GRP 75 CRISPR Activation Plasmid (h): sc-400471-ACT

HSPA9 encodes GRP75 (mortalin), a mitochondria-associated HSP70 family chaperone that supports protein import, folding, and proteostasis within the mitochondrial matrix. GRP75 contributes to mitochondrial bioenergetics, iron–sulfur cluster assembly, and stress-adaptive signaling, and it also participates in mitochondria–ER communication that influences calcium handling and redox homeostasis. Through these functions, HSPA9 is linked to pathways governing mitochondrial quality control, apoptotic sensitivity, and cellular responses to oxidative and proteotoxic stress. Dysregulated GRP75 activity and expression have been associated with altered mitochondrial function and phenotypes relevant to neurodegeneration, hematopoietic dysfunction, and cancer biology in experimental systems.

GRP 75 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous HSPA9 expression without altering the underlying DNA sequence.

GRP 75 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the HSPA9 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 HSPA9 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous GRP 75 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native HSPA9 locus and enabling the study of GRP 75-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of GRP 75 pathway restoration in tumor cells with silenced or reduced HSPA9 expression.

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