FUS/TLS CRISPR Activation Plasmid (h): sc-400612-ACT

FUS (FUS/TLS) encodes an RNA/DNA-binding protein that coordinates multiple steps of gene expression, including transcriptional regulation, pre-mRNA splicing, mRNA transport, and microRNA biogenesis. FUS participates in the DNA damage response and genome maintenance through interactions with repair factors and coupling of transcription to RNA processing, and it dynamically redistributes to stress granules under cellular stress. Through these roles, FUS influences neuronal homeostasis and proteostasis pathways where precise control of RNA metabolism is critical. Dysregulation, aggregation, or mutation of FUS is strongly linked to neurodegenerative disease biology, including amyotrophic lateral sclerosis and frontotemporal lobar degeneration, making it a widely used model for studying RNA-binding proteinopathies.

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

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

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