FRYL CRISPR Activation Plasmid (h): sc-415428-ACT

Human FRYL encodes a large cytoplasmic protein implicated in regulation of cell morphology and cytoskeletal organization, with reported links to polarity and actin-dependent processes that shape migration and tissue architecture. FRYL has been associated with transcriptional control programs and signaling networks that coordinate growth and differentiation, suggesting roles in maintaining cellular homeostasis during development and stress responses. Altered FRYL expression or disruption of its regulatory context has been observed in genomic and transcriptomic studies of cancer and other complex disorders, motivating investigation of how FRYL influences proliferation, invasion, and lineage-specific gene expression. As a relatively understudied factor, FRYL provides a useful node for mapping gene regulatory interactions and cytoskeleton-coupled pathways in human cell models.

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

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

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