HAI-1 CRISPR Activation Plasmid (h): sc-404011-ACT

SPINT1 encodes hepatocyte growth factor activator inhibitor-1 (HAI-1), a membrane-associated Kunitz-type serine protease inhibitor that constrains pericellular proteolysis by inhibiting proteases such as HGFA and matriptase (ST14). By regulating activation of HGF and downstream signaling through the MET pathway, HAI-1 influences epithelial homeostasis, cell–cell junction integrity, and extracellular matrix remodeling. SPINT1–protease networks intersect with processes including epithelial differentiation, wound repair, and inflammatory protease cascades. Dysregulated HAI-1 expression or imbalance with cognate proteases has been linked to altered epithelial barrier function and protease-driven phenotypes reported across multiple cancer and fibrotic disease research contexts.

HAI-1 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous SPINT1 expression without altering the underlying DNA sequence.

HAI-1 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the SPINT1 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 SPINT1 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous HAI-1 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native SPINT1 locus and enabling the study of HAI-1-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of HAI-1 pathway restoration in tumor cells with silenced or reduced SPINT1 expression.

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