GABARAP CRISPR Activation Plasmid (h): sc-416746-ACT

GABARAP (GABA type A receptor-associated protein) is a ubiquitin-like ATG8 family member that regulates autophagy by promoting autophagosome maturation, cargo recruitment, and fusion with lysosomes through interactions with LC3-interacting region (LIR)–containing proteins. It also contributes to intracellular membrane trafficking and receptor clustering, linking autophagic flux to vesicular transport and cytoskeletal organization. Dysregulation of GABARAP-associated pathways is relevant to proteostasis imbalance, neurodegenerative processes, and cancer cell stress adaptation, where altered autophagy can influence survival signaling and metabolic remodeling. As a compact scaffold protein with broad binding partners, GABARAP is frequently studied to dissect selective autophagy mechanisms and organelle quality control.

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

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

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