RAMP3 CRISPR Activation Plasmid (h): sc-402235-ACT

Receptor activity–modifying protein 3 (RAMP3) is a single-pass membrane accessory protein that heterodimerizes with select class B GPCRs, including calcitonin receptor-like receptor and the calcitonin receptor, to dictate ligand specificity, receptor trafficking, and downstream signaling bias. By shaping adrenomedullin/CGRP-family receptor complexes, RAMP3 influences cAMP-dependent pathways and broader processes such as endothelial signaling, barrier function, and cellular homeostasis through modulation of receptor internalization and recycling. Altered RAMP3 expression has been associated with dysregulated GPCR signaling programs relevant to cardiovascular physiology and tumor-associated signaling networks, supporting its use as a node for mechanistic studies of receptor pharmacology and cell-state regulation. These properties make human RAMP3 a useful target for dissecting receptor complex assembly and signal transduction dynamics in physiologically relevant cell models.

RAMP3 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous RAMP3 expression without altering the underlying DNA sequence.

RAMP3 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the RAMP3 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 RAMP3 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous RAMP3 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native RAMP3 locus and enabling the study of RAMP3-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of RAMP3 pathway restoration in tumor cells with silenced or reduced RAMP3 expression.

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