title: COP9-associated CSN5 regulates exosomal protein deubiquitination and sorting.
abstract: Ubiquitinated endosomal proteins that are deposited into the lumens of multivesicular bodies are either sorted for lysosomal-mediated degradation or secreted as exosomes into the extracellular milieu. The mechanisms that underlie the sorting of cellular cargo proteins are currently unknown. In this study, we show that the COP9 signalosome (CSN)-associated protein CSN5 quantitatively regulated proteins that were sorted into exosomes. Western blot analysis of exosomal proteins indicated that small interfering (si)RNA knockdown of CSN5 results in increased levels of both ubiquitinated and non-ubiquitinated exosomal proteins, including heat shock protein 70, in comparison with exosomes isolated from the supernatants of 293 cells transfected with scrambled siRNA. Furthermore, 293 cells transfected with JAB1/MPN/Mov34 metalloenzyme domain-deleted CSN5 produced exosomes with higher levels of ubiquitinated heat shock protein 70, which did not affect non-ubiquitinated heat shock protein 70 levels. The loss of COP9-associated deubiquitin activity of CSN5 also led to the enhancement of HIV Gag that was sorted into exosomes as well as the promotion of HIV-1 release, suggesting that COP9-associated CSN5 regulates the sorting of a number of exosomal proteins in both a CSN5 JAB1/MPN/Mov34 metalloenzyme domain-dependent and -independent manner. We propose that COP9-associated CSN5 regulates exosomal protein sorting in both a deubiquitinating activity-dependent and -independent manner, which is contrary to the current idea of ubiquitin-dependent sorting of proteins to exosomes.
authors.affiliation: Division of Clinical Immunology and Rheumatology, University of Alabama at Birmingham, Birmingham, AL 35294, USA.
authors.forename: Yuelong
authors.initials: Y
authors.lastname: Liu
authors.forename: Spandan V
authors.initials: SV
authors.lastname: Shah
authors.forename: Xiaoyu
authors.initials: X
authors.lastname: Xiang
authors.forename: Jianhua
authors.initials: J
authors.lastname: Wang
authors.forename: Zhong-bin
authors.initials: ZB
authors.lastname: Deng
authors.forename: Cunren
authors.initials: C
authors.lastname: Liu
authors.forename: Liming
authors.initials: L
authors.lastname: Zhang
authors.forename: Jianming
authors.initials: J
authors.lastname: Wu
authors.forename: Tara
authors.initials: T
authors.lastname: Edmonds
authors.forename: Christina
authors.initials: C
authors.lastname: Jambor
authors.forename: John C
authors.initials: JC
authors.lastname: Kappes
authors.forename: Huang-Ge
authors.initials: HG
authors.lastname: Zhang
mesh_terms: D015153:Blotting, Western; D000075686:COP9 Signalosome Complex; D055354:Exosomes / Q000378:metabolism*; D016655:HIV Core Protein p24 / Q000378:metabolism; D015658:HIV Infections / Q000378:metabolism; D015497:HIV-1; D018840:HSP70 Heat-Shock Proteins / Q000378:metabolism; D006801:Humans; D047468:Immunoprecipitation; D047908:Intracellular Signaling Peptides and Proteins / Q000378:metabolism*; D046912:Multiprotein Complexes / Q000378:metabolism*; D010446:Peptide Fragments / Q000378:metabolism; D010447:Peptide Hydrolases / Q000378:metabolism*; D021381:Protein Transport / Q000502:physiology; D011506:Proteins / Q000378:metabolism*; D034741:RNA, Small Interfering; D014162:Transfection; D054875:Ubiquitination / Q000502:physiology*
chemical_list: D016655:HIV Core Protein p24; C489635:HIV Gag p24 protein (29-60); D018840:HSP70 Heat-Shock Proteins; D047908:Intracellular Signaling Peptides and Proteins; D046912:Multiprotein Complexes; D010446:Peptide Fragments; D011506:Proteins; D034741:RNA, Small Interfering; D010447:Peptide Hydrolases; C501488:COPS5 protein, human; D000075686:COP9 Signalosome Complex