Abstract
The skeleton forms from multipotent human mesenchymal stem cells (hMSCs) competent to commit to specific lineages. Long noncoding RNAs (lncRNAs) have been identified as key epigenetic regulators of tissue development. However, regulation of osteogenesis by lncRNAs as mediators of commitment to the bone phenotype is largely unexplored. We focused on LINC01638, which is highly expressed in hMSCs and has been studied in cancers, but not in regulating osteogenesis. We demonstrated that LINC01638 promotes initiation of the osteoblast phenotype. Our findings reveal that LINC01638 is present at low levels during the induction of osteoblast differentiation. CRISPRi knockdown of LINC01638 in MSCs prevents osteogenesis and alkaline phosphatase expression, inhibiting osteoblast differentiation. This resulted in decreased MSC growth rate, accompanied by double-strand breaks, DNA damage, and cell senescence. Transcriptome profiling of control and LINC01638-depleted hMSCs identified > 2000 differentially expressed mRNAs related to cell cycle, cell division, spindle formation, DNA repair, and osteogenesis. Using ChIRP-qPCR, molecular mechanisms of chromatin interactions revealed the LINC01638 locus (Chr 22) includes many lncRNAs and bone-related genes. These novel findings identify the obligatory role for LINC01638 to sustain MSC pluripotency regulating osteoblast commitment and growth, as well as for physiological remodeling of bone tissue.
| Original language | English |
|---|---|
| Article number | 20314 |
| Journal | Scientific Reports |
| Volume | 13 |
| Issue number | 1 |
| DOIs | |
| State | Published - 20 Nov 2023 |
| Externally published | Yes |
Bibliographical note
Publisher Copyright:© 2023, The Author(s).
Funding
This work is supported by National Institutes of Health grants R01AR039588 to Gary Stein and Jane Lian, R37DE012528 to Jane Lian, 5R01DE029311 to Jane Lian, Gary Stein and Janet Stein; the Arthur Jason Perelman Professorship and the Charlotte Perelman Cancer Fund, and U54GM115516 for the Northern New England Clinical and Translational Research network. The next-generation sequencing was performed in the Vermont Integrative Genomics Resource Massively Parallel Sequencing Facility and was supported by the University of Vermont Cancer Center, and the UVM Larner College of Medicine. Imaging work was performed at the Microscopy Imaging Center at the University of Vermont. We would like to thank the Harry Hood Bassett Flow Cytometry and Cell Sorting Facility at the University of Vermont Larner College of Medicine for the use of BDFACSAria and BD LSRII in the generation of our flow cytometry data. The BD FACSAria is supported by NIH grant S10ODO18175. We would also like to thank Dr. Roxana del Rio-Guerra for assistance with the flow cytometry experiments. Also thank you to Joseph Boyd for support with bioinformatics analysis. This work is supported by National Institutes of Health grants R01AR039588 to Gary Stein and Jane Lian, R37DE012528 to Jane Lian, 5R01DE029311 to Jane Lian, Gary Stein and Janet Stein; the Arthur Jason Perelman Professorship and the Charlotte Perelman Cancer Fund, and U54GM115516 for the Northern New England Clinical and Translational Research network. The next-generation sequencing was performed in the Vermont Integrative Genomics Resource Massively Parallel Sequencing Facility and was supported by the University of Vermont Cancer Center, and the UVM Larner College of Medicine. Imaging work was performed at the Microscopy Imaging Center at the University of Vermont. We would like to thank the Harry Hood Bassett Flow Cytometry and Cell Sorting Facility at the University of Vermont Larner College of Medicine for the use of BDFACSAria and BD LSRII in the generation of our flow cytometry data. The BD FACSAria is supported by NIH grant S10ODO18175. We would also like to thank Dr. Roxana del Rio-Guerra for assistance with the flow cytometry experiments. Also thank you to Joseph Boyd for support with bioinformatics analysis.
| Funders | Funder number |
|---|---|
| Charlotte Perelman Cancer Fund | U54GM115516 |
| Harry Hood Bassett Flow Cytometry | |
| Northern New England Clinical and Translational Research network | |
| University of Vermont Cancer Center | |
| National Institutes of Health | 5R01DE029311, R37DE012528, S10ODO18175, R01AR039588 |
