MARK4 CRISPR Activation Plasmid (h): sc-404386-ACT

MARK4 (microtubule affinity-regulating kinase 4) is a serine/threonine kinase of the AMPK-related MARK family that modulates microtubule stability by phosphorylating microtubule-associated proteins such as TAU/MAPT. Through regulation of cytoskeletal dynamics, MARK4 contributes to cell polarity, intracellular transport, and mitotic processes, and it interfaces with signaling networks that coordinate stress and metabolic cues with microtubule remodeling. Dysregulated MARK4 activity has been linked to altered neuronal microtubule organization and has been investigated in contexts of neurodegeneration, as well as cancer-associated phenotypes including changes in proliferation and migration. These features make MARK4 a useful target for mechanistic studies of microtubule-dependent pathways and kinase-regulated cellular organization.

MARK4 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous MARK4 expression without altering the underlying DNA sequence.

MARK4 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the MARK4 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 MARK4 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous MARK4 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native MARK4 locus and enabling the study of MARK4-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of MARK4 pathway restoration in tumor cells with silenced or reduced MARK4 expression.

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