eIF2Bε CRISPR Activation Plasmid (h): sc-403917-ACT

EIF2B5 encodes the ε subunit of eukaryotic initiation factor 2B (eIF2Bε), the catalytic guanine nucleotide exchange factor that regenerates active eIF2–GTP to sustain cap-dependent translation initiation. eIF2Bε is a central node in the integrated stress response, linking phosphorylation of eIF2α to global translational repression while enabling selective translation of stress-adaptive transcripts. Through this regulation of proteostasis and stress granule dynamics, EIF2B5 influences neuronal and glial homeostasis, myelination programs, and cellular resilience to metabolic and oxidative stress. Pathogenic disruption of eIF2B complex function is associated with vanishing white matter leukodystrophy and related eIF2B-spectrum disorders, making EIF2B5 a key target for mechanistic studies of stress-regulated translation.

eIF2Bε CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous EIF2B5 expression without altering the underlying DNA sequence.

eIF2Bε CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the EIF2B5 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 EIF2B5 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous eIF2Bε expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native EIF2B5 locus and enabling the study of eIF2Bε-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of eIF2Bε pathway restoration in tumor cells with silenced or reduced EIF2B5 expression.

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