Francesco Scavone

All Publications

• UFMylation orchestrates spatiotemporal coordination of RQC at the ER. Science advances Penchev, I., Gumbin, S., Scavone, F., Berninghausen, O., Becker, T., Kopito, R., Beckmann, R. 2025; 11 (18): eadv0435

  Abstract

  Degradation of arrest peptides from endoplasmic reticulum (ER) translocon-bound 60S ribosomal subunits via the ribosome-associated quality control (ER-RQC) pathway requires covalent modification of RPL26/uL24 on 60S ribosomal subunits with UFM1. However, the underlying mechanism that coordinates the UFMylation and RQC pathways remains elusive. Structural analysis of ER-RQC intermediates revealed concomitant binding and direct interaction of the UFMylation and RQC machineries on the 60S. In the presence of an arrested peptidyl-transfer RNA, the RQC factor NEMF and the UFM1 E3 ligase (E3UFM1) form a direct interaction via the UFL1 subunit of E3UFM1, and UFL1 adopts a conformation distinct from that previously observed for posttermination 60S. While this concomitant binding occurs on translocon-bound 60S, LTN1 recruitment and arrest peptide degradation require UFMylation-dependent 60S dissociation from the translocon. These data reveal a mechanism by which the UFMylation cycle orchestrates ER-RQC.

  View details for DOI 10.1126/sciadv.adv0435

  View details for PubMedID 40315331

  View details for PubMedCentralID PMC12047416

• Trafficking of K63-polyubiquitin modified membrane proteins in a macroautophagy-independent pathway is linked to ATG9A. Molecular biology of the cell Scavone, F., Lian, S., Eskelinen, E. L., Cohen, R. E., Yao, T. 2025: mbcE24120535

  Abstract

  Cytoplasmic K63-linked polyubiquitin signals have well-established roles in endocytosis and selective autophagy. However, how these signals help to direct different cargos to different intracellular trafficking routes is unclear. Here we report that, when the K63-polyubiquitin signal is blocked by intracellular expression of a high-affinity sensor (named Vx3), many proteins originating from the plasma membrane are found trapped in clusters of small vesicles that co-localize with ATG9A, a transmembrane protein that plays an essential role in autophagy. Importantly, whereas ATG9A is required for cluster formation, other core autophagy machinery as well as selective autophagy cargo receptors are not required. Although the cargos are sequestered in the vesicular clusters in an ATG9-dependent manner, additional signals are needed to induce LC3 conjugation. Upon removal of the Vx3 block, K63-polyubiquitylated cargos are rapidly delivered to lysosomes. These observations suggest that ATG9A plays an unexpected role in the trafficking of K63-polyubiquitin modified membrane proteins. [Media: see text] [Media: see text] [Media: see text].

  View details for DOI 10.1091/mbc.E24-12-0535

  View details for PubMedID 39969968

• STING induces HOIP-mediated synthesis of M1 ubiquitin chains to stimulate NF-κB signaling. The EMBO journal Fischer, T. D., Bunker, E. N., Zhu, P. P., Le Guerroué, F., Hadjian, M., Dominguez-Martin, E., Scavone, F., Cohen, R., Yao, T., Wang, Y., Werner, A., Youle, R. J. 2024

  Abstract

  STING activation by cyclic dinucleotides induces IRF3- and NF-κB-mediated gene expression in mammals, as well as lipidation of LC3B at Golgi-related membranes. While mechanisms of the IRF3 response are well understood, the mechanisms of NF-κB activation via STING remain unclear. We report here that STING activation induces linear/M1-linked ubiquitin chain (M1-Ub) formation and recruitment of the LUBAC E3 ligase, HOIP, to LC3B-associated Golgi membranes where ubiquitin is also localized. Loss of HOIP prevents formation of M1-Ub chains and reduces STING-induced NF-κB and IRF3 signaling in human THP1 monocytes and mouse bone marrow-derived macrophages, without affecting STING activation. STING-induced LC3B lipidation is not required for M1-Ub chain formation or for immune-related gene expression, but the recently reported STING function in neutralizing Golgi pH may be involved. Thus, LUBAC synthesis of M1-linked ubiquitin chains mediates STING-induced innate immune signaling.

