DENND5B CRISPR Activation Plasmid (h): sc-414817-ACT

DENND5B encodes a DENN domain–containing protein implicated in regulation of Rab GTPase–dependent membrane trafficking, helping coordinate vesicle budding, endosomal transport, and compartmentalized signaling. By influencing small GTPase cycling and cargo sorting, DENND5B can affect receptor turnover, intracellular protein localization, and broader cellular processes such as polarity and proliferation. Altered expression or disruption of trafficking regulators like DENND5B has been associated with dysregulated signaling networks and genomic/functional changes observed across human disease contexts, including cancer-related phenotypes. As a result, DENND5B is frequently studied in pathways that link endomembrane dynamics to cell-state control and stress adaptation.

DENND5B CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous DENND5B expression without altering the underlying DNA sequence.

DENND5B CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the DENND5B 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 DENND5B transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous DENND5B expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native DENND5B locus and enabling the study of DENND5B-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of DENND5B pathway restoration in tumor cells with silenced or reduced DENND5B expression.

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