melanopsin CRISPR Activation Plasmid (h): sc-402783-ACT

Human OPN4 encodes melanopsin, a retinal opsin and G protein–coupled photopigment best known for mediating intrinsic photosensitivity in specialized retinal ganglion cells. Upon light activation, melanopsin couples primarily to Gq/11 signaling to stimulate PLCβ, elevate intracellular calcium, and modulate downstream pathways that influence neuronal excitability and transcriptional programs. This phototransduction axis contributes to non-image-forming visual functions, including circadian photoentrainment and pupillary light reflex regulation, linking OPN4 activity to sleep–wake timing and neuroendocrine rhythms. Altered melanopsin signaling has been studied in the context of circadian disruption and light-dependent physiological responses relevant to neurobehavioral and retinal research.

melanopsin CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous OPN4 expression without altering the underlying DNA sequence.

melanopsin CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the OPN4 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 OPN4 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous melanopsin expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native OPN4 locus and enabling the study of melanopsin-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of melanopsin pathway restoration in tumor cells with silenced or reduced OPN4 expression.

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