SPRED2 CRISPR Activation Plasmid (m): sc-431133-ACT

Spred2 encodes SPRED2, a Sprouty-related adaptor protein that negatively regulates receptor tyrosine kinase signaling by constraining the RAS–RAF–MEK–ERK/MAPK cascade. Through interactions with RAF and associated signaling complexes, SPRED2 helps shape growth factor–driven outputs such as proliferation, differentiation, migration, and inflammatory gene expression. In mouse systems, altered SPRED2 activity has been linked to dysregulated MAPK signaling dynamics and downstream changes in immune and tissue homeostasis. As a pathway brake, SPRED2 is frequently studied in contexts where ERK signaling amplitude and duration influence phenotypes relevant to oncogenic transformation models, vascular biology, and inflammation-associated pathologies.

SPRED2 CRISPR Activation Plasmid (m) provides a targeted, non-destructive approach to upregulating endogenous Spred2 expression without altering the underlying DNA sequence.

SPRED2 CRISPR Activation Plasmid (m) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the Spred2 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 Spred2 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous SPRED2 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native Spred2 locus and enabling the study of SPRED2-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of SPRED2 pathway restoration in tumor cells with silenced or reduced Spred2 expression.

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