RNase L CRISPR Activation Plasmid (h): sc-403412-ACT

Human RNASEL encodes RNase L, an interferon-inducible endoribonuclease that is activated by 2′-5′ oligoadenylates generated by OAS proteins during innate immune sensing. Once engaged, RNase L cleaves cellular and viral RNA to limit replication, remodel the transcriptome, and amplify antiviral signaling through RNA decay products that stimulate pattern-recognition pathways. This activity intersects with interferon/JAK-STAT programs, stress granule biology, translational control, and apoptosis, linking RNase L to regulation of inflammation and cell fate decisions. Genetic and functional alterations in RNASEL have been investigated in the context of infection biology and immune-associated disease mechanisms, as well as tumor-associated phenotypes related to dysregulated RNA homeostasis.

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

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

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