p35 CRISPR Activation Plasmid (h): sc-400431-ACT

Human CDK5R1 encodes p35, a neuron-enriched regulatory subunit that binds and activates cyclin-dependent kinase 5 (CDK5), shaping kinase activity critical for neuronal development and synaptic function. The CDK5–p35 complex coordinates processes including neurite outgrowth, neuronal migration, cytoskeletal remodeling, and activity-dependent signaling through phosphorylation of substrates that influence microtubule dynamics and vesicle trafficking. Dysregulated CDK5R1/p35 homeostasis, including altered proteolysis and aberrant CDK5 signaling, has been associated with neurodegenerative and neurodevelopmental phenotypes and is frequently studied in models of neuronal stress and synaptic dysfunction. As a pathway node linking calcium-dependent signaling to cytoskeletal and transcriptional outputs, p35 provides a tractable handle for mechanistic studies in neuronal and glial systems.

p35 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous CDK5R1 expression without altering the underlying DNA sequence.

p35 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the CDK5R1 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 CDK5R1 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous p35 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native CDK5R1 locus and enabling the study of p35-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of p35 pathway restoration in tumor cells with silenced or reduced CDK5R1 expression.

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