UNC45A CRISPR Activation Plasmid (m): sc-430401-ACT

Unc45a encodes UNC45A, a conserved myosin-directed chaperone that cooperates with HSP90 to promote proper folding, stability, and functional assembly of myosin motor proteins. Through regulation of actomyosin contractility and cytoskeletal organization, UNC45A contributes to processes such as intracellular transport, cell division, and cell migration. In mouse systems, perturbation of UNC45A activity can impact tissue morphogenesis and cellular mechanics by altering motor protein homeostasis and proteostasis pathways. Dysregulated UNC45A expression or function has been linked in the literature to cytoskeletal defects and aberrant proliferative or migratory phenotypes, supporting its relevance for mechanistic studies in disease-associated cell behavior.

UNC45A CRISPR Activation Plasmid (m) provides a targeted, non-destructive approach to upregulating endogenous Unc45a expression without altering the underlying DNA sequence.

UNC45A CRISPR Activation Plasmid (m) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the Unc45a 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 Unc45a transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous UNC45A expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native Unc45a locus and enabling the study of UNC45A-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of UNC45A pathway restoration in tumor cells with silenced or reduced Unc45a expression.

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