GCN1L1 CRISPR Activation Plasmid (h): sc-411545-ACT

Human GCN1 (GCN1L1) encodes a ribosome-associated cofactor that couples translational stress to the integrated stress response by facilitating activation of the eIF2α kinase GCN2 during amino acid limitation and related nutrient stresses. Through this role, GCN1L1 helps coordinate global translation attenuation with selective translation programs that support proteostasis and metabolic adaptation. Perturbation of GCN1L1-regulated stress signaling can influence ATF4-dependent transcriptional outputs, autophagy, and cellular survival decisions under proteotoxic or nutrient-deprived conditions. Dysregulation of these pathways has been implicated in contexts such as tumor cell stress tolerance, neurodegeneration-associated proteostasis defects, and metabolic disease mechanisms, making GCN1L1 a useful node for pathway-focused studies.

GCN1L1 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous GCN1 expression without altering the underlying DNA sequence.

GCN1L1 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the GCN1 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 GCN1 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous GCN1L1 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native GCN1 locus and enabling the study of GCN1L1-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of GCN1L1 pathway restoration in tumor cells with silenced or reduced GCN1 expression.

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