Generating endogenous Myh11-driven Cre mice for sex-independent gene deletion in smooth muscle cells
In: JCI Insight, Jg. 8 (2023-07-01), Heft 14
Online
academicJournal
Zugriff:
Specific and efficient smooth muscle cell–targeted (SMC-targeted) gene deletion is typically achieved by pairing SMMHC-CreERT2-Tg mice with mice carrying the loxP-flanked gene. However, the transgene, CreERT2, is not controlled by the endogenous Myh11 gene promoter, and the codon-modified iCreERT2 exhibits significant tamoxifen-independent leakage. Furthermore, because the Cre-bearing bacterial artificial chromosome (BAC) is inserted onto the Y chromosome, the SMMHC-CreERT2-Tg mice strain can only exhibit gene deletions in male mice. Additionally, there is a lack of Myh11-driven constitutive Cre mice when tamoxifen usage is a concern. We used CRISPR/Cas9-mediated homologous recombination between a donor vector carrying the CreNLSP2A or CreERT2–P2A sequence and homologous arm surrounding the translation start site of the Myh11 gene to generate Cre-knockin mice. The P2A sequence enables the simultaneous translation of Cre and endogenous proteins. Using reporter mice, we assessed Cre-mediated recombination efficiency, specificity, tamoxifen-dependent controllability, and functionality in both sexes. Both constitutive (Myh11-CreNLSP2A) and inducible (Myh11-CreERT2–P2A) Cre mice demonstrated efficient, SMC-specific, sex-independent Cre recombinase activity without confounding endogenous gene expression. Combined with recently generated BAC transgenic Myh11-CreERT2-RAD mice and the Itga8-CreERT2 mouse models, our models will help expand the research toolbox, facilitating unbiased and comprehensive research in SMCs and SMC-dependent cardiovascular diseases.
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Generating endogenous Myh11-driven Cre mice for sex-independent gene deletion in smooth muscle cells
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Autor/in / Beteiligte Person: | Zhao, Yang ; Zhao, Guizhen ; Chang, Ziyi ; Zhu, Tianqing ; Zhao, Ying ; Lu, Haocheng ; Xue, Chao ; Saunders, Thomas L. ; Guo, Yanhong ; Chang, Lin ; Y. Eugene Chen ; Zhang, Jifeng |
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Zeitschrift: | JCI Insight, Jg. 8 (2023-07-01), Heft 14 |
Veröffentlichung: | American Society for Clinical investigation, 2023 |
Medientyp: | academicJournal |
ISSN: | 2379-3708 (print) |
DOI: | 10.1172/jci.insight.171661 |
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