PMCA4 CRISPR Activation Plasmid (h): sc-402888-ACT

ATP2B4 encodes plasma membrane Ca2+-ATPase 4 (PMCA4), a high-affinity P-type ATPase that exports cytosolic Ca2+ to maintain calcium homeostasis and shape spatiotemporal Ca2+ signals. By tuning intracellular Ca2+ dynamics, PMCA4 influences excitation–contraction coupling, nitric oxide signaling via proximity to endothelial nitric oxide synthase, and downstream transcriptional programs controlled by Ca2+-responsive pathways such as calcineurin–NFAT and CaMK signaling. PMCA4 activity contributes to regulation of membrane potential, cell migration, and apoptotic susceptibility through control of Ca2+-dependent enzymes and ion channels. Dysregulated ATP2B4/PMCA4 expression or function has been linked in genetic and mechanistic studies to cardiovascular and erythroid phenotypes, supporting its relevance for modeling Ca2+-driven biology in human cells.

PMCA4 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous ATP2B4 expression without altering the underlying DNA sequence.

PMCA4 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the ATP2B4 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 ATP2B4 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous PMCA4 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native ATP2B4 locus and enabling the study of PMCA4-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of PMCA4 pathway restoration in tumor cells with silenced or reduced ATP2B4 expression.

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