Lyst CRISPR Activation Plasmid (h): sc-410730-ACT

Human LYST (lysosomal trafficking regulator) encodes Lyst, a large cytosolic protein that controls lysosome-related organelle size, positioning, and membrane dynamics required for late endosome–lysosome fusion and cargo trafficking. Through regulation of vesicle fission and fusion events, Lyst influences degradative lysosomal function, antigen processing, and secretion from specialized granules in immune and pigment cells. Dysregulated LYST activity is linked to defects in lysosomal homeostasis and abnormal giant granule formation that compromise innate and adaptive immune cell function, making it relevant to studies of immunodeficiency and pigmentation biology. LYST is therefore frequently examined in pathways governing intracellular trafficking, autophagy-lysosome crosstalk, and stress responses to impaired organelle turnover.

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

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

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