rpn-10 Antibody

Definition and Biological Context

Rpn10 (Proteasome 26S Subunit, Non-ATPase 4) is a subunit of the 19S regulatory particle of the 26S proteasome. It functions as a ubiquitin receptor, recognizing polyubiquitinated substrates for proteasomal degradation via its ubiquitin-interacting motifs (UIMs) and von Willebrand factor A (VWA) domain . Antibodies against Rpn10 enable researchers to investigate its expression, localization, and interactions in diverse biological systems.

Mechanistic Insights into Proteasomal Function

• Ubiquitin Receptor Dynamics: Rpn10 antibodies have been used to demonstrate its equilibrium between proteasome-bound and free cytosolic pools, highlighting its dual role as a substrate receptor and adaptor .

• Structural Studies: Crystallography and immunoblotting revealed that Rpn10 monoubiquitination (at K84) alters its interaction with proteasomal subunits, promoting dissociation for substrate shuttling .

Disease Relevance

• Multiple Myeloma (MM):

  • Rpn10 is overexpressed in MM cells compared to normal plasma cells, correlating with poor prognosis .

  • Knockdown/knockout of Rpn10 via CRISPR/Cas9 reduced MM cell viability by 60–80%, induced apoptosis, and activated unfolded protein response pathways .

  • Proteomic profiling in Rpn10-deficient cells showed dysregulation of cell cycle (G2-M transition) and autophagy pathways .

• Therapeutic Targeting:

  • High-throughput screening identified SB699551, a small-molecule inhibitor of Rpn10, which reduced MM cell viability without affecting normal peripheral blood mononuclear cells .

Model Organism Studies

• C. elegans: Rpn10 knockdown transformed hermaphrodites into females, implicating it in sex determination pathways .

• S. cerevisiae: Human Rpn10 rescued proteasomal defects in rpn10Δ yeast, confirming functional conservation .

Technical Validation and Protocols

• Immunoblotting: Anti-Rpn10 antibodies detect a ~55 kDa band in humans (expected MW: 41 kDa) , with higher molecular weight bands indicating post-translational modifications like ubiquitination .

• Immunoprecipitation: Used to study Rpn10 interactions with substrates (e.g., FAT10) and shuttling factors (e.g., Rad23) .

• Knockdown Validation: siRNA targeting Rpn10 reduced its mRNA levels by >70% in HEK293 cells, confirmed via RT-PCR .

Clinical and Functional Implications

• Biomarker Potential: Elevated Rpn10 expression in MM and hepatocellular carcinoma suggests diagnostic utility .

• Ubiquitin-Independent Degradation: Rpn10’s VWA domain facilitates degradation of FAT10-tagged proteins, expanding its role beyond canonical ubiquitin signaling .

Limitations and Future Directions

• Redundancy: Rpn13 partially compensates for Rpn10 in ubiquitin recognition, complicating phenotype interpretation .

• Therapeutic Challenges: Targeting Rpn10 requires balancing proteasomal inhibition with off-target effects on autophagy and immune activation .