HSPA5/BiP/GRP78 CRISPR Activation Plasmid (h): sc-400073-ACT

HSPA5 encodes the endoplasmic reticulum (ER) chaperone BiP/GRP78, a central regulator of protein folding, ER quality control, and proteostasis. HSPA5 binds nascent polypeptides and misfolded proteins, coordinates ER-associated degradation (ERAD), and modulates unfolded protein response (UPR) signaling through key sensors including PERK, IRE1, and ATF6. By integrating ER stress with calcium homeostasis, redox balance, and secretory pathway throughput, HSPA5 influences cell survival and metabolic adaptation. Dysregulated HSPA5 expression and UPR activation are implicated in cancer stress tolerance, neurodegenerative proteinopathies, diabetes-related ER stress, and inflammatory states.

HSPA5/BiP/GRP78 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous HSPA5 expression without altering the underlying DNA sequence.

HSPA5/BiP/GRP78 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the HSPA5 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 HSPA5 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous HSPA5/BiP/GRP78 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native HSPA5 locus and enabling the study of HSPA5/BiP/GRP78-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of HSPA5/BiP/GRP78 pathway restoration in tumor cells with silenced or reduced HSPA5 expression.

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