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Viruses Dec 2022Baculoviruses are enveloped, insect-specific viruses with large double-stranded DNA genomes. Among all the baculovirus species, Autographa californica multiple... (Review)
Review
Baculoviruses are enveloped, insect-specific viruses with large double-stranded DNA genomes. Among all the baculovirus species, Autographa californica multiple nucleopolyhedrovirus (AcMNPV) is the most studied. Due to its characteristics regarding biosafety, narrow host range and the availability of different platforms for modifying its genome, AcMNPV has become a powerful biotechnological tool. In this review, we will address the most widespread technological applications of baculoviruses. We will begin by summarizing their natural cycle both in larvae and in cell culture and how it can be exploited. Secondly, we will explore the different baculovirus-based protein expression systems (BEVS) and their multiple applications in the pharmaceutical and biotechnological industry. We will focus particularly on the production of vaccines, many of which are either currently commercialized or in advanced stages of development (e.g., Novavax, COVID-19 vaccine). In addition, recombinant baculoviruses can be used as efficient gene transduction and protein expression vectors in vertebrate cells (e.g., BacMam). Finally, we will extensively describe various gene therapy strategies based on baculoviruses applied to the treatment of different diseases. The main objective of this work is to provide an extensive up-to-date summary of the different biotechnological applications of baculoviruses, emphasizing the genetic modification strategies used in each field.
Topics: Animals; Humans; Baculoviridae; COVID-19 Vaccines; COVID-19; Nucleopolyhedroviruses; Spodoptera
PubMed: 36680120
DOI: 10.3390/v15010080 -
Annual Review of Entomology Jan 2023Silkworm () is not only an economic insect but also a model organism for life science research. nucleopolyhedrovirus (BmNPV) disease is a major infectious disease in... (Review)
Review
Silkworm () is not only an economic insect but also a model organism for life science research. nucleopolyhedrovirus (BmNPV) disease is a major infectious disease in the world's sericulture industry. The cocoon loss caused by this disease accounts for more than 60% of the total loss caused by all silkworm diseases. To date, there has been no effective solution for preventing and treating this disease. The most effective measure is to breed disease-resistant varieties. The quickest way to breed disease-resistant varieties is to apply genetic modification. However, this requires that we obtain disease resistance genes and know the mechanism of disease resistance. Since the discovery of disease-resistant resources in 1989, scholars in the sericulture industry around the world have been inspired to search for resistance genes. In the past two decades, with the help of multi-omics technologies, screening of resistance genes, gene localization, protein modification, virus-host interactions, etc., researchers have found some candidate genes that have been proposed to function at the cellular or individual level. Several disease-resistant varieties have been obtained and used in production through hybrid breeding, RNA interference, and genetic modification. This article summarizes and reviews the discovery of and research advances related to silkworm resistance to BmNPV. It is anticipated that the review will inspire scientific researchers to continue searching for disease resistance genes, clarify the molecular mechanism of silkworm disease resistance, and promote disease-resistant silkworm breeding.
Topics: Animals; Baculoviridae; Bombyx; Disease Resistance; Nucleopolyhedroviruses
PubMed: 36689303
DOI: 10.1146/annurev-ento-120220-112317 -
Viruses Apr 2023Accurate and rapid quantification of (infectious) virus titers is of paramount importance in the manufacture of viral vectors and vaccines. Reliable quantification data...
Accurate and rapid quantification of (infectious) virus titers is of paramount importance in the manufacture of viral vectors and vaccines. Reliable quantification data allow efficient process development at a laboratory scale and thorough process monitoring in later production. However, current gold standard applications, such as endpoint dilution assays, are cumbersome and do not provide true process analytical monitoring. Accordingly, flow cytometry and quantitative polymerase chain reaction have attracted increasing interest in recent years, offering various advantages for rapid quantification. Here, we compared different approaches for the assessment of infectious viruses, using a model baculovirus. Firstly, infectivity was estimated by the quantification of viral nucleic acids in infected cells, and secondly, different flow cytometric approaches were investigated regarding analysis times and calibration ranges. The flow cytometry technique included a quantification based on post-infection fluorophore expression and labeling of a viral surface protein using fluorescent antibodies. Additionally, the possibility of viral (m)RNA labeling in infected cells was investigated as a proof of concept. The results confirmed that infectivity assessment based on qPCR is not trivial and requires sophisticated method optimization, whereas staining of viral surface proteins is a fast and feasible approach for enveloped viruses. Finally, labeling of viral (m)RNA in infected cells appears to be a promising opportunity but will require further research.
Topics: Baculoviridae; Genetic Vectors; RNA
PubMed: 37112978
DOI: 10.3390/v15040998 -
Virus Research Dec 2022Transfer RNAs (tRNAs) genes are both coded for and arranged along some viral genomes representing the entire virosphere and seem to play different biological functions...
