TRPV5 CRISPR Activation Plasmid (h): sc-403668-ACT

TRPV5 encodes a highly calcium-selective, constitutively active TRP cation channel predominantly localized to the apical membrane of epithelial cells, where it mediates transcellular Ca2+ entry. Channel activity is coordinated with calcium homeostasis pathways, including calbindin buffering and basolateral extrusion by Na+/Ca2+ exchange and Ca2+-ATPases, and is modulated by factors such as intracellular Ca2+, pH, and calciotropic signaling. TRPV5 function is tightly linked to epithelial ion transport programs and calcium reabsorption physiology, making it a relevant node for studying mineral metabolism and epithelial barrier transport. Dysregulated TRPV5 expression or activity has been associated with altered calcium handling phenotypes and has been investigated in contexts of renal and systemic calcium balance disorders.

TRPV5 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous TRPV5 expression without altering the underlying DNA sequence.

TRPV5 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the TRPV5 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 TRPV5 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous TRPV5 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native TRPV5 locus and enabling the study of TRPV5-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of TRPV5 pathway restoration in tumor cells with silenced or reduced TRPV5 expression.

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