Quantitation of adenovirus type 5 empty capsids

Takahashi, Eddie; Cohen, Steven L.; Tsai, Pei-Kuo; Sweeney, Joyce

doi:10.1016/j.ab.2005.11.014

Cited by 28 publications

(41 citation statements)

References 43 publications

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“…4b, arrows). The morphologies of the fluorescently labeled adenoviral particles did not indicate empty viral capsids 28. Collectively, the TEM images revealed modification of adenoviral capsid and core proteins causes various morphologies of viral particles.…”

Section: Resultsmentioning

confidence: 87%

In Vitro Dynamic Visualization Analysis of Fluorescently Labeled Minor Capsid Protein IX and Core Protein V by Simultaneous Detection

Ugai

Wang

et al. 2010

Journal of Molecular Biology

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Oncolytic adenoviruses represent a promising therapeutic medicine for human cancer therapy, but successful translation to human clinical trials requires careful evaluation of these viral characteristics. While the function of the adenovirus proteins have been analyzed in detail, the dynamics of adenovirus infection remain largely unknown due to technological constraints which prevent adequate tracking of the adenovirus particles after infection. Fluorescent labeling of the adenoviral particles is one new strategy designed to directly analyze dynamic processes of viral infection in virus-host cell interactions. We hypothesized that the double labeling technique of adenovirus with fluorescent proteins would allow us to properly analyze intracellular viruses and the fate of viral proteins in live analysis of adenovirus as compared to a single labeling. Thus, we generated a fluorescently labeled adenovirus with both a red fluorescent minor capsid protein IX (pIX-mRFP1) and a green fluorescent minor core protein V (pV-EGFP), resulting in Ad5-IX-mRFP1-E3-V-EGFP. The fluorescent signals for pIX-mRFP1 and pV-EGFP were detected within 10 min in living cells. However, the growth curve analysis of Ad5-IX-mRFP1-E3-V-EGFP showed approximately 150-fold reduced production of the viral progeny at 48 hours post-infection (h.p.i.) as compared to Ad5. Interestingly, pIX-mRFP1 and pV-EGFP were initially localized in the cytoplasm and the nucleolus, respectively, at 18 h.p.i. These proteins were observed in the nucleus during the late stage of infection and the relocalization of the proteins was observed in an adenoviral replication-dependent manner. These results indicate that the simultaneous detection of adenovirus using dual-fluorescent proteins is suitable for real-time analysis, including identification of infected cells, and monitoring viral spread, which will be required for complete evaluation of oncolytic adenoviruses.

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Section: Resultsmentioning

confidence: 87%

In Vitro Dynamic Visualization Analysis of Fluorescently Labeled Minor Capsid Protein IX and Core Protein V by Simultaneous Detection

Ugai

Wang

et al. 2010

Journal of Molecular Biology

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“…Polypeptide VIII is synthesized in precursor form (pVIII, 227 residues), present in assembly intermediates [130,131]. It has three potential cleavage sites by AVP at residues 112, 131 and 157 [132], but it was not clear which ones were actually cleaved, nor which fragments remained in the virion (Figure 6a).…”

Section: Atomic Structure Of the Mature Adenovirus Virionmentioning

confidence: 99%

Latest Insights on Adenovirus Structure and Assembly

Martín

2012

Viruses

140

129

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Adenovirus (AdV) capsid organization is considerably complex, not only because of its large size (~950 Å) and triangulation number (pseudo T = 25), but also because it contains four types of minor proteins in specialized locations modulating the quasi-equivalent icosahedral interactions. Up until 2009, only its major components (hexon, penton, and fiber) had separately been described in atomic detail. Their relationships within the virion, and the location of minor coat proteins, were inferred from combining the known crystal structures with increasingly more detailed cryo-electron microscopy (cryoEM) maps. There was no structural information on assembly intermediates. Later on that year, two reports described the structural differences between the mature and immature adenoviral particle, starting to shed light on the different stages of viral assembly, and giving further insights into the roles of core and minor coat proteins during morphogenesis [1,2]. Finally, in 2010, two papers describing the atomic resolution structure of the complete virion appeared [3,4]. These reports represent a veritable tour de force for two structural biology techniques: X-ray crystallography and cryoEM, as this is the largest macromolecular complex solved at high resolution by either of them. In particular, the cryoEM analysis provided an unprecedented clear picture of the complex protein networks shaping the icosahedral shell. Here I review these latest developments in the field of AdV structural studies.

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“…4B) revealed that these particles had very similar levels of outer capsid proteins (i.e., hexon, penton base, and fiber), whereas EC particles selectively lacked the DNA-associated capsid proteins V and VII. EC but not MC also contained the scaffold proteins L1 52/55 that are normally found in immature particles prior to viral capsid maturation and assembly (17,49). Unlike mature Ad particles, Ad5.F16 EC were incapable of inducing IFN-␣ expression (Fig.…”

Section: Ifn-␣ Expression By Cd46-utilizing Ads Requires Tlr Activationmentioning

confidence: 99%

Preferential Activation of Toll-Like Receptor Nine by CD46-Utilizing Adenoviruses

Iacobelli-Martinez

Nemerow

2007

J Virol

104

101

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Quantitation of adenovirus type 5 empty capsids

Cited by 28 publications

References 43 publications

In Vitro Dynamic Visualization Analysis of Fluorescently Labeled Minor Capsid Protein IX and Core Protein V by Simultaneous Detection

In Vitro Dynamic Visualization Analysis of Fluorescently Labeled Minor Capsid Protein IX and Core Protein V by Simultaneous Detection

Latest Insights on Adenovirus Structure and Assembly

Preferential Activation of Toll-Like Receptor Nine by CD46-Utilizing Adenoviruses

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