WTX CRISPR Activation Plasmid (h2): sc-409039-ACT-2

Human AMER1 encodes the WTX protein, a tumor suppressor–associated factor that regulates canonical Wnt/β-catenin signaling by promoting β-catenin turnover and coordinating components of the destruction complex. WTX also participates in cytoskeletal organization and cell–cell adhesion processes, linking membrane-associated scaffolding to transcriptional control of developmental programs. Loss-of-function alterations in AMER1 are implicated in Wilms tumor and osteopathia striata with cranial sclerosis, and AMER1 disruption has been observed across additional cancer contexts where Wnt pathway dysregulation drives aberrant proliferation. AMER1/WTX gene editing and functional perturbation tools support mechanistic studies of Wnt-dependent transcription, developmental signaling, and genotype–phenotype relationships in renal and skeletal model systems.

WTX CRISPR Activation Plasmid (h2) provides a targeted, non-destructive approach to upregulating endogenous AMER1 expression without altering the underlying DNA sequence.

WTX CRISPR Activation Plasmid (h2) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the AMER1 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 AMER1 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous WTX expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native AMER1 locus and enabling the study of WTX-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of WTX pathway restoration in tumor cells with silenced or reduced AMER1 expression.

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