Enhancing Cardiomyocyte Transcription Using In Vivo CRISPR/Cas9 Systems

Area of Science:

• Molecular Biology
• Genetics
• Cardiovascular Research

Background:

• Programmable transcription factors allow gene-dose-dependent phenotyping within natural tissue environments.
• CRISPR/Cas9 systems have been modified into guide RNA (gRNA)-programmable transcriptional activation platforms (CRISPRa) for in vitro and in vivo applications.
• Targeted gene activation offers new avenues for studying cellular functions and disease mechanisms.

Purpose of the Study:

• To develop and validate a protocol for designing and implementing gRNA-based CRISPRa systems for endogenous gene activation.
• To establish a dCas9VPR-expressing mouse model under the Myosin heavy chain 6 (Myh6) promoter for cardiomyocyte-specific gene activation.
• To provide insights into the downstream applications of CRISPRa technology in cardiomyocyte biology.

Main Methods:

• Design and validation of novel gRNAs for specific gene targets.
• Development of CRISPRa tools for in vitro and in vivo applications.
• Generation of a transgenic mouse model expressing dCas9VPR in cardiomyocytes, driven by the Myh6 promoter.

Main Results:

• Efficient design and validation of gRNAs for enhanced transcriptional activity of selected genes.
• Successful implementation of CRISPRa in a dCas9VPR mouse model for cardiomyocyte studies.
• Demonstration of gene-dose-dependent phenotyping in vivo.

Conclusions:

• CRISPRa technology provides a powerful tool for endogenous gene activation with precise control.
• The developed protocol and mouse model are valuable for advancing research in cardiomyocyte biology and gene regulation.
• This approach facilitates in vivo, gene-dose-dependent phenotyping for a deeper understanding of cellular functions.