Effects of deletion of the ac109 gene of Autographa californica nucleopolyhedrovirus on interactions with mammalian cells

Alfonso, Victoria; Amalfi, Sabrina; López, María Gabriela; Taboga, Oscar

doi:10.1007/s00705-016-3142-y

Cited by 3 publications

(4 citation statements)

References 28 publications

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“…However, nuclear importation or cytoplasmic traffic suffers a blockage in these cells (11). Regarding the accessibility to gene transcription, we have previously demonstrated the presence of viral genomes in the cytoplasm of BHK-21 cells; nevertheless, we were not able to rule out a disassembling of inner cellular membranes or disruption of actin microfilaments by the vaccinia virus used as a tool and subsequent detection of endosomal DNA (42). Fluorescent in situ hybridization (FISH) assays in PK1 cells (6) and DNA detection in the cytoplasm of rat chondrocytes ( 16) by qPCR might allow the detection of endosomal content.…”

Section: Discussionmentioning

confidence: 69%

“…Wild-type AcMNPV was obtained from Pharmingen (BD Biosciences). The AcCAGeGFP virus was constructed previously (42). AcT7IRESmCherry was generated using the Bac-to-Bac system.…”

Section: Methodsmentioning

confidence: 99%

“…The linear PCR fragment was gel purified and cloned into pGEM-T Easy vector systems (Promega) to obtain pGemT-T7IRES. This plasmid was digested with the enzymes PvuII and SpHI and cloned into the pFBDmCherry plasmid (42) digested with the same enzymes, to obtain pFBD-T7IRESmCherry.…”

Section: Methodsmentioning

confidence: 99%

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Baculovirus Transduction in Mammalian Cells Is Affected by the Production of Type I and III Interferons, Which Is Mediated Mainly by the cGAS-STING Pathway

Amalfi

Molina

Bevacqua

et al. 2020

J Virol

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The baculovirus Autographa californica multiple nucleopolyhedrovirus is an insect virus with a circular double-stranded DNA genome, which, among other multiple biotechnological applications, is used as an expression vector for gene delivery in mammalian cells. Nevertheless, the nonspecific immune response triggered by viral vectors often suppresses transgene expression. To understand the mechanisms involved in that response, in the present study, we studied the cyclic GMP-AMP synthase-stimulator of interferon genes (cGAS-STING) pathway by using two approaches: the genetic edition through CRISPR/Cas9 technology of genes encoding STING or cGAS in NIH/3T3 murine fibroblasts and the infection of human epithelial cells HEK293 and HEK293 T, deficient in cGAS or in cGAS and STING expression, respectively. Overall, our results suggest the existence of two different pathways involved in the establishment of the antiviral response, both dependent on STING expression. Particularly, the cGAS-STING pathway resulted more relevant in the production of interferon (IFN)-β and IFN-λ1 in response to baculovirus infection. In human epithelial cells, IFN-λ1 production was also induced in a cGAS-independent and DNA-PK-dependent manner. Finally, we demonstrated that these cellular responses towards baculovirus infection affect the efficiency of transduction of baculovirus vectors. IMPORTANCE Baculoviruses are non-pathogenic viruses that infect mammals, which, among other applications, are used as vehicles for gene delivery. Here, we demonstrated that the cytosolic DNA sensor cGAS recognizes baculoviral DNA and that the cGAS-STING axis is primarily responsible for the attenuation of transduction in human and mouse cell lines through type I and III IFNs. Furthermore, we identified DNA-dependent protein kinase (DNA-PK) as a cGAS-independent and alternative DNA cytosolic sensor that contributes less to the antiviral state in baculovirus infection in human epithelial cells than cGAS. Knowledge of the pathways involved in the response of mammalian cells to baculovirus infection will improve the use of this vector as a tool for gene therapy.

