LRRK2 CRISPR Activation Plasmid (h): sc-402602-ACT

Leucine-rich repeat kinase 2 (LRRK2) is a multidomain serine/threonine kinase and GTPase that integrates signaling at membranous compartments to coordinate vesicle trafficking, endolysosomal dynamics, cytoskeletal organization, and autophagy. LRRK2 phosphorylates multiple Rab GTPases and interfaces with microtubule- and actin-associated processes, linking kinase activity to organelle transport and proteostasis pathways. In human cells, dysregulated LRRK2 signaling has been extensively associated with neuronal vulnerability and immune-related stress responses, making it a key node for mechanistic studies of Parkinson’s disease–relevant biology. Experimental modulation of LRRK2 expression is used to interrogate pathway crosstalk involving lysosomal function, mitochondrial homeostasis, and inflammatory signaling cascades.

LRRK2 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous LRRK2 expression without altering the underlying DNA sequence.

LRRK2 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the LRRK2 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 LRRK2 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous LRRK2 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native LRRK2 locus and enabling the study of LRRK2-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of LRRK2 pathway restoration in tumor cells with silenced or reduced LRRK2 expression.

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