CRISPR activation identifies a novel miR-2861 binding site that facilitates the osteogenesis of human mesenchymal stem cells
Abstract We investigated the regulation of histone deacetylases (HDACs) by miR-2861 in the osteoblastic differentiation of human mesenchymal stem cells (MSCs) and miR-2861 binding site by CRISPR activation (CRISPRa). Transfection of miR-2861 into human MSCs was performed and the effect on osteoblast...
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| Format: | Article |
| Language: | English |
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BMC
2024-11-01
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| Series: | Journal of Orthopaedic Surgery and Research |
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| Online Access: | https://doi.org/10.1186/s13018-024-05163-3 |
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| author | Seong-Ho Park Jungwoo Kim Hee-Jin Yang Ju Yeon Lee Chi Heon Kim Junho K. Hur Sung Bae Park |
| author_facet | Seong-Ho Park Jungwoo Kim Hee-Jin Yang Ju Yeon Lee Chi Heon Kim Junho K. Hur Sung Bae Park |
| author_sort | Seong-Ho Park |
| collection | DOAJ |
| description | Abstract We investigated the regulation of histone deacetylases (HDACs) by miR-2861 in the osteoblastic differentiation of human mesenchymal stem cells (MSCs) and miR-2861 binding site by CRISPR activation (CRISPRa). Transfection of miR-2861 into human MSCs was performed and the effect on osteoblast differentiation was analyzed. Using catalytically inactive Cas12a, the CRISPRa system induced targeted overexpression of endogenous miRNA and repressed the luciferase activities of reporters that contained functional miRNA target sites. The delivery of miR-2861 into MSCs enhanced osteoblast differentiation by decreased expressions of the HDAC1, 4 and 5 genes. The mechanism of HDAC5 repression by miR-2861 in humans has not been fully elucidated. To this end, the HDAC5 mRNA sequence was analyzed and a putative primate-specific miR-2861 binding site was identified in the 3’ untranslated region (3’-UTR). CRISPRa was applied to validate the putative binding site and an increase in endogenous miR-2861 was found to repress the expression of a reporter that contained the novel miR-2861 binding site. The delivery of miR-2861 to human MSCs enhanced osteoblast differentiation. In the 3’-UTR, the HDAC5 repression was mediated by the miR-2861 binding site, and miR-2861 promoted osteoblast differentiation via the inhibition of HDAC5 through a primate-specific miRNA binding site. Therefore, miRNAmiR-2861 with the CRISPRa methods might be a good biomaterial for osteogenesis augmentation. |
| format | Article |
| id | doaj-art-bd5020c75b0546cfad82b9f47d7ba51e |
| institution | Kabale University |
| issn | 1749-799X |
| language | English |
| publishDate | 2024-11-01 |
| publisher | BMC |
| record_format | Article |
| series | Journal of Orthopaedic Surgery and Research |
| spelling | doaj-art-bd5020c75b0546cfad82b9f47d7ba51e2024-11-24T12:36:00ZengBMCJournal of Orthopaedic Surgery and Research1749-799X2024-11-011911910.1186/s13018-024-05163-3CRISPR activation identifies a novel miR-2861 binding site that facilitates the osteogenesis of human mesenchymal stem cellsSeong-Ho Park0Jungwoo Kim1Hee-Jin Yang2Ju Yeon Lee3Chi Heon Kim4Junho K. Hur5Sung Bae Park6Department of Medicine, Major in Medical Genetics, Graduate School, Hanyang UniversityDepartment of Neurosurgery, Seoul National University Boramae Medical CenterDepartment of Neurosurgery, Seoul National University Boramae Medical CenterHanyang Institute of Bioscience and Biotechnology, Hanyang UniversityDepartment of Neurosurgery, Seoul National University College of MedicineHanyang Institute of Bioscience and Biotechnology, Hanyang UniversityDepartment of Neurosurgery, Seoul National University Boramae Medical CenterAbstract We investigated the regulation of histone deacetylases (HDACs) by miR-2861 in the osteoblastic differentiation of human mesenchymal stem cells (MSCs) and miR-2861 binding site by CRISPR activation (CRISPRa). Transfection of miR-2861 into human MSCs was performed and the effect on osteoblast differentiation was analyzed. Using catalytically inactive Cas12a, the CRISPRa system induced targeted overexpression of endogenous miRNA and repressed the luciferase activities of reporters that contained functional miRNA target sites. The delivery of miR-2861 into MSCs enhanced osteoblast differentiation by decreased expressions of the HDAC1, 4 and 5 genes. The mechanism of HDAC5 repression by miR-2861 in humans has not been fully elucidated. To this end, the HDAC5 mRNA sequence was analyzed and a putative primate-specific miR-2861 binding site was identified in the 3’ untranslated region (3’-UTR). CRISPRa was applied to validate the putative binding site and an increase in endogenous miR-2861 was found to repress the expression of a reporter that contained the novel miR-2861 binding site. The delivery of miR-2861 to human MSCs enhanced osteoblast differentiation. In the 3’-UTR, the HDAC5 repression was mediated by the miR-2861 binding site, and miR-2861 promoted osteoblast differentiation via the inhibition of HDAC5 through a primate-specific miRNA binding site. Therefore, miRNAmiR-2861 with the CRISPRa methods might be a good biomaterial for osteogenesis augmentation.https://doi.org/10.1186/s13018-024-05163-3CRISPRMicroRNAOsteogenesisHistone deacetylase |
| spellingShingle | Seong-Ho Park Jungwoo Kim Hee-Jin Yang Ju Yeon Lee Chi Heon Kim Junho K. Hur Sung Bae Park CRISPR activation identifies a novel miR-2861 binding site that facilitates the osteogenesis of human mesenchymal stem cells Journal of Orthopaedic Surgery and Research CRISPR MicroRNA Osteogenesis Histone deacetylase |
| title | CRISPR activation identifies a novel miR-2861 binding site that facilitates the osteogenesis of human mesenchymal stem cells |
| title_full | CRISPR activation identifies a novel miR-2861 binding site that facilitates the osteogenesis of human mesenchymal stem cells |
| title_fullStr | CRISPR activation identifies a novel miR-2861 binding site that facilitates the osteogenesis of human mesenchymal stem cells |
| title_full_unstemmed | CRISPR activation identifies a novel miR-2861 binding site that facilitates the osteogenesis of human mesenchymal stem cells |
| title_short | CRISPR activation identifies a novel miR-2861 binding site that facilitates the osteogenesis of human mesenchymal stem cells |
| title_sort | crispr activation identifies a novel mir 2861 binding site that facilitates the osteogenesis of human mesenchymal stem cells |
| topic | CRISPR MicroRNA Osteogenesis Histone deacetylase |
| url | https://doi.org/10.1186/s13018-024-05163-3 |
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