Optimization of miR-22 expression cassette for rAAV delivery on diabetes
Abstract MicroRNA-22 (miR-22) was suggested to be important for type 2 diabetes but its functions for this disease remained unclear. Recombinant adeno-associated virus (rAAV)-mediated miR delivery is a powerful approach to study miR functions in vivo, however, the overexpression of miR-22 by rAAV re...
| Main Authors: | , , , , , , , , , , , , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
Springer
2022-01-01
|
| Series: | Molecular Biomedicine |
| Subjects: | |
| Online Access: | https://doi.org/10.1186/s43556-021-00063-y |
| _version_ | 1818979415349002240 |
|---|---|
| author | Li Yang Wenya Du Zhaoyue Zheng Li Wang Lin Xiao Qingzhe Yang Qiukui Hao Jiao Zhou Jintao Du Jun Li C. Alexander Valencia Birong Dong Hoi Yee Chow Xianghui Fu Biao Dong |
| author_facet | Li Yang Wenya Du Zhaoyue Zheng Li Wang Lin Xiao Qingzhe Yang Qiukui Hao Jiao Zhou Jintao Du Jun Li C. Alexander Valencia Birong Dong Hoi Yee Chow Xianghui Fu Biao Dong |
| author_sort | Li Yang |
| collection | DOAJ |
| description | Abstract MicroRNA-22 (miR-22) was suggested to be important for type 2 diabetes but its functions for this disease remained unclear. Recombinant adeno-associated virus (rAAV)-mediated miR delivery is a powerful approach to study miR functions in vivo, however, the overexpression of miR-22 by rAAV remains challenging because it is one of the most abundant miRs in the liver. In this study, a series of expression cassettes were designed and compared. It was shown that different lengths of primary miR-22 were overexpressed in HEK293 and HeLa cells but the longer ones were more efficiently expressed. miR-22 may be placed in either introns or the 3′ UTR of a transgene for efficient overexpression. RNA polymerase III or II promoters were successfully utilized for miR expression but the latter showed higher expression levels in cell lines. Specifically, miR-22 was expressed efficiently together with an EGFP gene. After screening, a liver-specific TTR promoter was chosen to overexpress miR-22 in diabetic mice fed a high-fat diet. It was shown that miR-22 was overexpressed 2-3 folds which improved the insulin sensitivity significantly. The approach utilized in this study to optimize miR overexpression is a powerful tool for the creation of efficient rAAV vectors for the other miRs. |
| first_indexed | 2024-12-20T16:59:10Z |
| format | Article |
| id | doaj.art-c41809fd0a524c129af69b32ce4f2e29 |
| institution | Directory Open Access Journal |
| issn | 2662-8651 |
| language | English |
| last_indexed | 2024-12-20T16:59:10Z |
| publishDate | 2022-01-01 |
| publisher | Springer |
| record_format | Article |
| series | Molecular Biomedicine |
| spelling | doaj.art-c41809fd0a524c129af69b32ce4f2e292022-12-21T19:32:37ZengSpringerMolecular Biomedicine2662-86512022-01-013111110.1186/s43556-021-00063-yOptimization of miR-22 expression cassette for rAAV delivery on diabetesLi Yang0Wenya Du1Zhaoyue Zheng2Li Wang3Lin Xiao4Qingzhe Yang5Qiukui Hao6Jiao Zhou7Jintao Du8Jun Li9C. Alexander Valencia10Birong Dong11Hoi Yee Chow12Xianghui Fu13Biao Dong14Department of Geriatrics and National Clinical Research Center for Geriatrics, West China Hospital, Sichuan UniversityState Key Laboratory of Biotherapy, West China Hospital, Sichuan UniversityState Key Laboratory of Biotherapy, West China Hospital, Sichuan UniversityState Key Laboratory of Biotherapy, West China Hospital, Sichuan UniversityState Key Laboratory of Biotherapy, West China Hospital, Sichuan UniversityState Key Laboratory of Biotherapy, West China Hospital, Sichuan UniversityDepartment of Geriatrics and National Clinical Research Center for Geriatrics, West China Hospital, Sichuan UniversityDepartment of Geriatrics and National Clinical Research Center for Geriatrics, West China Hospital, Sichuan UniversityDepartment of Otorhinolaryngology Head & Neck Surgery, West China Hospital, Sichuan UniversityDepartment of Geriatrics and National Clinical Research Center for Geriatrics, West China Hospital, Sichuan UniversityDepartment of Geriatrics and National Clinical