title: The Proteasome Ubiquitin Receptor hRpn13 and Its Interacting Deubiquitinating Enzyme Uch37 Are Required for Proper Cell Cycle Progression.
abstract: Recently, we reported that bisbenzylidine piperidone RA190 adducts to Cys-88 of the proteasome ubiquitin receptor hRpn13, triggering accumulation of ubiquitinated proteins and endoplasmic reticulum stress-related apoptosis in various cancer cell lines. hRpn13 contains an N-terminal pleckstrin-like receptor for ubiquitin domain that binds ubiquitin and docks it into the proteasome as well as a C-terminal deubiquitinase adaptor (DEUBAD) domain that binds the deubiquitinating enzyme Uch37. Here we report that hRpn13 and Uch37 are required for proper cell cycle progression and that their protein knockdown leads to stalling at G0/G1 Moreover, serum-starved cells display reduced hRpn13 and Uch37 protein levels with hallmarks of G0/G1 stalling and recovery to their steady-state protein levels following release from nutrient deprivation. Interestingly, loss of hRpn13 correlates with a small but statistically significant reduction in Uch37 protein levels, suggesting that hRpn13 interaction may stabilize this deubiquitinating enzyme in human cells. We also find that RA190 treatment leads to a loss of S phase, suggesting a block of DNA replication, and G2 arrest by using fluorescence-activated cell sorting. Uch37 deprivation further indicated a reduction of DNA replication and G0/G1 stalling. Overall, this work implicates hRpn13 and Uch37 in cell cycle progression, providing a rationale for their function in cellular proliferation and for the apoptotic effect of the hRpn13-targeting molecule RA190.
authors.affiliation: From the Protein Processing Section, Structural Biophysics Laboratory, Center for Cancer Research, NCI, National Institutes of Health, Frederick, Maryland 21702 and.
authors.forename: Leah
authors.initials: L
authors.lastname: Randles
authors.affiliation: the Departments of Oncology.
authors.forename: Ravi K
authors.initials: RK
authors.lastname: Anchoori
authors.affiliation: the Departments of Oncology, Pathology, and Gynecology and Obstetrics, The Johns Hopkins University, Baltimore, Maryland 21231.
authors.forename: Richard B S
authors.initials: RB
authors.lastname: Roden
authors.affiliation: From the Protein Processing Section, Structural Biophysics Laboratory, Center for Cancer Research, NCI, National Institutes of Health, Frederick, Maryland 21702 and kylie.walters@nih.gov.
authors.forename: Kylie J
authors.initials: KJ
authors.lastname: Walters
mesh_terms: D002453:Cell Cycle / Q000187:drug effects / Q000502:physiology*; D004261:DNA Replication / Q000187:drug effects / Q000502:physiology*; D006367:HeLa Cells; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D008562:Membrane Glycoproteins / Q000235:genetics / Q000378:metabolism*; D043222:Ubiquitin Thiolesterase / Q000235:genetics / Q000378:metabolism*
chemical_list: C500781:ADRM1 protein, human; D047908:Intracellular Signaling Peptides and Proteins; D008562:Membrane Glycoproteins; C421245:UCHL5 protein, human; D043222:Ubiquitin Thiolesterase
keywords: RA190; Uch37; cancer biology; cell cycle progression; cell proliferation; hRpn13; p27Kip1; proteasome; ubiquitin; ubiquitylation (ubiquitination)