TIRAP CRISPR Activation Plasmid (h): sc-402650-ACT

Human TIRAP (also known as MAL) encodes an adaptor protein that couples Toll-like receptors, particularly TLR2 and TLR4, to downstream signaling cascades in innate immunity. By bridging receptor-proximal complexes to MyD88-dependent signaling, TIRAP promotes activation of NF-κB and MAPK pathways and shapes inflammatory cytokine and chemokine programs. Its regulation influences antimicrobial responses, immune cell activation, and crosstalk with interferon-associated networks. Dysregulated TIRAP signaling has been linked to inflammatory and infectious disease susceptibility and is frequently studied for its impact on tumor-associated inflammation and immune evasion mechanisms.

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

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

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