FAT10 CRISPR Activation Plasmid (m): sc-423986-ACT

Mouse Ubd encodes FAT10, a ubiquitin-like modifier that is conjugated to protein substrates through an E1/E2/E3 cascade and directs them toward proteasomal turnover in a manner that can be distinct from canonical ubiquitination. FAT10 expression is strongly induced by inflammatory cytokines and contributes to immune signaling, antigen processing, and cellular stress adaptation through regulation of protein homeostasis. By modulating degradation of key regulatory proteins, FAT10 influences cell cycle control, apoptosis, and innate immune responses. Dysregulated Ubd/FAT10 activity has been associated with inflammation-linked phenotypes and altered proteostasis, making it a useful node for studying immune-mediated tissue remodeling and disease-relevant signaling networks in mouse models.

FAT10 CRISPR Activation Plasmid (m) provides a targeted, non-destructive approach to upregulating endogenous Ubd expression without altering the underlying DNA sequence.

FAT10 CRISPR Activation Plasmid (m) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the Ubd 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 Ubd transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous FAT10 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native Ubd locus and enabling the study of FAT10-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of FAT10 pathway restoration in tumor cells with silenced or reduced Ubd expression.

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