CYP3A5 CRISPR Activation Plasmid (h): sc-401306-ACT

CYP3A5 encodes a cytochrome P450 monooxygenase that contributes to oxidative metabolism of endogenous steroids and xenobiotics, shaping cellular exposure to bioactive compounds and metabolites. As part of the CYP3A subfamily, CYP3A5 participates in NADPH-dependent electron transfer via cytochrome P450 reductase and impacts lipid homeostasis, hormone signaling, and cellular stress responses through metabolite-driven regulation. Expression is tissue- and genotype-dependent, with notable activity in liver and kidney influencing interindividual variability in metabolic capacity. Dysregulated CYP3A5 expression or function has been associated with altered pharmacokinetics and susceptibility to metabolic and renal pathophysiology, supporting its use in mechanistic studies of detoxification pathways.

CYP3A5 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous CYP3A5 expression without altering the underlying DNA sequence.

CYP3A5 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the CYP3A5 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 CYP3A5 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous CYP3A5 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native CYP3A5 locus and enabling the study of CYP3A5-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of CYP3A5 pathway restoration in tumor cells with silenced or reduced CYP3A5 expression.

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