FAT1 CRISPR Activation Plasmid (h): sc-410778-ACT

FAT1 encodes an atypical cadherin that contributes to cell–cell adhesion, epithelial polarity, and regulation of cytoskeletal organization during tissue morphogenesis. Through interactions that influence Hippo pathway signaling and planar cell polarity programs, FAT1 helps coordinate contact-dependent growth control, migration, and directional organization of cells. Dysregulated FAT1 expression or mutation has been associated with altered adhesion dynamics and aberrant signaling states implicated in cancer biology and developmental disorders. In human cell models, FAT1 is frequently studied for its roles in invasion-associated phenotypes, mechanotransduction, and transcriptional control downstream of junctional cues.

FAT1 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous FAT1 expression without altering the underlying DNA sequence.

FAT1 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the FAT1 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 FAT1 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous FAT1 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native FAT1 locus and enabling the study of FAT1-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of FAT1 pathway restoration in tumor cells with silenced or reduced FAT1 expression.

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