Optineurin CRISPR Activation Plasmid (h): sc-401851-ACT

OPTN encodes optineurin, a multifunctional adaptor protein that coordinates selective autophagy, vesicular trafficking, and innate immune signaling. Optineurin binds ubiquitinated cargo and autophagy machinery to support mitophagy and xenophagy, and it interfaces with NF-κB and TBK1-dependent pathways that regulate inflammatory responses. In the endocytic system, optineurin contributes to membrane dynamics and receptor trafficking through interactions with Rab GTPases and cytoskeletal components. Genetic perturbation or altered expression of OPTN has been linked to neurodegenerative and ocular phenotypes, including amyotrophic lateral sclerosis and glaucoma, making it a valuable target for mechanistic studies of proteostasis and inflammation.

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

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

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