EAAT1 CRISPR Activation Plasmid (h): sc-400917-ACT

SLC1A3 encodes excitatory amino acid transporter 1 (EAAT1), a high-affinity, sodium-dependent glutamate and aspartate transporter that maintains extracellular glutamate homeostasis and limits excitotoxic signaling. EAAT1 is prominently expressed in astrocytes and contributes to the glutamate–glutamine cycle, coupling neurotransmitter clearance to cellular energy metabolism and ion gradients. By shaping synaptic transmission and protecting neuronal circuits from excessive glutamatergic activity, SLC1A3 influences neurodevelopmental and neurodegenerative processes. Altered EAAT1 expression or function has been associated with disturbed glutamate signaling in neurological and psychiatric disease contexts, supporting its use in mechanistic studies of excitatory neurotransmission.

EAAT1 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous SLC1A3 expression without altering the underlying DNA sequence.

EAAT1 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the SLC1A3 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 SLC1A3 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous EAAT1 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native SLC1A3 locus and enabling the study of EAAT1-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of EAAT1 pathway restoration in tumor cells with silenced or reduced SLC1A3 expression.

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