SIP1/ZEB2 CRISPR Activation Plasmid (h): sc-400655-ACT

ZEB2, also known as SIP1, is a zinc finger E-box–binding transcription factor that regulates epithelial–mesenchymal transition (EMT) by repressing epithelial gene programs, including CDH1, and coordinating cell fate and differentiation. Through its interactions with SMAD proteins, SIP1/ZEB2 integrates TGF-β/BMP signaling with chromatin remodeling to control lineage specification, migration, and developmental patterning. ZEB2 activity shapes neural crest and hematopoietic development and modulates immune cell differentiation, linking it to pathways governing tissue morphogenesis and inflammatory responses. Dysregulation of ZEB2-associated transcriptional networks has been implicated in developmental disorders and in cellular plasticity programs relevant to tumor progression and metastasis-associated phenotypes.

SIP1/ZEB2 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous ZEB2 expression without altering the underlying DNA sequence.

SIP1/ZEB2 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the ZEB2 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 ZEB2 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous SIP1/ZEB2 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native ZEB2 locus and enabling the study of SIP1/ZEB2-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of SIP1/ZEB2 pathway restoration in tumor cells with silenced or reduced ZEB2 expression.

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