MYH4 CRISPR Activation Plasmid (h): sc-400467-ACT

MYH4 encodes myosin heavy chain 4, a fast-twitch skeletal muscle contractile motor that partners with actin to generate force through ATP-dependent cross-bridge cycling. As a core component of sarcomeric thick filaments, MYH4 contributes to myofibril assembly, fiber-type specification, and muscle performance programs regulated by calcium handling and metabolic signaling networks. Although MYH4 is best characterized in fast skeletal muscle physiology, altered myosin isoform expression and sarcomere remodeling are recurrent features of neuromuscular and myopathic disease states, supporting its use as a marker and mechanistic node in muscle biology studies. Modulating MYH4 expression provides a tractable route to interrogate pathways governing contractility, muscle adaptation, and stress responses in human cellular models.

MYH4 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous MYH4 expression without altering the underlying DNA sequence.

MYH4 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the MYH4 locus in human cell lines. The system is built around a catalytically inactive Cas9 (dCas9) carrying two inactivating mutations (D10A and N863A) that eliminate nuclease activity while preserving DNA binding. This dCas9 is fused to VP64, a potent transcriptional activator, and is co-expressed with a blasticidin resistance gene for selection. The second plasmid encodes the MS2-p65-HSF1 fusion protein, a secondary activator complex that works in concert with dCas9-VP64, alongside a hygromycin resistance gene. The third plasmid encodes a target-specific 20 nt sgRNA fused to two MS2 RNA aptamers that recruit the MS2-p65-HSF1 complex to the activation site, accompanied by a puromycin resistance gene. The three plasmids are delivered at a 1:1:1 mass ratio for balanced expression of all system components.

Once assembled at the target locus, the SAM complex binds within approximately 200 bp upstream of the MYH4 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous MYH4 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native MYH4 locus and enabling the study of MYH4-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of MYH4 pathway restoration in tumor cells with silenced or reduced MYH4 expression.

For Research Use Only. Not Intended for Diagnostic or Therapeutic Use.