BioE3 identifies specific substrates of ubiquitin E3 ligases

  • Nat Commun. 2023 Nov 23;14(1):7656. doi: 10.1038/s41467-023-43326-8.
Orhi Barroso-Gomila  #  1 ,  Laura Merino-Cacho  #  1 ,  Veronica Muratore  1 ,  Coralia Perez  1 ,  Vincenzo Taibi  2 ,  Elena Maspero  2 ,  Mikel Azkargorta  1  3 ,  Ibon Iloro  1  3 ,  Fredrik Trulsson  4 ,  Alfred C O Vertegaal  4 ,  Ugo Mayor  5  6 ,  Felix Elortza  1  3 ,  Simona Polo  2  7 ,  Rosa Barrio  8 ,  James D Sutherland  9
Affiliations
  • 1. Center for Cooperative Research in Biosciences (CIC bioGUNE), Basque Research and Technology Alliance (BRTA), Bizkaia Technology Park, Building 801A, 48160, Derio, Spain.
  • 2. IFOM ETS, The AIRC Institute of Molecular Oncology, Milan, Italy.
  • 3. CIBERehd, Instituto de Salud Carlos III, C/ Monforte de Lemos 3-5, Pabellón 11, Planta 0, 28029, Madrid, Spain.
  • 4. Cell and Chemical Biology, Leiden University Medical Center (LUMC), 2333, ZA, Leiden, The Netherlands.
  • 5. Ikerbasque, Basque Foundation for Science, 48011, Bilbao, Spain.
  • 6. Biochemistry and Molecular Biology Department, University of the Basque Country (UPV/EHU), E-48940, Leioa, Spain.
  • 7. Dipartimento di oncologia ed emato-oncologia, Università degli Studi di Milano, Milan, Italy.
  • 8. Center for Cooperative Research in Biosciences (CIC bioGUNE), Basque Research and Technology Alliance (BRTA), Bizkaia Technology Park, Building 801A, 48160, Derio, Spain. [email protected].
  • 9. Center for Cooperative Research in Biosciences (CIC bioGUNE), Basque Research and Technology Alliance (BRTA), Bizkaia Technology Park, Building 801A, 48160, Derio, Spain. [email protected].
  • # Contributed equally.
PMID: 37996419 DOI: 10.1038/s41467-023-43326-8
Abstract

Hundreds of E3 Ligases play a critical role in recognizing specific substrates for modification by ubiquitin (Ub). Separating genuine targets of E3s from E3-interactors remains a challenge. We present BioE3, a powerful approach for matching substrates to Ub E3 Ligases of interest. Using BirA-E3 Ligase fusions and bioUb, site-specific biotinylation of Ub-modified substrates of particular E3s facilitates proteomic identification. We show that BioE3 identifies both known and new targets of two RING-type E3 ligases: RNF4 (DNA damage response, PML bodies), and MIB1 (endocytosis, Autophagy, centrosome dynamics). Versatile BioE3 identifies targets of an organelle-specific E3 (MARCH5) and a relatively uncharacterized E3 (RNF214). Furthermore, BioE3 works with NEDD4, a HECT-type E3, identifying new targets linked to vesicular trafficking. BioE3 detects altered specificity in response to chemicals, opening avenues for targeted protein degradation, and may be applicable for Other Ub-likes (UbLs, e.g., SUMO) and E3 types. BioE3 applications shed light on cellular regulation by the complex UbL network.

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