CD44 CRISPR Activation Plasmid (h): sc-400209-ACT

Human CD44 encodes a multifunctional cell-surface glycoprotein that binds hyaluronan and other extracellular matrix ligands to regulate cell adhesion, migration, and lymphocyte homing. CD44 participates in cytoskeletal remodeling and signal transduction through pathways involving ERM proteins, Rho-family GTPases, and receptor tyrosine kinase crosstalk, shaping responses to inflammatory cues and tissue microenvironments. Alternative splicing and post-translational modifications diversify CD44 isoforms and tune ligand binding and signaling outputs. Dysregulated CD44 expression and isoform usage are frequently studied in contexts such as tumor invasion and metastasis, epithelial–mesenchymal transition, fibrosis, and immune cell trafficking.

CD44 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous CD44 expression without altering the underlying DNA sequence.

CD44 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the CD44 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 CD44 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous CD44 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native CD44 locus and enabling the study of CD44-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of CD44 pathway restoration in tumor cells with silenced or reduced CD44 expression.

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