Amphiphysin I CRISPR Activation Plasmid (h): sc-403904-ACT

AMPH encodes Amphiphysin I, a BAR domain–containing endocytic adaptor that senses and induces membrane curvature and coordinates clathrin-mediated endocytosis. It interacts with dynamin and other SH3 domain partners to regulate synaptic vesicle recycling, membrane trafficking, and actin-linked remodeling at presynaptic terminals. Through its role in vesicle budding and recycling, Amphiphysin I contributes to neuronal signaling homeostasis and is commonly studied in pathways governing synaptic transmission and endosome dynamics. Altered AMPH expression or function has been investigated in the context of neurobiology and neurodegenerative disease mechanisms where defects in trafficking and synaptic maintenance are prominent.

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

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

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