IRAK-4 CRISPR Activation Plasmid (h): sc-416405-ACT

Interleukin-1 receptor-associated kinase 4 (IRAK4) encodes IRAK-4, a serine/threonine kinase that functions as a proximal signaling node downstream of MyD88-dependent Toll-like receptors and IL-1 receptor family members. Upon receptor activation, IRAK-4 helps assemble the myddosome and propagates signaling to TRAF6 and TAK1, driving NF-κB and MAPK pathway activation and shaping cytokine and chemokine transcriptional programs. This pathway is central to innate immune sensing, inflammasome-associated responses, and crosstalk with interferon signaling networks. Dysregulated IRAK4 activity or expression has been linked to aberrant inflammatory signaling and immune-driven pathology, and altered pathway output is studied in infection biology, autoimmunity, and hematologic malignancy contexts.

IRAK-4 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous IRAK4 expression without altering the underlying DNA sequence.

IRAK-4 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the IRAK4 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 IRAK4 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous IRAK-4 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native IRAK4 locus and enabling the study of IRAK-4-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of IRAK-4 pathway restoration in tumor cells with silenced or reduced IRAK4 expression.

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