Ero1-Lα CRISPR Activation Plasmid (m): sc-424456-ACT

Mouse Ero1l encodes ER oxidoreductin 1 alpha (Ero1-Lα), an endoplasmic reticulum flavoprotein that re-oxidizes protein disulfide isomerases to support disulfide bond formation during oxidative protein folding. By coupling thiol oxidation to FAD-dependent electron transfer, Ero1-Lα contributes to ER redox homeostasis and intersects with the unfolded protein response, ER-associated degradation, and secretory pathway proteostasis. Altered Ero1-Lα activity can shift oxidative stress and calcium/ER stress signaling, linking Ero1l regulation to contexts such as hypoxia adaptation, inflammation, and tumor-associated secretory demands. These properties make Ero1l a useful target for studying redox-controlled maturation of secreted and membrane proteins and stress-responsive remodeling of ER function.

Ero1-Lα CRISPR Activation Plasmid (m) provides a targeted, non-destructive approach to upregulating endogenous Ero1l expression without altering the underlying DNA sequence.

Ero1-Lα CRISPR Activation Plasmid (m) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the Ero1l 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 Ero1l transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous Ero1-Lα expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native Ero1l locus and enabling the study of Ero1-Lα-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of Ero1-Lα pathway restoration in tumor cells with silenced or reduced Ero1l expression.

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