TOM40 CRISPR Activation Plasmid (h): sc-400967-ACT

Human TOMM40 encodes TOM40, the central β-barrel pore of the translocase of the outer mitochondrial membrane (TOM) complex that mediates import of nuclear-encoded proteins into mitochondria. By enabling precursor recognition and translocation across the outer membrane, TOM40 supports mitochondrial proteostasis, oxidative phosphorylation capacity, and coordination of mitochondrial biogenesis with cellular metabolism. TOMM40 function intersects with pathways governing mitochondrial dynamics, stress signaling, and apoptosis through its role in maintaining efficient protein trafficking to the organelle. Genetic variation and dysregulated mitochondrial import have been linked to neurodegeneration and other disorders characterized by mitochondrial dysfunction, making TOMM40 a key node for mechanistic studies of mitochondrial homeostasis.

TOM40 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous TOMM40 expression without altering the underlying DNA sequence.

TOM40 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the TOMM40 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 TOMM40 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous TOM40 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native TOMM40 locus and enabling the study of TOM40-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of TOM40 pathway restoration in tumor cells with silenced or reduced TOMM40 expression.

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