RhoB CRISPR Activation Plasmid (h): sc-400875-ACT

Human RHOB encodes RhoB, a prenylated small GTPase of the Rho family that functions as a rapid-response regulator of actin cytoskeleton remodeling, endosomal trafficking, and membrane receptor turnover. RhoB integrates signaling from growth factor receptors and cellular stress pathways to modulate focal adhesion dynamics, cell shape, and motility through downstream effectors that interface with ROCK- and formin-associated programs. It also contributes to transcriptional and survival responses by influencing MAPK and PI3K/AKT pathway outputs, linking membrane-proximal signaling to nuclear decisions. Altered RHOB expression or regulation has been associated with cancer-related phenotypes including invasion, therapy stress responses, and changes in apoptotic susceptibility, making it relevant for mechanistic studies of tumor cell plasticity and microenvironmental adaptation.

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

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

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