Deletion of a putative promoter-proximal Tnfsf11 regulatory region in mice does not alter bone mass or Tnfsf11 expression in vivo
| Autor: | Charles A. O'Brien, Yu Liu, Jeff D. Thostenson, Ryan S. MacLeod, Jinhu Xiong, Mark B. Meyer, Melda Onal, J. Wesley Pike, Keisha M. Cawley, Nancy A. Benkusky |
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| Rok vydání: | 2021 |
| Předmět: |
Male
0301 basic medicine Vertebrae Physiology medicine.medical_treatment Osteoclasts Parathyroid hormone Artificial Gene Amplification and Extension Regulatory Sequences Nucleic Acid Biochemistry Polymerase Chain Reaction Mice 0302 clinical medicine Bone Density Animal Cells Medicine and Health Sciences Transcriptional regulation Lymphocytes Femur Promoter Regions Genetic Musculoskeletal System Cells Cultured Connective Tissue Cells Mice Knockout Mice Inbred BALB C Multidisciplinary biology Animal Models medicine.anatomical_structure Cytokine Experimental Organism Systems Parathyroid Hormone Connective Tissue Regulatory sequence RANKL 030220 oncology & carcinogenesis Models Animal Medicine Female Bone Remodeling Anatomy Cellular Types Research Article musculoskeletal diseases medicine.medical_specialty Science Bone Marrow Cells Mouse Models Research and Analysis Methods Bone resorption 03 medical and health sciences Model Organisms Osteoclast Internal medicine medicine Animals Bone Resorption Bone Molecular Biology Techniques Enhancer Molecular Biology Skeleton RANK Ligand Biology and Life Sciences Cell Biology Hormones Spine Mice Inbred C57BL Biological Tissue 030104 developmental biology Endocrinology Animal Studies biology.protein CRISPR-Cas Systems Physiological Processes |
| Zdroj: | PLoS ONE, Vol 16, Iss 5, p e0250974 (2021) PLoS ONE |
| ISSN: | 1932-6203 |
| DOI: | 10.1371/journal.pone.0250974 |
| Popis: | The cytokine RANKL is essential for osteoclast formation during physiological and pathological bone resorption. RANKL also contributes to lymphocyte production, development of lymph nodes and mammary glands, as well as other biological activities. Transcriptional control of the Tnfsf11 gene, which encodes RANKL, is complex and involves distant regulatory regions. Nevertheless, cell culture studies suggest that an enhancer region near the transcription start site is involved in the control of Tnfsf11 expression by hormones such as 1,25-(OH)2 vitamin D3 and parathyroid hormone, as well as the sympathetic nervous system. To address the significance of this region in vivo, we deleted the sequence between -510 to -1413 bp, relative to Tnfsf11 exon 1, from mice using CRISPR-based gene editing. MicroCT analysis of the femur and fourth lumbar vertebra of enhancer knockout mice showed no differences in bone mass compared to wild type littermates at 5 weeks and 6 months of age, suggesting no changes in osteoclast formation. RNA extracted from the tibia, fifth lumbar vertebra, thymus, and spleen at 6 months of age also showed no reduction in Tnfsf11 mRNA abundance between these groups. However, maximal stimulation of Tnfsf11 mRNA abundance in cultured stromal cells by PTH was reduced approximately 40% by enhancer deletion, while stimulation by 1,25-(OH)2 vitamin D3 was unaffected. The abundance of B and T lymphocytes in the bone marrow did not differ between genotypes. These results demonstrate that the region between -510 and -1413 does not contribute to Tnfsf11 expression, osteoclast support, or lymphocyte production in mice under normal physiological conditions but may be involved in situations of elevated parathyroid hormone. |
| Databáze: | OpenAIRE |
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