VPS41 CRISPR Activation Plasmid (h): sc-403666-ACT

VPS41 encodes a core subunit of the HOPS tethering complex that coordinates endosome–lysosome fusion and late endosomal maturation through interactions with Rab GTPases and SNARE machinery. By supporting delivery of cargo to lysosomes, VPS41 contributes to autophagic flux, receptor downregulation, and turnover of damaged organelles, integrating with vesicle trafficking networks that maintain cellular proteostasis. Disruption of HOPS-dependent trafficking is linked to lysosomal dysfunction, impaired autophagy, and altered signaling outputs, processes frequently implicated in neurodegeneration and cancer cell biology. Human VPS41 is therefore widely studied in pathways governing endolysosomal homeostasis, stress responses, and intracellular transport.

VPS41 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous VPS41 expression without altering the underlying DNA sequence.

VPS41 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the VPS41 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 VPS41 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous VPS41 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native VPS41 locus and enabling the study of VPS41-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of VPS41 pathway restoration in tumor cells with silenced or reduced VPS41 expression.

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