APPL1 CRISPR Activation Plasmid (h): sc-402243-ACT

APPL1 (adaptor protein, phosphotyrosine interacting with PH domain and leucine zipper 1) is a cytosolic endosomal adaptor that links membrane trafficking to signal transduction downstream of receptors such as adiponectin receptors and insulin/IGF pathways. By binding Rab5 and phosphoinositides and engaging AKT and AMPK signaling, APPL1 modulates endosomal dynamics, glucose homeostasis, and cellular responses to metabolic cues. APPL1-dependent signaling influences proliferation, survival, and oxidative stress responses through PI3K–AKT and related networks. Altered APPL1 expression or function has been associated with insulin resistance and metabolic syndrome phenotypes, and it is frequently studied in contexts connecting endocytosis with oncogenic and cardiometabolic signaling.

APPL1 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous APPL1 expression without altering the underlying DNA sequence.

APPL1 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the APPL1 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 APPL1 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous APPL1 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native APPL1 locus and enabling the study of APPL1-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of APPL1 pathway restoration in tumor cells with silenced or reduced APPL1 expression.

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