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Knocking out TMEM38B in human foetal osteoblasts hFOB 1.19 by CRISPR/Cas9: A model for recessive OI type XIV
Osteogenesis imperfecta (OI) type XIV is a rare recessive bone disorder characterized by variable degree of severity associated to osteopenia. It is caused by mutations in TMEM38B encoding for the trimeric intracellular cation channel TRIC-B, specific for potassium and ubiquitously present in the en...
Autores principales: | , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Public Library of Science
2021
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8478202/ https://www.ncbi.nlm.nih.gov/pubmed/34582479 http://dx.doi.org/10.1371/journal.pone.0257254 |
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author | Leoni, Laura Tonelli, Francesca Besio, Roberta Gioia, Roberta Moccia, Francesco Rossi, Antonio Forlino, Antonella |
author_facet | Leoni, Laura Tonelli, Francesca Besio, Roberta Gioia, Roberta Moccia, Francesco Rossi, Antonio Forlino, Antonella |
author_sort | Leoni, Laura |
collection | PubMed |
description | Osteogenesis imperfecta (OI) type XIV is a rare recessive bone disorder characterized by variable degree of severity associated to osteopenia. It is caused by mutations in TMEM38B encoding for the trimeric intracellular cation channel TRIC-B, specific for potassium and ubiquitously present in the endoplasmic reticulum (ER) membrane. OI type XIV molecular basis is largely unknown and, due to the rarity of the disease, the availability of patients’ osteoblasts is challenging. Thus, CRISPR/Cas9 was used to knock out (KO) TMEM38B in the human Foetal Osteoblast hFOB 1.19 to obtain an OI type XIV model. CRISPR/Cas9 is a powerful technology to generate in vitro and in vivo models for heritable disorders. Its limited cost and ease of use make this technique widely applicable in most laboratories. Nevertheless, to fully take advantage of this approach, it is important to be aware of its strengths and limitations. Three gRNAs were used and several KO clones lacking the expression of TRIC-B were obtained. Few clones were validated as good models for the disease since they reproduce the altered ER calcium flux, collagen I structure and impaired secretion and osteoblastic markers expression detected in patients’ cells. Impaired proliferation and mineralization in KO clones unveiled the relevance of TRIC-B in osteoblasts functionality. |
format | Online Article Text |
id | pubmed-8478202 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2021 |
publisher | Public Library of Science |
record_format | MEDLINE/PubMed |
spelling | pubmed-84782022021-09-29 Knocking out TMEM38B in human foetal osteoblasts hFOB 1.19 by CRISPR/Cas9: A model for recessive OI type XIV Leoni, Laura Tonelli, Francesca Besio, Roberta Gioia, Roberta Moccia, Francesco Rossi, Antonio Forlino, Antonella PLoS One Research Article Osteogenesis imperfecta (OI) type XIV is a rare recessive bone disorder characterized by variable degree of severity associated to osteopenia. It is caused by mutations in TMEM38B encoding for the trimeric intracellular cation channel TRIC-B, specific for potassium and ubiquitously present in the endoplasmic reticulum (ER) membrane. OI type XIV molecular basis is largely unknown and, due to the rarity of the disease, the availability of patients’ osteoblasts is challenging. Thus, CRISPR/Cas9 was used to knock out (KO) TMEM38B in the human Foetal Osteoblast hFOB 1.19 to obtain an OI type XIV model. CRISPR/Cas9 is a powerful technology to generate in vitro and in vivo models for heritable disorders. Its limited cost and ease of use make this technique widely applicable in most laboratories. Nevertheless, to fully take advantage of this approach, it is important to be aware of its strengths and limitations. Three gRNAs were used and several KO clones lacking the expression of TRIC-B were obtained. Few clones were validated as good models for the disease since they reproduce the altered ER calcium flux, collagen I structure and impaired secretion and osteoblastic markers expression detected in patients’ cells. Impaired proliferation and mineralization in KO clones unveiled the relevance of TRIC-B in osteoblasts functionality. Public Library of Science 2021-09-28 /pmc/articles/PMC8478202/ /pubmed/34582479 http://dx.doi.org/10.1371/journal.pone.0257254 Text en © 2021 Leoni et al https://creativecommons.org/licenses/by/4.0/This is an open access article distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. |
spellingShingle | Research Article Leoni, Laura Tonelli, Francesca Besio, Roberta Gioia, Roberta Moccia, Francesco Rossi, Antonio Forlino, Antonella Knocking out TMEM38B in human foetal osteoblasts hFOB 1.19 by CRISPR/Cas9: A model for recessive OI type XIV |
title | Knocking out TMEM38B in human foetal osteoblasts hFOB 1.19 by CRISPR/Cas9: A model for recessive OI type XIV |
title_full | Knocking out TMEM38B in human foetal osteoblasts hFOB 1.19 by CRISPR/Cas9: A model for recessive OI type XIV |
title_fullStr | Knocking out TMEM38B in human foetal osteoblasts hFOB 1.19 by CRISPR/Cas9: A model for recessive OI type XIV |
title_full_unstemmed | Knocking out TMEM38B in human foetal osteoblasts hFOB 1.19 by CRISPR/Cas9: A model for recessive OI type XIV |
title_short | Knocking out TMEM38B in human foetal osteoblasts hFOB 1.19 by CRISPR/Cas9: A model for recessive OI type XIV |
title_sort | knocking out tmem38b in human foetal osteoblasts hfob 1.19 by crispr/cas9: a model for recessive oi type xiv |
topic | Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8478202/ https://www.ncbi.nlm.nih.gov/pubmed/34582479 http://dx.doi.org/10.1371/journal.pone.0257254 |
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