RBFOX3/NeuN CRISPR Activation Plasmid (h): sc-402976-ACT

RBFOX3, also known as NeuN, encodes a neuron-enriched RNA-binding protein that recognizes UGCAUG motifs to regulate alternative splicing programs essential for neuronal differentiation, synaptic maturation, and maintenance of neuronal identity. Through control of exon inclusion and mRNA isoform balance, RBFOX3 integrates into post-transcriptional networks that shape cytoskeletal dynamics, neurite outgrowth, and activity-dependent gene expression. RBFOX family–directed splicing is linked to pathways governing excitatory/inhibitory balance and neuronal connectivity, making RBFOX3 a widely used marker and regulator in studies of neural development and function. Dysregulated neuronal splicing programs involving RBFOX factors have been associated with neurodevelopmental and neurodegenerative disease mechanisms, supporting its relevance for modeling RNA processing defects in human systems.

RBFOX3/NeuN CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous RBFOX3 expression without altering the underlying DNA sequence.

RBFOX3/NeuN CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the RBFOX3 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 RBFOX3 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous RBFOX3/NeuN expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native RBFOX3 locus and enabling the study of RBFOX3/NeuN-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of RBFOX3/NeuN pathway restoration in tumor cells with silenced or reduced RBFOX3 expression.

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