Rab 8A CRISPR Activation Plasmid (h): sc-404285-ACT

RAB8A encodes the small GTPase Rab8A, a central regulator of post-Golgi membrane trafficking that coordinates polarized exocytosis, vesicle budding, and tethering during delivery of cargo to the plasma membrane and primary cilium. Rab8A functions in concert with GEFs and effectors to control recycling endosome dynamics, ciliogenesis, and cytoskeletal organization, thereby influencing cell polarity and directed migration. Through these roles, RAB8A participates in pathways linked to ciliopathy biology and developmental signaling, and its dysregulation has been associated with altered secretion programs and invasive cell behaviors in cancer-related contexts. Rab8A-dependent trafficking also interfaces with neuronal transport and autophagy-related membrane remodeling, making it relevant for mechanistic studies in diverse tissues.

Rab 8A CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous RAB8A expression without altering the underlying DNA sequence.

Rab 8A CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the RAB8A 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 RAB8A transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous Rab 8A expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native RAB8A locus and enabling the study of Rab 8A-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of Rab 8A pathway restoration in tumor cells with silenced or reduced RAB8A expression.

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