The effect of beta-globin intron and WPRE on transient expression of SARS-CoV-2 spike and generation of pseudotyped lentiviruses
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 (...
Saved in:
| Main Authors: | , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
Frontiers Media S.A.
2025-04-01
|
| Series: | Acta Virologica |
| Subjects: | |
| Online Access: | https://www.frontierspartnerships.org/articles/10.3389/av.2025.14262/full |
| Tags: |
Add Tag
No Tags, Be the first to tag this record!
|
| _version_ | 1849738054574538752 |
|---|---|
| author | Hyeop Jin Ye Rae Cho Yong Tae Jung |
| author_facet | Hyeop Jin Ye Rae Cho Yong Tae Jung |
| author_sort | Hyeop Jin |
| collection | DOAJ |
| description | 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. |
| format | Article |
| id | doaj-art-dc62ca3bb8a341e9b085b6e92b379d30 |
| institution | DOAJ |
| issn | 1336-2305 |
| language | English |
| publishDate | 2025-04-01 |
| publisher | Frontiers Media S.A. |
| record_format | Article |
| series | Acta Virologica |
| spelling | doaj-art-dc62ca3bb8a341e9b085b6e92b379d302025-08-20T03:06:43ZengFrontiers Media S.A.Acta Virologica1336-23052025-04-016910.3389/av.2025.1426214262The effect of beta-globin intron and WPRE on transient expression of SARS-CoV-2 spike and generation of pseudotyped lentivirusesHyeop JinYe Rae ChoYong Tae JungSevere 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.https://www.frontierspartnerships.org/articles/10.3389/av.2025.14262/fullβ-globin intronCOVID-19pseudovirusSars-CoV-2WPRE |
| spellingShingle | Hyeop Jin Ye Rae Cho Yong Tae Jung The effect of beta-globin intron and WPRE on transient expression of SARS-CoV-2 spike and generation of pseudotyped lentiviruses Acta Virologica β-globin intron COVID-19 pseudovirus Sars-CoV-2 WPRE |
| title | The effect of beta-globin intron and WPRE on transient expression of SARS-CoV-2 spike and generation of pseudotyped lentiviruses |
| title_full | The effect of beta-globin intron and WPRE on transient expression of SARS-CoV-2 spike and generation of pseudotyped lentiviruses |
| title_fullStr | The effect of beta-globin intron and WPRE on transient expression of SARS-CoV-2 spike and generation of pseudotyped lentiviruses |
| title_full_unstemmed | The effect of beta-globin intron and WPRE on transient expression of SARS-CoV-2 spike and generation of pseudotyped lentiviruses |
| title_short | The effect of beta-globin intron and WPRE on transient expression of SARS-CoV-2 spike and generation of pseudotyped lentiviruses |
| title_sort | effect of beta globin intron and wpre on transient expression of sars cov 2 spike and generation of pseudotyped lentiviruses |
| topic | β-globin intron COVID-19 pseudovirus Sars-CoV-2 WPRE |
| url | https://www.frontierspartnerships.org/articles/10.3389/av.2025.14262/full |
| work_keys_str_mv | AT hyeopjin theeffectofbetaglobinintronandwpreontransientexpressionofsarscov2spikeandgenerationofpseudotypedlentiviruses AT yeraecho theeffectofbetaglobinintronandwpreontransientexpressionofsarscov2spikeandgenerationofpseudotypedlentiviruses AT yongtaejung theeffectofbetaglobinintronandwpreontransientexpressionofsarscov2spikeandgenerationofpseudotypedlentiviruses AT hyeopjin effectofbetaglobinintronandwpreontransientexpressionofsarscov2spikeandgenerationofpseudotypedlentiviruses AT yeraecho effectofbetaglobinintronandwpreontransientexpressionofsarscov2spikeandgenerationofpseudotypedlentiviruses AT yongtaejung effectofbetaglobinintronandwpreontransientexpressionofsarscov2spikeandgenerationofpseudotypedlentiviruses |