Rent1 CRISPR Activation Plasmid (h): sc-402462-ACT

Human UPF1 (Rent1) is an ATP-dependent RNA helicase and central effector of nonsense-mediated mRNA decay (NMD), a conserved pathway that detects and degrades transcripts containing premature termination codons. Through interactions with SMG proteins and the exon junction complex, UPF1 couples translation termination surveillance to mRNP remodeling, RNA quality control, and regulation of normal transcript isoforms. Beyond NMD, UPF1 contributes to telomere maintenance, genome stability, and stress-responsive RNA metabolism, linking RNA surveillance to broader cellular homeostasis. Dysregulated UPF1 activity has been associated with altered transcriptome integrity and aberrant proteome output in contexts including tumorigenesis and neurodevelopmental phenotypes, supporting its utility for mechanistic studies of RNA regulation.

Rent1 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous UPF1 expression without altering the underlying DNA sequence.

Rent1 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the UPF1 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 UPF1 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous Rent1 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native UPF1 locus and enabling the study of Rent1-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of Rent1 pathway restoration in tumor cells with silenced or reduced UPF1 expression.

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