Endothelin-3/EDN3/ET-3 CRISPR Activation Plasmid (h): sc-404436-ACT

EDN3 encodes endothelin-3 (ET-3), a secreted vasoactive peptide of the endothelin family that signals primarily through endothelin receptor type B (EDNRB) to regulate neural crest–derived cell migration, melanocyte lineage specification, and enteric nervous system development. ET-3 contributes to paracrine control of smooth muscle tone and vascular homeostasis via GPCR-mediated pathways that engage phospholipase C, intracellular calcium flux, and MAPK signaling. In development and tissue patterning contexts, EDN3–EDNRB signaling intersects with pathways controlling proliferation and differentiation, including crosstalk with WNT and KIT-dependent programs in pigment cell biology. Dysregulation of EDN3 signaling is linked to disorders of neural crest development and pigmentation, and it is frequently studied for its roles in gastrointestinal innervation and microenvironmental signaling dynamics.

Endothelin-3/EDN3/ET-3 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous EDN3 expression without altering the underlying DNA sequence.

Endothelin-3/EDN3/ET-3 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the EDN3 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 EDN3 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous Endothelin-3/EDN3/ET-3 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native EDN3 locus and enabling the study of Endothelin-3/EDN3/ET-3-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of Endothelin-3/EDN3/ET-3 pathway restoration in tumor cells with silenced or reduced EDN3 expression.

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