Transfer RNAs (tRNAs) genes are both coded for and arranged along some viral genomes representing the entire virosphere and seem to play different biological functions during infection, other than transferring the correct amino acid to a growing peptide chain. Baculovirus genome description and annotation has focused mostly on protein-coding genes, microRNA, and homologous regions. Here we carried out a large-scale in silico search for putative tRNA genes in baculovirus genomes. Ninety-six of 257 baculovirus genomes analyzed was found to contain at least one putative tRNA gene. We found great diversity in primary and secondary structure, in location within the genome, in intron presence and size, and in anti-codon identity. In some cases, genes of tRNA-containing genomes were found to have a bias for the codons specified by the tRNAs present in such genomes. Moreover, analysis revealed that most of the putative tRNA genes possessed conserved motifs for tRNA type 2 promoters, including the A-box and B-box motifs with few mismatches from the eukaryotic canonical motifs. From publicly available small RNA deep sequencing datasets of baculovirus-infected insect cells, we found evidence that a putative Autographa californica multiple nucleopolyhedrovirus Gln-tRNA gene was transcribed and modified with the addition of the non-templated 3'-CCA tail found at the end of all tRNAs. Further research is needed to determine the expression and functionality of these viral tRNAs.
Topics: Baculoviridae; RNA, Transfer; Eukaryota; Base Sequence; Codon
PubMed: 36181979
DOI: 10.1016/j.virusres.2022.198949 -
Current Issues in Molecular Biology 2020Over 35 years since it was established to make recombinant proteins, the baculovirus expression vector system continues to develop and improve. Early systems for... (Review)
Review
Over 35 years since it was established to make recombinant proteins, the baculovirus expression vector system continues to develop and improve. Early systems for recombinant virus selection were laborious, but better methods were rapidly devised that enabled non-virologists to use baculovirus vectors successfully in a wide range of applications. These applications include multiple gene expression for complex molecules, production of adeno-associated virus-like particles for gene therapy, the use of baculovirus budded virus for the same purpose, numerous potential human and animal vaccines, and for other therapeutic proteins. A number of products for human and veterinary use are now on the market, which attests to the utility of the systems. Despite these successes, baculovirus vectors essentially remain in a relatively primitive state of development. Many proteins, particularly membrane-bound or secreted products, continue to be difficult to produce. Various research groups are working to identify potential areas of improvement, which if combined into an ideal vector might offer considerable advances to the system. This chapter will review some of the most recent reports and highlight those that might have generic application for recombinant protein synthesis in insect cells. We also summarize parallel developments in host cells used for baculovirus expression and how culture conditions can influence protein production.
Topics: Animals; Baculoviridae; Gene Expression; Genetic Engineering; Genetic Vectors; Humans; Protein Engineering; RNA Interference; RNA, Small Interfering; Recombinant Proteins
PubMed: 31167962
DOI: 10.21775/cimb.034.215 -
Molecular Therapy : the Journal of the... Apr 2013Insect-derived baculoviruses have emerged as versatile and safe workhorses of biotechnology. Baculovirus expression vectors (BEVs) have been applied widely for crop and... (Review)
Review
Insect-derived baculoviruses have emerged as versatile and safe workhorses of biotechnology. Baculovirus expression vectors (BEVs) have been applied widely for crop and forest protection, as well as safe tools for recombinant protein production in insect cells. However, BEVs ability to efficiently transduce noninsect cells is still relatively poorly recognized despite the fact that efficient baculovirus-mediated in vitro and ex vivo gene delivery into dormant and dividing vertebrate cells of diverse origin has been described convincingly by many authors. Preliminary proof of therapeutic potential has also been established in preclinical studies. This review summarizes the advantages and current status of baculovirus-mediated gene delivery. Stem cell transduction, preclinical animal studies, tissue engineering, vaccination, cancer gene therapy, viral vector production, and drug discovery are covered.
Topics: Animals; Baculoviridae; Gene Transfer Techniques; Genetic Therapy; Genetic Vectors; Humans; Insect Vectors
PubMed: 23439502
DOI: 10.1038/mt.2012.286 -
Genes Apr 2022Apoptosis plays an important role in virus-host interactions and is a major element of the insect immune response. Exploring the regulatory mechanisms of virus-induced...
