mAChR M5 CRISPR Activation Plasmid (h): sc-402943-ACT

CHRM5 encodes the human muscarinic acetylcholine receptor M5 (mAChR M5), a G protein–coupled receptor that preferentially couples to Gq/11 to stimulate phospholipase C signaling, inositol phosphate production, intracellular Ca2+ mobilization, and protein kinase C activation. Through these pathways, mAChR M5 modulates neuronal excitability and synaptic transmission, with additional roles in vascular tone and secretory processes depending on cell context. CHRM5-regulated signaling intersects with MAPK/ERK and other second-messenger networks that shape gene expression and adaptive cellular responses. Dysregulated cholinergic GPCR signaling has been linked to neuropsychiatric and neurodegenerative phenotypes as well as cardiometabolic and inflammatory processes, making CHRM5 a useful node for mechanistic studies of pathway coupling and receptor-driven transcriptional programs.

mAChR M5 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous CHRM5 expression without altering the underlying DNA sequence.

mAChR M5 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the CHRM5 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 CHRM5 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous mAChR M5 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native CHRM5 locus and enabling the study of mAChR M5-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of mAChR M5 pathway restoration in tumor cells with silenced or reduced CHRM5 expression.

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