Mrf-1 CRISPR Activation Plasmid (h2): sc-406689-ACT-2

Human ARID5A (Mrf-1) encodes an AT-rich interaction domain–containing RNA-binding regulator that modulates inflammatory gene expression by stabilizing selective cytokine mRNAs, including IL6, thereby shaping innate and adaptive immune responses. ARID5A integrates signals downstream of TLR/IL-1R and NF-κB pathways and counterbalances RNase-mediated decay programs to control the magnitude and duration of immune activation. Dysregulated ARID5A activity has been linked to aberrant cytokine production and immune-driven pathology in contexts such as autoimmunity, chronic inflammation, and tumor-associated inflammation. Gene editing of ARID5A supports mechanistic studies of post-transcriptional control, immune cell differentiation and activation, and pathway mapping of cytokine networks in human cellular models.

Mrf-1 CRISPR Activation Plasmid (h2) provides a targeted, non-destructive approach to upregulating endogenous ARID5A expression without altering the underlying DNA sequence.

Mrf-1 CRISPR Activation Plasmid (h2) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the ARID5A 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 ARID5A transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous Mrf-1 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native ARID5A locus and enabling the study of Mrf-1-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of Mrf-1 pathway restoration in tumor cells with silenced or reduced ARID5A expression.

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