Cbl CRISPR Activation Plasmid (h): sc-400561-ACT

Human CBL encodes Cbl, an E3 ubiquitin ligase and adaptor protein that downregulates activated receptor and non-receptor tyrosine kinases by promoting ubiquitination and endocytic trafficking. Through its TKB and RING finger domains, Cbl modulates signaling flux in EGFR, PDGFR, and immune receptor pathways, shaping MAPK/ERK, PI3K–AKT, and JAK/STAT outputs that control proliferation, differentiation, and cytokine responses. CBL integrates negative feedback in hematopoietic and immune cell signaling and contributes to regulation of cytoskeletal dynamics and cell migration. Dysregulation of CBL, including altered expression or functional variants, is associated with aberrant tyrosine kinase signaling in malignancy and immune-related disorders, supporting its use in mechanistic studies of signal attenuation.

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

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

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