Abstract
Dengue is a viral disease that represents a significant threat to global public health since billions of people are now at risk of infection by this mosquito-borne virus. The implementation of extensive screening tests is indispensable to control this disease, and the Dengue virus non-structural protein 1 (NS1) is a promising antigen for the serological diagnosis of dengue fever. Plant-based systems can be a safe and cost-effective alternative for the production of dengue virus antigens. In this work, two strategies to produce the dengue NS1 protein in Nicotiana benthamiana leaves were evaluated: Targeting NS1 to five different subcellular compartments to assess the best subcellular organelle for the expression and accumulation of NS1, and the addition of elastin-like polypeptide (ELP) or hydrophobin (HFBI) fusion tags to NS1. The transiently expressed proteins in N. benthamiana were quantified by Western blot analysis. The NS1 fused to ELP and targeted to the ER (NS1 ELP-ER) showed the highest yield (445 mg/kg), approximately a forty-fold increase in accumulation levels compared to the non-fused protein (NS1-ER), representing the first example of transient expression of DENV NS1 in plant. We also demonstrated that NS1 ELP-ER was successfully recognized by a monoclonal anti-dengue virus NS1 glycoprotein antibody, and by sera from dengue virus-infected patients. Interestingly, it was found that transient production of NS1-ER and NS1 ELP-ER using vacuum infiltration of whole plants, which is easier to scale up, rather than syringe infiltration of leaves, greatly improved the accumulation of NS1 proteins. The generated plant made NS1, even without extensive purification, showed potential to be used for the development of the NS1 diagnostic tests in resource-limited areas where dengue is endemic.
Highlights
The Dengue virus (DENV) is a positive-sense RNA virus belonging to the Flaviviridae family and to the genus Flavivirus
Each of the seven different constructs of non-structural protein 1 (NS1) were transiently expressed into the leaves of 6- to 7-weekold N. benthamiana plants infiltrated with a construct containing the suppressor of gene silencing, p19 (Silhavy et al, 2002)
Our results show that when the endoplasmic reticulum (ER)-retention signal KDEL was fused to the C-terminus of DENV NS1, the highest level of protein accumulation was achieved
Summary
The Dengue virus (DENV) is a positive-sense RNA virus belonging to the Flaviviridae family and to the genus Flavivirus. The virus is the etiologic agent of dengue fever, a neglected disease which incidence has increased 30-fold over the last 50 years (Guzman and Harris, 2014). The Dengvaxia vaccine developed by Sanofi Pasteur was licensed in some countries, including Brazil, Mexico, the Philippines, Indonesia, Costa Rica, Paraguay, and El Salvador. It displayed low efficacy against serotype 2, and it has shown declined protection against DENV, especially in seronegative individuals (Hadinegoro et al, 2015; Halstead and Russell, 2016)
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
