AUTHOR=Jin Hyeop , Cho Ye Rae , Jung Yong Tae TITLE=The effect of beta-globin intron and WPRE on transient expression of SARS-CoV-2 spike and generation of pseudotyped lentiviruses JOURNAL=Acta Virologica VOLUME=Volume 69 - 2025 YEAR=2025 URL=https://www.frontierspartnerships.org/journals/acta-virologica/articles/10.3389/av.2025.14262 DOI=10.3389/av.2025.14262 ISSN=1336-2305 ABSTRACT=

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), an enveloped, single-stranded RNA virus, is causative of coronavirus disease 2019 (COVID-19). The viral entry process is facilitated by the highly antigenic spike protein of SARS-CoV-2, which binds to the angiotensin-converting enzyme 2 (ACE2) receptor. However, research involving live SARS-CoV-2 is limited to biosafety level (BSL)-3 facilities, hindering the progress in vaccine and therapeutic development. Pseudotyped viruses have emerged as valuable tools for studying entry inhibitors, fusion inhibitors, and neutralizing assays in BSL-2 facilities. Several modifications to the spike protein have been identified to enhance the yield of pseudotyped viruses. Notably, the removal of the last 19 amino acids from the spike protein and the D614G modification have been shown to increase pseudovirus titers. In this study, we aimed to identify an effective expression vector for pseudotyping by inserting SARS-CoV-2 S Δ19 into the intron-containing pcDNA3.1 vector. The inclusion of the β-globin intron sequence led to a significant augmentation in spike protein expression and a threefold improvement in pseudotyped virus production. Furthermore, we examined whether the woodchuck hepatitis virus post-transcriptional regulatory element (WPRE) could enhance spike protein expression by subcloning the spike protein gene into a lentiviral vector containing WPRE. Our results demonstrated that vectors containing the WPRE exhibited increased spike protein expression compared to vectors lacking the WPRE in HEK293-hACE2 cells. This study demonstrates that the inclusion of a β-globin intron and WPRE enhances the production of SARS-CoV-2 spike protein pseudotyped lentiviruses, a valuable tool for COVID-19 research.