RIP4 CRISPR Activation Plasmid (h): sc-404910-ACT

Human RIPK4 encodes receptor-interacting serine/threonine-protein kinase 4 (RIP4), a signaling kinase that functions downstream of protein kinase C to regulate keratinocyte differentiation, epidermal morphogenesis, and epithelial barrier formation. RIP4 participates in pathways controlling NF-κB and MAPK signaling and interfaces with cytoskeletal and adhesion programs that shape epithelial architecture. Dysregulated RIPK4 activity has been linked to altered epithelial homeostasis, inflammation-associated signaling, and tumor biology in squamous epithelia, making it relevant for studies of differentiation states and epithelial stress responses. Its pathway connectivity also supports mechanistic interrogation of transcriptional programs that govern stratified epithelium development and disease-associated remodeling.

RIP4 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous RIPK4 expression without altering the underlying DNA sequence.

RIP4 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the RIPK4 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 RIPK4 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous RIP4 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native RIPK4 locus and enabling the study of RIP4-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of RIP4 pathway restoration in tumor cells with silenced or reduced RIPK4 expression.

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