MATH-5 CRISPR Activation Plasmid (h): sc-415110-ACT

ATOH7 encodes the basic helix–loop–helix transcription factor MATH-5, a key regulator of retinal neurogenesis that promotes retinal ganglion cell competence and differentiation while influencing broader neuronal lineage decisions. MATH-5 integrates with developmental transcriptional networks involving Notch signaling, neurogenic bHLH factors, and downstream axon guidance and cell cycle exit programs to coordinate timing of neuronal specification. Dysregulated ATOH7 expression or function is linked to altered retinal development and has been associated with congenital optic nerve and retinal anomalies as well as susceptibility to neurodevelopmental visual disorders. As a node in early eye field patterning and neuronal fate determination, ATOH7 is widely used to interrogate gene regulatory circuits governing sensory neuron formation.

MATH-5 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous ATOH7 expression without altering the underlying DNA sequence.

MATH-5 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the ATOH7 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 ATOH7 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous MATH-5 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native ATOH7 locus and enabling the study of MATH-5-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of MATH-5 pathway restoration in tumor cells with silenced or reduced ATOH7 expression.

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