14-3-3 ζ CRISPR Activation Plasmid (h): sc-400490-ACT

YWHAZ encodes the human 14-3-3 zeta adaptor protein, a phosphoserine/phosphothreonine-binding regulator that stabilizes client proteins and coordinates signal transduction across multiple compartments. 14-3-3 zeta participates in PI3K–AKT, MAPK, and cell-cycle checkpoint control by modulating the localization, phosphorylation state, and activity of diverse kinases, transcription factors, and apoptotic regulators. Through these interactions it influences proliferation, survival, stress responses, cytoskeletal dynamics, and metabolic signaling. Dysregulated YWHAZ/14-3-3 zeta function has been associated with altered oncogenic signaling, therapy resistance phenotypes, and neurobiological processes relevant to synaptic function and proteostasis.

14-3-3 ζ CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous YWHAZ expression without altering the underlying DNA sequence.

14-3-3 ζ CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the YWHAZ 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 YWHAZ transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous 14-3-3 ζ expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native YWHAZ locus and enabling the study of 14-3-3 ζ-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of 14-3-3 ζ pathway restoration in tumor cells with silenced or reduced YWHAZ expression.

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