tPA CRISPR Activation Plasmid (h): sc-400922-ACT

Human PLAT encodes tissue-type plasminogen activator (tPA), a secreted serine protease that converts plasminogen to plasmin to regulate fibrinolysis and extracellular matrix proteolysis. Through plasmin generation, tPA influences pericellular remodeling, cell migration, and vascular homeostasis, interfacing with pathways that control coagulation balance and protease-activated signaling. PLAT expression is tightly controlled in endothelial and other cell types, and dysregulation of the plasminogen activation system has been associated with thrombotic and hemorrhagic phenotypes, neurovascular injury, and tumor microenvironment remodeling. These attributes make PLAT a useful target for studying protease networks, vascular biology, and context-dependent extracellular proteolysis.

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

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

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