Human rhinovirus 2 induces the autophagic pathway and replicates more efficiently in autophagic cells - PubMed (original) (raw)

Human rhinovirus 2 induces the autophagic pathway and replicates more efficiently in autophagic cells

Kathryn A Klein et al. J Virol. 2011 Sep.

Abstract

Picornaviruses rearrange cellular membranes to form cytosolic replication sites. In the case of poliovirus and several other picornaviruses, these membranes are derived from subversion of the cellular autophagy pathway. We also reported observation of autophagosome-like structures during infection by two human rhinoviruses (HRVs), HRV-2 and HRV-14 (W. T. Jackson et al., PLoS Biol. 3:e156, 2005). Another group reported that HRV-2 does not induce autophagosomes or respond to changes in cellular autophagy (M. Brabec-Zaruba, U. Berka, D. Blaas, and R. Fuchs, J. Virol. 81:10815-10817, 2007). In this study, we tested HRV-2-infected cells for activation of autophagic signaling and changes in virus growth in response to changes in autophagy levels. Our data indicate that HRV-2 induces and subverts the autophagic machinery to promote its own replication.

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Figures

Fig. 1.

Fig. 1.

Induction of autophagosomes by HRV-2. 293T cells were transfected with a GFP-LC3 expression plasmid. Twenty-four hours later, cells were infected with virus at an MOI of 50, mock infected, or treated with 10 μM rapamycin (RAPA) or DMSO carrier. After 6 h of infection or treatment, cells were fixed for imaging and analysis. (A) Sample images of GFP-LC3 in cells. Arrows indicate a representative cell scored as punctate. (B) Quantification of cells displaying GFP-LC3 puncta. Three random fields, each containing at least 100 cells, were counted. Results from one representative experiment are shown.

Fig. 2.

Fig. 2.

LC3-II formation in response to HRV-2 infection. 293T cells were infected with PV, HRV-2, or HRV-1A at an MOI of 50. For controls, cells were mock infected or treated with 100 nM bafA for 6 h. At the designated hpi, cell lysates were collected and run on a 13% SDS-PAGE gel, which was probed for LC3 and actin or GAPDH. Unmodified LC3-I and lipid-modified LC3-II are labeled. Quantitation of lipid modification is below each band; intensities were normalized to loading controls, and then the ratio of LC3-II to LC3-I was expressed in arbitrary units.

Fig. 3.

Fig. 3.

HRV-2 replication correlates with autophagic activation. (A) H1-HeLa cells were treated with 10 μM rapamycin for 5 h, then infected with PV or HRV-2 at an MOI of 0.1, and fed with EMEM containing 10 μM rapamycin or an equivalent volume of DMSO carrier. At 6 hpi, cells were collected and cell-associated virus was measured by plaque assay. (B) H1-HeLa cells were infected with PV, HRV-1A, or HRV-2 at an MOI of 0.1 and fed with EMEM containing 10 mM 3-MA. Virus was collected at 0, 2, 4, and 6 h and assayed as described for panel A. Three replicate infections were assayed in each experiment; asterisks indicate significant differences from infections without 3-MA (Student's t test P values of <0.05).

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