Smurf1 CRISPR Activation Plasmid (h): sc-401196-ACT

SMURF1 encodes Smurf1, a HECT-domain E3 ubiquitin ligase that regulates protein turnover in key signaling networks, including TGF-β/BMP-SMAD pathways, WNT/planar cell polarity, and cytoskeletal remodeling via RhoA signaling. By catalyzing ubiquitination of pathway components, Smurf1 influences epithelial–mesenchymal transition, cell migration, and osteogenic differentiation. Altered SMURF1 activity has been linked to dysregulated growth factor signaling and aberrant tissue remodeling processes implicated in cancer biology, fibrosis, and skeletal homeostasis. These functions make SMURF1 a useful target for mechanistic studies of ubiquitin-mediated proteostasis and signaling crosstalk.

Smurf1 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous SMURF1 expression without altering the underlying DNA sequence.

Smurf1 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the SMURF1 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 SMURF1 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous Smurf1 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native SMURF1 locus and enabling the study of Smurf1-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of Smurf1 pathway restoration in tumor cells with silenced or reduced SMURF1 expression.

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