How does rhinovirus reproduce

This association plays an important role in holding pentamers together in the mature virion as this contact region includes more than half the total short non-bonded contact between pentamers. Contacts between protomers within pentamers are more extensive than the contacts betwen pentamers, explaining in part the stability of pentamers.

The immunogenic regions are correlated with high solvent accessibility: accessibility to large probes and high thermal parameters.

Surface residues in the putative cellular receptor recognition site, the canyon, have lower thermal parameters than other portions of the human RV 14 surface.

A number of unsual crevices exist in the protein shell of human RV 14, including the hydrophobic pocket in VP1 which is the locus binding for the WIN antiviral agents. These may be required for conformational flexibility during both assembly and disassembly. A number of unusual crevices exist in the protein shell of human rhinovirus 14, including the hydrophobic pocket in VP1 which is the locus of binding for the WIN antiviral agents.

These may be required for conformational flexibility during assembly and disassembly. The RV is inhaled directly or finds its way to the nassal passage through hands that are contaminated with the virus. The virus is then transported to the back of the nose by the regular functioning of the nose itself. Here, the RV attaches itself to the receptors at the surface of the nasal cells, where it reproduces.

The reproductive cycle of the RV is hours long. Once the cells have been attacked, in the infected cells, the virus replicates as rapidly as possible, and continually sheds progeny viruses. These progeny viruses can propagate the infection by invading neighboring cells. The infection remains localized in the upper respiratory tract. The virus can also be swallowed and end up in the stomach where both increased temperature and decreased pH work to prevent infection.

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However, in all cases, levels of negative strand were approximately 10,fold lower than those of positive strand genome. Comparison of total copy numbers HRV intracellular positive genomic and negative template RNA strands at multiple times after infection with varying levels of HRV We then used the same negative strand copy number data to assess the relationship of dsRNA levels to production of a number of other molecules from the same cultures.

In contrast to the close association between levels of negative strand template and genomic positive strand, no such relationship was observed for production of IFNs. Relationship between copy number of negative strand template taken from Fig.

Expression of mRNA and protein secretion predominantly into the basolateral medium of the chemokine CXCL10 showed similar temporal and quantitative patterns to the IFNs, with a clear relationship between levels of induction and initial infectious dose Fig. Moreover, the dependence on infectious dose was less clear, as comparable viperin expression was observed at all initial infectious doses except 10 4 TCID 50 of HRV , and the reduction of ISG15 expression at lower doses was less marked.

Generation of all of the host defense molecules examined depend upon viral replication, since exposure of HBE to 10 4 TCID 50 of HRV that was rendered replication deficient by exposure to UV light did not induce epithelial expression of any of the molecules studied not shown.

The HeLa cervical carcinoma cell line was invaluable for the development of vaccines against poliovirus [ 20 ], and the ease of infection of HeLa cells by picornavirus family members has led to the frequent use of these cells for studies of picornavirus replication.

The physiological relevance of such studies of HRV replication must be questioned, however, since HRV selectively infects the airways. HRV infection studies in vivo demonstrate that epithelial cells are the natural site of HRV infection and replication [ 5 , 6 , 21 ], and the use of human airway epithelial cell lines [ 22 ], and subsequently primary HBE grown in submersion culture [ 23 , 24 ], or at ALI [ 25 , 26 ], have provided important insights into the inflammatory and innate immune responses to HRV infection.

HBE in submersion culture are not suited, however, for studying the complete HRV replication cycle, as confluent cultures cannot be maintained for extended periods. Moreover, these cultures can be maintained for months at ALI, permitting the current studies examining the HRV replication cycle in response to varying initial infectious doses of HRV.

Although well differentiated cultures of human airway epithelial cells have been used to study kinetic responses to infection with respiratory syncytial virus [ 27 , 28 ], to our knowledge this is the first use of such cells for extended kinetic studies of responses to HRV infection. Our data clearly show that similar peak intracellular and shed levels of HRV genomic RNA are observed regardless of the initial infectious dose of virus, although the peak of viral replication was delayed at lower infectious doses.

Whether the peak levels of intracellular and shed virus observed with all infectious doses represent the maximal viral replication that can be supported by the cells is unclear. Given that the initial infectious material removed from the cells contains levels of HRV that are not distinguishable from the dose applied not shown , it would appear that few virions are taken up by cells.

Thus, it is unclear why a delay in replication is observed with lower doses. One could speculate that fewer cells are initially infected with lower doses and that it takes longer for all cells that can support replication to be infected. This is reflected both by intracellular and shed virus levels. It should be noted that the general time course of shed virus in these cultures resembles that observed in nasal secretions following experimental HRV infection of normal volunteers [ 17 ].

Overall, these data indicate that epithelial antiviral responses on their own are adequate to clear HRV infections and that, while recruitment of inflammatory and immune cells to the airway mucosa in vivo may contribute to symptoms, these cells are not essential for virus clearance. The generation of negative strand template is a prerequisite for transcription of new genomic RNA strands.

