Sec5 CRISPR Activation Plasmid (h): sc-405088-ACT

Human EXOC2 encodes Sec5, an essential subunit of the octameric exocyst complex that tethers secretory vesicles to the plasma membrane to support polarized exocytosis. Sec5 coordinates vesicle targeting with actin remodeling and small GTPase signaling, contributing to membrane expansion, cell migration, cytokinesis, and regulated delivery of receptors and transporters. Through its role in exocyst-dependent trafficking, EXOC2 influences pathways governing epithelial polarity and neuronal outgrowth, and altered exocyst function has been associated with dysregulated proliferation and invasive phenotypes in multiple disease contexts. Sec5-linked trafficking defects are therefore relevant for mechanistic studies of membrane dynamics, signaling compartmentalization, and cell-state transitions.

Sec5 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous EXOC2 expression without altering the underlying DNA sequence.

Sec5 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the EXOC2 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 EXOC2 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous Sec5 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native EXOC2 locus and enabling the study of Sec5-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of Sec5 pathway restoration in tumor cells with silenced or reduced EXOC2 expression.

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