Crk II CRISPR Activation Plasmid (h): sc-402415-ACT

Human CRK encodes Crk II, an SH2/SH3 domain–containing adaptor protein that couples tyrosine-phosphorylated receptors and scaffold proteins to downstream signaling complexes. Crk II coordinates assembly of pathways controlling integrin signaling, focal adhesion dynamics, actin cytoskeletal remodeling, and cell migration through interactions with partners such as p130CAS/BCAR1, paxillin, and DOCK180/RAC1 modules. These networks integrate cues from receptor tyrosine kinases and adhesion receptors to regulate proliferation, survival, and directional motility. Dysregulated CRK/Crk II signaling has been implicated in aberrant invasion and metastasis biology and altered responses to growth factor and adhesion-dependent signaling in cancer and other proliferative disorders.

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

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

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