C3 CRISPR Activation Plasmid (h): sc-400622-ACT

Human complement component 3 (C3) is a central hub of the complement cascade, generated as a secreted zymogen that is proteolytically cleaved into C3a and C3b to amplify innate immune responses. C3b covalently opsonizes targets to promote phagocytosis and immune complex clearance, while C3a functions as an anaphylatoxin that modulates leukocyte recruitment and inflammatory signaling. Through integration of the classical, lectin, and alternative pathways, C3 regulates host defense, synapse pruning, and crosstalk with coagulation and cytokine networks. Dysregulated C3 activity or processing is implicated in complement-mediated inflammation and tissue injury, with relevance to autoimmune disease, infection biology, and neuroinflammatory and degenerative conditions.

C3 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous C3 expression without altering the underlying DNA sequence.

C3 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the C3 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 C3 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous C3 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native C3 locus and enabling the study of C3-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of C3 pathway restoration in tumor cells with silenced or reduced C3 expression.

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