibody, Dr. Richard Kuhn for the polyclonal cdE2 antibody, and Dr. Ritu Aneja for the noscapine and paclitaxel. We acknowledge Yu-Han Tsai, Meng-Chun Hsieh, Chia-Hsuan Chan, and Fen-Hua Chang, undergraduate exchange students from China Medical University, Tiachung, Taiwan, who were supervised in the lab by Dr. Yumei Zhou, for their efforts in the preliminary stages of this study.
- Pollard TD, Cooper JA: Actin, a central player in cell shape and movement. Science 2009, 326: 1208-1212. 10.1126/science.1175862PubMedPubMed CentralView Article
- Goldman RD, Grin B, Mendez MG, Kuczmarski ER: Intermediate filaments: versatile building blocks of cell structure. Curr Opin Cell Biol 2008, 20: 28-34. 10.1016/j.ceb.2007.11.003PubMedPubMed CentralView Article
- Wade RH: Microtubules: an overview. Methods Mol Med 2007, 137: 1-16. 10.1007/978-1-59745-442-1_1PubMedView Article
- Radtke K, Dohner K, Sodeik B: Viral interactions with the cytoskeleton: a hitchhiker’s guide to the cell. Cell Microbiol 2006, 8: 387-400. 10.1111/j.1462-5822.2005.00679.xPubMedView Article
- Matthews JD, Tzeng WP, Frey TK: Analysis of the function of cytoplasmic fibers formed by the rubella virus nonstructural replicase proteins. Virology 2010, 406: 212-227. 10.1016/j.virol.2010.07.025PubMedPubMed CentralView Article
- Barton DJ, Sawicki SG, Sawicki DL: Solubilization and immunoprecipitation of alphavirus replication complexes. J Virol 1991, 65: 1496-1506.PubMedPubMed Central
- Frolova EI, Gorchakov R, Pereboeva L, Atasheva S, Frolov I: Functional Sindbis virus replicative complexes are formed at the plasma membrane. J Virol 2010, 84: 11679-11695. 10.1128/JVI.01441-10PubMedPubMed CentralView Article
- Hardy WR, Strauss JH: Processing the nonstructural polyproteins of Sindbis virus: study of the kinetics in vivo by using monospecific antibodies. J Virol 1988, 62: 998-1007.PubMedPubMed Central
- Frolova E, Gorchakov R, Garmashova N, Atasheva S, Vergara LA, Frolov I: Formation of nsP3-specific protein complexes during Sindbis virus replication. J Virol 2006, 80: 4122-4134. 10.1128/JVI.80.8.4122-4134.2006PubMedPubMed CentralView Article
- Wirth DF, Katz F, Small B, Lodish HF: How a single Sindbis virus mRNA directs the synthesis of one soluble protein and two integral membrane glycoproteins. Cell 1977, 10: 253-263. 10.1016/0092-8674(77)90219-7PubMedView Article
- Knipfer ME, Brown DT: Intracellular transport and processing of Sindbis virus glycoproteins. Virology 1989, 170: 117-122. 10.1016/0042-6822(89)90358-9PubMedView Article
- Yang W, Storrie B: Scattered Golgi elements during microtubule disruption are initially enriched in trans-Golgi proteins. Mol Biol Cell 1998, 9: 191-207.PubMedPubMed CentralView Article
- Molla A, Paul AV, Wimmer E: Cell-free, de novo synthesis of poliovirus. Science 1991, 254: 1647-1651. 10.1126/science.1661029PubMedView Article
This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.