Research Center for Geriatrics, West China Hospital, Sichuan UniversityDepartment of Geriatrics and National Clinical Research Center for Geriatrics, West China Hospital, Sichuan UniversityDepartment of Geriatrics and National Clinical Research Center for Geriatrics, West China Hospital, Sichuan UniversityState Key Laboratory of Biotherapy, West China Hospital, Sichuan UniversityDepartment of Geriatrics and National Clinical Research Center for Geriatrics, West China Hospital, Sichuan UniversityAbstract MicroRNA-22 (miR-22) was suggested to be important for type 2 diabetes but its functions for this disease remained unclear. Recombinant adeno-associated virus (rAAV)-mediated miR delivery is a powerful approach to study miR functions in vivo, however, the overexpression of miR-22 by rAAV remains challenging because it is one of the most abundant miRs in the liver. In this study, a series of expression cassettes were designed and compared. It was shown that different lengths of primary miR-22 were overexpressed in HEK293 and HeLa cells but the longer ones were more efficiently expressed. miR-22 may be placed in either introns or the 3′ UTR of a transgene for efficient overexpression. RNA polymerase III or II promoters were successfully utilized for miR expression but the latter showed higher expression levels in cell lines. Specifically, miR-22 was expressed efficiently together with an EGFP gene. After screening, a liver-specific TTR promoter was chosen to overexpress miR-22 in diabetic mice fed a high-fat diet. It was shown that miR-22 was overexpressed 2-3 folds which improved the insulin sensitivity significantly. The approach utilized in this study to optimize miR overexpression is a powerful tool for the creation of efficient rAAV vectors for the other miRs.https://doi.org/10.1186/s43556-021-00063-yRecombinant adeno-associated virusMicroRNAExpression cassettesMiR-22Insulin |
| spellingShingle | Li Yang Wenya Du Zhaoyue Zheng Li Wang Lin Xiao Qingzhe Yang Qiukui Hao Jiao Zhou Jintao Du Jun Li C. Alexander Valencia Birong Dong Hoi Yee Chow Xianghui Fu Biao Dong Optimization of miR-22 expression cassette for rAAV delivery on diabetes Molecular Biomedicine Recombinant adeno-associated virus MicroRNA Expression cassettes MiR-22 Insulin |
| title | Optimization of miR-22 expression cassette for rAAV delivery on diabetes |
| title_full | Optimization of miR-22 expression cassette for rAAV delivery on diabetes |
| title_fullStr | Optimization of miR-22 expression cassette for rAAV delivery on diabetes |
| title_full_unstemmed | Optimization of miR-22 expression cassette for rAAV delivery on diabetes |
| title_short | Optimization of miR-22 expression cassette for rAAV delivery on diabetes |
| title_sort | optimization of mir 22 expression cassette for raav delivery on diabetes |
| topic | Recombinant adeno-associated virus MicroRNA Expression cassettes MiR-22 Insulin |
| url | https://doi.org/10.1186/s43556-021-00063-y |
| work_keys_str_mv | AT liyang optimizationofmir22expressioncassetteforraavdeliveryondiabetes AT wenyadu optimizationofmir22expressioncassetteforraavdeliveryondiabetes AT zhaoyuezheng optimizationofmir22expressioncassetteforraavdeliveryondiabetes AT liwang optimizationofmir22expressioncassetteforraavdeliveryondiabetes AT linxiao optimizationofmir22expressioncassetteforraavdeliveryondiabetes AT qingzheyang optimizationofmir22expressioncassetteforraavdeliveryondiabetes AT qiukuihao optimizationofmir22expressioncassetteforraavdeliveryondiabetes AT jiaozhou optimizationofmir22expressioncassetteforraavdeliveryondiabetes AT jintaodu optimizationofmir22expressioncassetteforraavdeliveryondiabetes AT junli optimizationofmir22expressioncassetteforraavdeliveryondiabetes AT calexandervalencia optimizationofmir22expressioncassetteforraavdeliveryondiabetes AT birongdong optimizationofmir22expressioncassetteforraavdeliveryondiabetes AT hoiyeechow optimizationofmir22expressioncassetteforraavdeliveryondiabetes AT xianghuifu optimizationofmir22expressioncassetteforraavdeliveryondiabetes AT biaodong optimizationofmir22expressioncassetteforraavdeliveryondiabetes |