MECL-1 CRISPR Activation Plasmid (h): sc-402101-ACT

PSMB10 encodes MECL-1, an inducible β subunit of the immunoproteasome that replaces constitutive proteasome components in response to inflammatory cues such as interferon-γ. MECL-1 contributes to ubiquitin-dependent protein turnover and shapes MHC class I antigen processing by influencing the generation of peptide epitopes presented to CD8+ T cells. Through its role in proteostasis and immune surveillance, altered immunoproteasome composition can affect inflammatory signaling, antigen presentation dynamics, and cellular stress responses. Dysregulation of immunoproteasome subunits, including MECL-1, has been associated with immune-mediated pathology and cancer-related immune evasion mechanisms in translational research contexts.

MECL-1 CRISPR Activation Plasmid (h) provides a targeted, non-destructive approach to upregulating endogenous PSMB10 expression without altering the underlying DNA sequence.

MECL-1 CRISPR Activation Plasmid (h) is a three-plasmid synergistic activation mediator (SAM) system engineered for highly efficient, site-specific transcriptional upregulation of the PSMB10 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 PSMB10 transcriptional start site, where VP64, p65, and HSF1 act in concert to recruit transcriptional machinery and drive upregulation of endogenous MECL-1 expression. Unlike nuclease-active Cas9, dCas9 does not introduce double-strand breaks or modify the genomic sequence, preserving the native PSMB10 locus and enabling the study of MECL-1-dependent transcriptional responses at the endogenous locus, making it a valuable tool for functional studies, target gene identification, and the modeling of MECL-1 pathway restoration in tumor cells with silenced or reduced PSMB10 expression.

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