Mel-CAM CRISPR Activation Plasmid (h): sc-401020-ACT

MCAM encodes Mel-CAM (CD146), a cell surface immunoglobulin superfamily adhesion molecule that regulates cell–cell interactions, endothelial junction dynamics, and cytoskeletal remodeling. In human tissues, Mel-CAM participates in processes linked to vascular biology, including endothelial activation, cell migration, and angiogenic signaling, and it can interface with pathways controlling adhesion-dependent signaling and permeability. Altered MCAM expression has been reported in multiple disease contexts, where it is frequently studied in relation to tumor cell invasiveness, metastatic behavior, and inflammatory remodeling of the vasculature. These features make MCAM a useful target for dissecting mechanisms of adhesion, motility, and microenvironmental crosstalk in relevant cellular models.

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

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

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