Ceramide Kinase CRISPR Activation Plasmid (h): sc-404164-ACT

Human CERK encodes ceramide kinase, a lipid kinase that phosphorylates ceramide to generate ceramide-1-phosphate (C1P), shifting sphingolipid balance from pro-apoptotic ceramide toward signaling-competent C1P pools. CERK activity supports membrane trafficking and inflammatory signaling, and it intersects with pathways regulating cell survival, proliferation, and stress responses through modulation of sphingolipid metabolism. By controlling the ceramide/C1P axis, CERK influences processes such as eicosanoid production, vesicular transport, and cytoskeletal dynamics. Dysregulated CERK-dependent sphingolipid signaling has been implicated in contexts including cancer biology, neuroinflammation, and metabolic dysfunction, making CERK a relevant node for mechanistic studies of lipid-mediated signaling.

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

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

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