SPPL2b CRISPR Activation Plasmid (h): sc-405646-ACT

Human SPPL2B encodes the intramembrane aspartyl protease SPPL2b, a signal peptide peptidase–like enzyme that cleaves select type II membrane proteins within the lipid bilayer. By regulating substrate turnover and membrane protein fragment signaling, SPPL2b contributes to proteostasis, endosomal/lysosomal trafficking, and receptor-associated processing events that shape cell communication. SPPL2b activity intersects with pathways controlling protein quality control and immune-relevant membrane dynamics, making it pertinent to studies of inflammatory regulation and neurobiology where intramembrane proteolysis can modulate signaling outputs. Dysregulated membrane protein processing and trafficking are frequently implicated in complex disorders, positioning SPPL2B as a useful node for mechanistic interrogation in disease-relevant cell models.

SPPL2b CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous SPPL2B expression without altering the underlying DNA sequence.

SPPL2b CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the SPPL2B 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 SPPL2B transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous SPPL2b expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native SPPL2B locus and enabling the study of SPPL2b-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of SPPL2b pathway restoration in tumor cells with silenced or reduced SPPL2B expression.

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