DLST CRISPR Activation Plasmid (h): sc-409604-ACT

DLST (dihydrolipoamide S-succinyltransferase) encodes the E2 core component of the mitochondrial 2-oxoglutarate dehydrogenase complex, a key enzyme node in the tricarboxylic acid (TCA) cycle. By catalyzing succinyl-CoA formation from α-ketoglutarate and coordinating lipoate-dependent acyl transfer, DLST supports oxidative metabolism, NADH production, and mitochondrial redox balance. DLST function links central carbon metabolism to amino acid catabolism and broader mitochondrial homeostasis, processes frequently studied in contexts of metabolic stress and mitochondrial dysfunction. Altered regulation of TCA cycle components, including DLST, has been associated in the literature with bioenergetic remodeling observed in neurodegenerative and proliferative disease biology, supporting its relevance for mechanistic research.

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

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

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