UFSP1

UFSP1 is a papain-like cysteine protease that regulates the UFM1 conjugation (UFMylation) system by controlling both maturation of the UFM1 modifier and recycling of UFMylated targets (PMID:17182609, PMID:18321862, PMID:35926457). It specifically cleaves the C-terminal extension of pro-UFM1 to generate the mature form competent for conjugation and releases UFM1 from conjugated proteins, acting on UFM1 but not on ubiquitin, SUMO-1, or ISG15; catalysis depends on an active-site cysteine within an Asp175-Pro-His177 triad and a Tyr41 oxyanion hole, and the enzyme binds UFM1 with sub-micromolar affinity (PMID:17182609, PMID:18321862). In human cells UFSP1 is produced from a non-canonical translation start site and is the principal protease for UFM1 maturation and for cleaving an autoinhibitory UFMylation on the E2 enzyme UFC1, thereby gating activation of the pathway; this is a division of labor distinct from UFSP2, which deUFMylates the ribosomal subunit RPL26, while both proteases together are required for efficient deUFMylation and neither efficiently cleaves UFM1 from histone H4K31 within the nucleosome owing to steric occlusion (PMID:35926457, PMID:41964563, PMID:41759738). Independently of its catalytic activity, UFSP1 binds the UBA domain of the autophagy receptor p62/SQSTM1 and promotes p62-body formation by enhancing p62 engagement with ubiquitinated proteins (PMID:37285312). Loss of Ufsp1 in mice is compatible with normal development and homeostasis, with only mild structural alterations at the neuromuscular junction where the protein is enriched (PMID:40748989).

1. 2006 High

   Established that UFSP1 is the dedicated protease for the UFM1 modifier, answering whether a specific deconjugase/maturase exists for this Ubl.

   Evidence In vitro cleavage assays against multiple Ubl substrates, active-site Cys→Ser mutagenesis, NEM inhibition, and Ufm1-vinylmethylester activity-based labeling (mouse)

   PMID:17182609

   Open questions at the time

   • Did not define structural basis of UFM1 recognition
   • Cellular substrates and physiological role not addressed
2. 2008 High

   Defined the catalytic architecture and UFM1-binding determinants, explaining how UFSP1 achieves its specificity as a novel papain-like protease.

   Evidence 1.7 Å crystal structure of mouse UfSP1 with active-site mutagenesis, ITC binding measurement, and NMR mapping of the UFM1 interface

   PMID:18321862

   Open questions at the time

   • Structure is of the apo/mouse enzyme without bound substrate complex
   • Human enzyme behavior not addressed
3. 2022 High

   Resolved how active human UFSP1 is generated and partitioned the pathway, showing it matures UFM1 and de-represses UFC1 while UFSP2 handles RPL26.

   Evidence Non-canonical start-site identification plus UFSP1/UFSP2 double-knockout cells with biochemical readouts for UFM1 maturation, UFC1 deUFMylation, and RPL26 deUFMylation

   PMID:35926457

   Open questions at the time

   • Mechanism of non-canonical translation regulation unresolved
   • Physiological consequences of UFC1 autoinhibition control not defined in vivo
4. 2023 Medium

   Uncovered a protease-independent function in selective autophagy, addressing whether UFSP1 acts only as a deconjugase.

   Evidence BioID proximity proteomics, reciprocal Co-IP, immunofluorescence colocalization, and catalytic-mutant comparison for p62-body formation

   PMID:37285312

   Open questions at the time

   • Single-lab finding
   • Structural basis of UBA-domain binding not defined
   • In vivo relevance of p62-body promotion unknown
5. 2025 Medium

   Tested the organismal requirement for UFSP1, showing it is dispensable for mouse development despite NMJ enrichment.

   Evidence Ufsp1 knockout and catalytic-dead point-mutant mice with Mendelian ratio analysis and NMJ structural/functional assays

   PMID:40748989

   Open questions at the time

   • Mild NMJ phenotype mechanism unexplained
   • Possible redundancy with UFSP2 not tested in vivo
   • Single lab, negative/mild result
6. 2026 Medium

   Defined a substrate context limit, showing nucleosomal H4K31-UFM1 resists both UFSPs due to steric hindrance.

   Evidence Semisynthetic ufmylated histone deUFMylation assays and cryo-EM of the H4K31UFM1-nucleosome

   PMID:41964563

   Open questions at the time

   • Whether another factor removes nucleosomal UFM1 is unknown
   • Single lab, no replication
   • In vivo relevance of persistent histone UFMylation not addressed

• How UFSP1's enzymatic and protease-independent (p62) activities are coordinated, and what its physiological substrate repertoire and regulation are in vivo, remain open.
• No in vivo substrate catalog
• Coordination of dual functions unresolved
• Regulation of non-canonical translation not mechanistically defined