Apoptosis plays an important role in virus-host interactions and is a major element of the insect immune response. Exploring the regulatory mechanisms of virus-induced apoptosis through the expression of apoptotic genes holds important research and application value. Functional research on the reported inhibitor of apoptosis proteins (IAPs) mainly focuses on the group I baculovirus, while the functions of the group II baculovirus IAPs remains unclear. To explore its role in the regulation of the apoptosis of insect cells, we constructed the transient expression vector (pIE1 vectors) and the recombinant baculovirus expressing genes (from the nucleopolyhedrovirus) of the group II baculovirus. Apoptosis gene expression results and the virus-induced apoptosis rate show that the overexpression of BsIAP1 could promote apoptosis in insect cells. However, the overexpression of BsIAP2 and BsIAP3 decreases the expression of apoptotic genes, revealing an inhibitory effect. Results on the impact of baculovirus-induced apoptosis also confirm that BsIAP1 reduces viral nucleocapsid expression and the baculovirus titer, while BsIAP2 and BsIAP3 increase them significantly. Furthermore, compared with single expression, the co-expression of BsIAP2 and BsIAP3 significantly reduces the rate of virus-induced apoptosis and improves the expression of nucleocapsids and the titer of offspring virus, indicating the synergistic effect on BsIAP2 and BsIAP3. In addition, combined expression of all three BsIAPs significantly reduced levels of intracellular apoptosis-related genes (including apoptosis and anti-apoptosis genes), as well as apoptosis rate and progeny virus titer, indicating that life activities in insect cells are also inhibited. These findings reveal the relationship between apoptosis and group II baculovirus IAP, which provide an experimental and theoretical basis for further exploration of the molecular mechanism between group II baculoviruses and insect cells.
Topics: Animals; Apoptosis; Baculoviridae; Inhibitor of Apoptosis Proteins; Insecta; Nucleopolyhedroviruses
PubMed: 35627135
DOI: 10.3390/genes13050750 -
Applied Microbiology and Biotechnology Nov 2021Baculoviruses are insect pathogens widely used as biotechnological tools in different fields of life sciences and technologies. The particular biology of these entities... (Review)
Review
Baculoviruses are insect pathogens widely used as biotechnological tools in different fields of life sciences and technologies. The particular biology of these entities (biosafety viruses 1; large circular double-stranded DNA genomes, infective per se; generally of narrow host range on insect larvae; many of the latter being pests in agriculture) and the availability of molecular-biology procedures (e.g., genetic engineering to edit their genomes) and cellular resources (availability of cell lines that grow under in vitro culture conditions) have enabled the application of baculoviruses as active ingredients in pest control, as systems for the expression of recombinant proteins (Baculovirus Expression Vector Systems-BEVS) and as viral vectors for gene delivery in mammals or to display antigenic proteins (Baculoviruses applied on mammals-BacMam). Accordingly, BEVS and BacMam technologies have been introduced in academia because of their availability as commercial systems and ease of use and have also reached the human pharmaceutical industry, as incomparable tools in the development of biological products such as diagnostic kits, vaccines, protein therapies, and-though still in the conceptual stage involving animal models-gene therapies. Among all the baculovirus species, the Autographa californica multiple nucleopolyhedrovirus has been the most highly exploited in the above utilities for the human-biotechnology field. This review highlights the main achievements (in their different stages of development) of the use of BEVS and BacMam technologies for the generation of products for infectious and noninfectious human diseases. KEY POINTS: • Baculoviruses can assist as biotechnological tools in human health problems. • Vaccines and diagnosis reagents produced in the baculovirus platform are described. • The use of recombinant baculovirus for gene therapy-based treatment is reviewed.
Topics: Animals; Baculoviridae; Cell Line; Genetic Vectors; Humans; Insecta; Recombinant Proteins
PubMed: 34618205
DOI: 10.1007/s00253-021-11615-1 -
Advances in Experimental Medicine and... 2016Multiprotein complexes regulate most if not all cellular functions. Elucidating the structure and function of these complex cellular machines is essential for... (Review)
Review
Multiprotein complexes regulate most if not all cellular functions. Elucidating the structure and function of these complex cellular machines is essential for understanding biology. Moreover, multiprotein complexes by themselves constitute powerful reagents as biologics for the prevention and treatment of human diseases. Recombinant production by the baculovirus/insect cell expression system is particularly useful for expressing proteins of eukaryotic origin and their complexes. MultiBac, an advanced baculovirus/insect cell system, has been widely adopted in the last decade to produce multiprotein complexes with many subunits that were hitherto inaccessible, for academic and industrial research and development. The MultiBac system, its development and numerous applications are presented. Future opportunities for utilizing MultiBac to catalyze discovery are outlined.
Topics: Animals; Baculoviridae; Computational Biology; Databases, Protein; Drug Discovery; Gene Expression Regulation, Viral; Genetic Vectors; Humans; Models, Molecular; Multiprotein Complexes; Protein Engineering; Protein Multimerization; Protein Structure, Quaternary; Protein Subunits; Recombinant Proteins; Structure-Activity Relationship; Transcription, Genetic; Transfection; Viral Proteins
PubMed: 27165327
DOI: 10.1007/978-3-319-27216-0_13 -
PLoS Pathogens 2013