Ribophorin I CRISPR Activation Plasmid (h): sc-402286-ACT

RPN1 encodes ribophorin I, an essential component of the oligosaccharyltransferase (OST) complex embedded in the rough endoplasmic reticulum membrane that supports co-translational N-linked glycosylation of nascent secretory and membrane proteins. By coordinating glycan transfer and protein maturation, ribophorin I contributes to ER proteostasis, quality control, and trafficking across the secretory pathway, with downstream impacts on receptor signaling and cell-surface proteome composition. Altered ER glycosylation capacity and OST subunit dysregulation have been linked to cellular stress responses and aberrant protein processing observed in multiple disease-relevant contexts, making RPN1 a useful node for studying glycoprotein biogenesis. Experimental modulation of RPN1 expression helps interrogate how ER-resident glycosylation machinery shapes folding efficiency, degradation via ER-associated degradation (ERAD), and broader proteome stability.

Ribophorin I CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous RPN1 expression without altering the underlying DNA sequence.

Ribophorin I CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the RPN1 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 RPN1 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous Ribophorin I expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native RPN1 locus and enabling the study of Ribophorin I-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of Ribophorin I pathway restoration in tumor cells with silenced or reduced RPN1 expression.

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