To examine the formation of dsRNA we used two complementary approaches. This perinuclear localization is consistent with the concept that HRV replicates on membrane fragments derived from endoplasmic reticulum or golgi [ 29 , 30 ]. To our knowledge there has been no prior quantitative assessment of the level of dsRNA achieved in airway epithelial cells infected with HRV.

We examined absolute copy numbers of negative strand as a means to quantify dsRNA. Prior studies of single strand genome viruses have shown that conventional RT-PCR targeting the negative strand does not provide strand-specificity. This technique showed that similar levels of negative strand and, thus, presumably, dsRNA were achieved regardless of the initial infectious dose, albeit with varying time-courses.

This implies that a defined level of negative strand is required for optimal replication of genomic RNA. If the concept put forward for poliovirus that genomes that have not been translated cannot function as templates for negative strand synthesis [ 37 , 38 ] also holds true for HRV, this would suggest that a defined level of virus polyprotein strands are translated after infection in order to support optimal viral replication. Given that copy numbers of negative strand were about 10,fold lower than those of positive strand, transcription of positive strand from template must be highly efficient.

Our data differ from a previous report of a ratio of only about 70 positive strands per negative strand in cells infected with poliovirus, assessed using an RNase protection assay [ 39 ]. This difference may be due to specificity of the natural cell host the cell type used in the prior poliovirus study was never defined , differences between viruses, or the sensitivity and specificity of the methods used.

The dsRNA generated during viral replication is recognized by pattern recognition receptors PRRs to induce innate immune responses. Studies have reported varying roles of the dsRNA receptors, toll-like receptor-3, retinoic acid inducible gene-I and melanoma differentiation associated gene-5 in inducing epithelial responses to HRV infection [ 19 , 40 , 41 ].

For each infectious dose of HRV, the kinetics, but not the quantity, of IFN induction mirrored that of negative strand template. Thus, the degree of induction of IFNs is not simply regulated by levels of dsRNA, as reflected by levels of negative strand.

This implies that other factors generated during the replication cycle must play a role. The identity of these factors is unknown but single-stranded RNA, other replications intermediates, or other components of the viral genome, may contribute. We have also observed this for other epithelial mediators not shown. At this point the reasons for these differences are unknown.

Such IFN-independent induction occurs via direct viral induction of key transcription factors, particularly members of the interferon regulatory factor IRF family. Similarly, it has previously been show that viperin can be induced independently of IFN signaling via an IRF-1 mediated mechanism [ 44 ]. Although textbooks frequently state that HRV virions escape cells via lysis, this is based on studies in HeLa cells.

It has been known for years, however, that HRV infections in vivo do not cause overt epithelial cytotoxicity [ 47 ]. We cannot rule out that death or shedding of a small percentage of cells occurs during infection, but if so, the epithelium either generates replacement cells or else adjacent cells expand to maintain an intact epithelium.

In terms of alternative means for HRV to escape epithelial cells, there is precedent for HRV and other enterovirus species to leave other cell types via phosphatidylserine rich membrane vesicles with properties of either exosomes or autophagosomal like vesicles [ 48 , 49 , 50 ].

Nonetheless, we must avoid generalizing our data to all strains of HRV. Additional studies will be required to determine if HRV-A strains that use the low density lipoprotein as a receptor, or HRV-C strains that require cadherin related family member 3 to gain cell entry [ 51 ], show the same patterns of virus clearance, epithelial integrity, negative strand template expression and antiviral gene expression observed for HRV It will also be of interest to determine if responses to a second consecutive infection, either with the same HRV strain or a different one, will show altered responses.

These HRV infections are cleared without the need for immune cells, and the time to clearance does not depend on levels of IFNs.

Regardless of initial infectious dose used, relatively constant levels of genomic and negative strand RNA are generated, albeit with varying kinetics. Copy numbers of negative-strand RNA are some 10,fold lower than numbers of positive genomic strands. Although relatively constant levels of negative strands are generated regardless of initial infectious dose, levels of type I and type III IFNs vary depending upon initial infectious dose, implying that factors other than levels of dsRNA regulate IFN induction.

These data challenge a number of widely held paradigms generated from earlier studies in HeLa cells and emphasize the importance of appropriate cell context when performing experiments using HRV infections. Viruses and bacteria in the etiology of the common cold. J Clin Microbiol. Storer, Megan E. Fitzgerald, Bethany R. Temperature-dependent innate defense against the common cold virus limits viral replication at warm temperature in mouse airway cells.

ScienceDaily, 5 January Yale University. Cold virus replicates better at cooler temperatures. Retrieved November 14, from www. Their finding could help solve the mystery of why some people exposed to This is most evident in temperate regions where forest trees shed their leaves to conserve energy