title: TRIM32-Cytoplasmic-Body Formation Is an ATP-Consuming Process Stimulated by HSP70 in Cells.
abstract: The spontaneous and energy-releasing reaction of protein aggregation is typically prevented by cellular quality control machinery (QC). TRIM32 is a member of the TRIM (tripartite motif-containing) ubiquitin E3 ligases, and when overexpressed in cultured cells, readily forms spherical inclusions designated as cytoplasmic bodies (CBs) even without proteasome inhibition. Here, we show that HSP70, a central QC component, is a primary binding factor of overexpressed TRIM32. Contrary to expectation, however, we find that this molecular chaperone facilitates and stabilizes CB assembly depending on intrinsic ATPase activity, rather than preventing CB formation. We also show that the HSP70-TRIM32 complex is biochemically distinct from the previously characterized 14-3-3-TRIM32 phospho-complex. Moreover, the two complexes have opposing roles, with HSP70 stimulating CB formation and 14-3-3 retaining TRIM32 in a diffuse form throughout the cytosol. Our results suggest that CB inclusion formation is actively controlled by cellular QC and requires ATP, similar to protein folding and degradation reactions.
authors.affiliation: Department of Applied Chemistry, National Defense Academy, Yokosuka, Kanagawa, Japan.
authors.forename: Yuki
authors.initials: Y
authors.lastname: Kawaguchi
authors.affiliation: Department of Chemistry, Tokyo Metropolitan University, Hachioji, Tokyo, Japan.
authors.forename: Masato
authors.initials: M
authors.lastname: Taoka
authors.affiliation: Department of Applied Chemistry, National Defense Academy, Yokosuka, Kanagawa, Japan.
authors.forename: Takahiro
authors.initials: T
authors.lastname: Takekiyo
authors.affiliation: Department of Applied Chemistry, National Defense Academy, Yokosuka, Kanagawa, Japan.
authors.forename: Takamasa
authors.initials: T
authors.lastname: Uekita
authors.affiliation: Division of Infectious Disease Control, Center for Infectious Diseases, Kobe University Graduate School of Medicine, Kobe, Japan.
authors.forename: Ikuo
authors.initials: I
authors.lastname: Shoji
authors.affiliation: Biotechnology Group, R&D Division, Tokyo Metropolitan Industrial Technology Research Institute, Koto-ku, Tokyo, Japan.
authors.forename: Naomi
authors.initials: N
authors.lastname: Hachiya
authors.affiliation: Department of Applied Chemistry, National Defense Academy, Yokosuka, Kanagawa, Japan.
authors.forename: Tohru
authors.initials: T
authors.lastname: Ichimura
mesh_terms: D048948:14-3-3 Proteins / Q000378:metabolism; D000251:Adenosine Triphosphatases / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism*; D057809:HEK293 Cells; D018840:HSP70 Heat-Shock Proteins / Q000378:metabolism*; D006801:Humans; D002479:Inclusion Bodies / Q000378:metabolism*; D008751:Methylene Blue / Q000378:metabolism; D008954:Models, Biological; D046912:Multiprotein Complexes / Q000378:metabolism; D009124:Muscle Proteins / Q000378:metabolism; D011485:Protein Binding; D055550:Protein Stability; D015203:Reproducibility of Results; D014157:Transcription Factors / Q000378:metabolism*; D000070584:Tripartite Motif Proteins / Q000378:metabolism*; D044767:Ubiquitin-Protein Ligases / Q000378:metabolism*
chemical_list: D048948:14-3-3 Proteins; D018840:HSP70 Heat-Shock Proteins; D046912:Multiprotein Complexes; D009124:Muscle Proteins; D014157:Transcription Factors; D000070584:Tripartite Motif Proteins; D000255:Adenosine Triphosphate; C094027:TRIM32 protein, human; C440908:TRIM63 protein, human; D044767:Ubiquitin-Protein Ligases; D000251:Adenosine Triphosphatases; D008751:Methylene Blue