EDD CRISPR Activation Plasmid (h): sc-402123-ACT

UBR5 encodes the human EDD protein, a HECT-type E3 ubiquitin ligase that regulates protein turnover and signaling through targeted ubiquitination. EDD participates in genome stability programs by coordinating DNA damage sensing and repair, modulating checkpoint control, and shaping transcriptional responses to cellular stress. Through these functions it influences cell-cycle progression, replication stress tolerance, and proteostasis pathways that intersect with ubiquitin–proteasome system dynamics. Dysregulated UBR5/EDD activity and copy-number changes have been associated with altered oncogenic signaling networks and aberrant DNA repair capacity, supporting its use as a mechanistic node in studies of tumor biology and stress-adaptive transcription.

EDD CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous UBR5 expression without altering the underlying DNA sequence.

EDD CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the UBR5 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 UBR5 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous EDD expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native UBR5 locus and enabling the study of EDD-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of EDD pathway restoration in tumor cells with silenced or reduced UBR5 expression.

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