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Section: Discussionmentioning

confidence: 69%

“…Wild-type AcMNPV was obtained from Pharmingen (BD Biosciences). The AcCAGeGFP virus was constructed previously (42). AcT7IRESmCherry was generated using the Bac-to-Bac system.…”

Section: Methodsmentioning

confidence: 99%

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Baculovirus Transduction in Mammalian Cells Is Affected by the Production of Type I and III Interferons, Which Is Mediated Mainly by the cGAS-STING Pathway

Amalfi

Molina

Bevacqua

et al. 2020

J Virol

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“…214,215 En particular, el estudio de mutantes que portan deleciones y los correspondientes virus reparados con una nueva copia del gen, la cual ha sido insertada en un locus distinto al original, representan el enfoque genético más empleado a la fecha para la caracterización de la función molecular de genes baculovirales no anotados. 74,[216][217][218] A nivel biotecnológico, los cultivos celulares también representa una alternativa atractiva debido a su compatibilidad natural con procesos de escalado y estandarización industriales. [219][220][221][222] Para el caso de varios granulovirus (AgseGV, CpGV, PhopGV, PrGV y TnGV), se ha experimentado en el pasado con diversas líneas celulares para su producción in vitro.…”

Section: Estudio De La Replicación De Sfgv En Cultivo Celular 41 Introducciónunclassified

Caracterización de baculovirus autóctonos para el control de la oruga militar tardía (<i>Spodoptera frugiperda</i>)

Masson¹

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El cogollero del maíz, también conocido como oruga militar tardía (Spodoptera frugiperda), es una plaga voraz que actualmente presenta una distribución global y representa una amenaza para los productores agrícolas. Dos miembros de la familia Baculoviridae son capaces de infectar a esta plaga, proporcionando una alternativa para su control biológico sin incurrir en el uso excesivo de pesticidas químicos. En esta tesis se abordó la caracterización dos aislamientos argentinos de baculovirus capaces de infectar S. frugiperda (SfGV y SfMNPV) a través de distintas tecnologías ómicas. El principal objetivo de esta trabajo fue profundizar nuestro conocimiento sobre los factores genómicos y proteómicos involucrados en la adaptación de estos aislamientos nativos a su hospedador. Sumado a esto, se exploró la posibilidad de multiplicar el granulovirus SfGV en cultivos celulares, de forma similar a las metodologías empleadas con nucleopoliedrovirus. En última instancia, se espera que los conocimientos y datos generados en esta tesis contribuyan a mejorar la aplicabilidad de estos baculovirus como biopesticidas. El estudio de la composición proteica de la partícula infectiva de granulovirus se desarrolla en el capítulo 2. En el capítulo 3 se discute la diversidad genética encontrada dentro de un aislamiento argentino de SfMNPV y sus posibles consecuencias en la adaptación del virus a su hospedador. En el capítulo 4 se describen los resultados preliminares de la caracterización del ciclo infectivo de SfGV en cultivos celulares de insecto. Por último, en el capítulo 5 se provee una discusión global de los resultados obtenidos y se proyectan las perspectivas de futuros estudios para caracterizar una proteína con capacidad agregante en SfGV y una colagenasa conservada en SfMNPV.

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Architecture of the baculovirus nucleocapsid revealed by cryo-EM

Jia,

Gao,

Huang

et al. 2023

Nat Commun

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Baculovirus Autographa californica multiple nucleopolyhedrovirus (AcMNPV) has been widely used as a bioinsecticide and a protein expression vector. Despite their importance, very little is known about the structure of most baculovirus proteins. Here, we show a 3.2 Å resolution structure of helical cylindrical body of the AcMNPV nucleocapsid, composed of VP39, as well as 4.3 Å resolution structures of both the head and the base of the nucleocapsid composed of over 100 protein subunits. AcMNPV VP39 demonstrates some features of the HK97-like fold and utilizes disulfide-bonds and a set of interactions at its C-termini to mediate nucleocapsid assembly and stability. At both ends of the nucleocapsid, the VP39 cylinder is constricted by an outer shell ring composed of proteins AC104, AC142 and AC109. AC101(BV/ODV-C42) and AC144(ODV-EC27) form a C14 symmetric inner layer at both capsid head and base. In the base, these proteins interact with a 7-fold symmetric capsid plug, while a portal-like structure is seen in the central portion of head. Additionally, we propose an application of AlphaFold2 for model building in intermediate resolution density.

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Effects of deletion of the ac109 gene of Autographa californica nucleopolyhedrovirus on interactions with mammalian cells

Cited by 3 publications

References 28 publications

Baculovirus Transduction in Mammalian Cells Is Affected by the Production of Type I and III Interferons, Which Is Mediated Mainly by the cGAS-STING Pathway

Baculovirus Transduction in Mammalian Cells Is Affected by the Production of Type I and III Interferons, Which Is Mediated Mainly by the cGAS-STING Pathway

Caracterización de baculovirus autóctonos para el control de la oruga militar tardía (<i>Spodoptera frugiperda</i>)

Architecture of the baculovirus nucleocapsid revealed by cryo-EM

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