saposin CRISPR Activation Plasmid (h): sc-401394-ACT

PSAP encodes prosaposin, a lysosomal precursor that is proteolytically processed into saposins A–D, essential activator proteins for sphingolipid hydrolases. By facilitating glycosphingolipid and ceramide catabolism, saposins support lysosome function, membrane lipid turnover, and endolysosomal homeostasis, linking PSAP to autophagy-lysosome pathways and cellular stress responses. Disruption of prosaposin processing or saposin activity perturbs sphingolipid metabolism, leading to lipid storage phenotypes and neurodegeneration-associated cellular pathology. Consequently, PSAP is widely studied in models of lysosomal storage disorders, myelin maintenance, and inflammation driven by altered lipid signaling.

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

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

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