  View details for DOI 10.1038/s44318-024-00291-2

  View details for PubMedID 39578541

  View details for PubMedCentralID 4019140

• UFM1 E3 ligase promotes recycling of 60S ribosomal subunits from the ER. Nature DaRosa, P. A., Penchev, I., Gumbin, S. C., Scavone, F., Wąchalska, M., Paulo, J. A., Ordureau, A., Peter, J. J., Kulathu, Y., Harper, J. W., Becker, T., Beckmann, R., Kopito, R. R. 2024

  Abstract

  Reversible modification of target proteins by ubiquitin and ubiquitin-like proteins (UBLs) is widely used by eukaryotic cells to control protein fate and cell behaviour1. UFM1 is a UBL that predominantly modifies a single lysine residue on a single ribosomal protein, uL24 (also called RPL26), on ribosomes at the cytoplasmic surface of the endoplasmic reticulum (ER)2,3. UFM1 conjugation (UFMylation) facilitates the rescue of 60S ribosomal subunits (60S) that are released after ribosome-associated quality-control-mediated splitting of ribosomes that stall during co-translational translocation of secretory proteins into the ER3,4. Neither the molecular mechanism by which the UFMylation machinery achieves such precise target selection nor how this ribosomal modification promotes 60S rescue is known. Here we show that ribosome UFMylation in vivo occurs on free 60S and we present sequential cryo-electron microscopy snapshots of the heterotrimeric UFM1 E3 ligase (E3(UFM1)) engaging its substrate uL24. E3(UFM1) binds the L1 stalk, empty transfer RNA-binding sites and the peptidyl transferase centre through carboxy-terminal domains of UFL1, which results in uL24 modification more than 150 Å away. After catalysing UFM1 transfer, E3(UFM1) remains stably bound to its product, UFMylated 60S, forming a C-shaped clamp that extends all the way around the 60S from the transfer RNA-binding sites to the polypeptide tunnel exit. Our structural and biochemical analyses suggest a role for E3(UFM1) in post-termination release and recycling of the large ribosomal subunit from the ER membrane.

  View details for DOI 10.1038/s41586-024-07073-0

  View details for PubMedID 38383785

  View details for PubMedCentralID 6347690

• RPL26/uL24 UFMylation is essential for ribosome-associated quality control at the endoplasmic reticulum. Proceedings of the National Academy of Sciences of the United States of America Scavone, F., Gumbin, S. C., Da Rosa, P. A., Kopito, R. R. 2023; 120 (16): e2220340120

  Abstract

  Ribosomes that stall while translating cytosolic proteins are incapacitated by incomplete nascent chains, termed "arrest peptides" (APs) that are destroyed by the ubiquitin proteasome system (UPS) via a process known as the ribosome-associated quality control (RQC) pathway. By contrast, APs on ribosomes that stall while translocating secretory proteins into the endoplasmic reticulum (ER-APs) are shielded from cytosol by the ER membrane and the tightly sealed ribosome-translocon junction (RTJ). How this junction is breached to enable access of cytosolic UPS machinery and 26S proteasomes to translocon- and ribosome-obstructing ER-APs is not known. Here, we show that UPS and RQC-dependent degradation of ER-APs strictly requires conjugation of the ubiquitin-like (Ubl) protein UFM1 to 60S ribosomal subunits at the RTJ. Therefore, UFMylation of translocon-bound 60S subunits modulates the RTJ to promote access of proteasomes and RQC machinery to ER-APs.

  View details for DOI 10.1073/pnas.2220340120

  View details for PubMedID 37036982

• RPL26/uL24 UFMylation is essential for ribosome-associated quality control at the endoplasmic reticulum. bioRxiv : the preprint server for biology Scavone, F., Gumbin, S. C., DaRosa, P. A., Kopito, R. R. 2023

  Abstract

  Ribosomes that stall while translating cytosolic proteins are incapacitated by incomplete nascent chains, termed "arrest peptides" (APs) that are destroyed by the ubiquitin proteasome system (UPS) via a process known as the ribosome-associated quality control (RQC) pathway. By contrast, APs on ribosomes that stall while translocating secretory proteins into the endoplasmic reticulum (ER-APs) are shielded from cytosol by the ER membrane and the tightly sealed ribosome-translocon junction (RTJ). How this junction is breached to enable access of cytosolic UPS machinery and 26S proteasomes to translocon- and ribosome-obstructing ER-APs is not known. Here, we show that UPS and RQC-dependent degradation of ER-APs strictly requires conjugation of the ubiquitin-like (Ubl) protein UFM1 to 60S ribosomal subunits at the RTJ. Therefore, UFMylation of translocon-bound 60S subunits modulates the RTJ to promote access of proteasomes and RQC machinery to ER-APs.UFM1 is a ubiquitin-like protein that is selectively conjugated to the large (60S) subunit of ribosomes bound to the endoplasmic reticulum (ER), but the specific biological function of this modification is unclear. Here, we show that UFMylation facilitates proteasome-mediated degradation of arrest polypeptides (APs) which are generated following splitting of ribosomes that stall during co-translational translocation of secretory proteins into the ER. We propose that UFMylation weakens the tightly sealed ribosome-translocon junction, thereby allowing the cytosolic ubiquitin-proteasome and ribosome-associated quality control machineries to access ER-APs.

  View details for DOI 10.1101/2023.03.08.531792

  View details for PubMedID 36945571

  View details for PubMedCentralID PMC10028864