CoChat
Sign Up
Cited by 16Ubiquitin and proteasome pathwaysCellular transport and secretionGenetics and Neurodevelopmental Disorders

Mono-ubiquitination of Rabphilin 3A by UBE3A serves a non-degradative function

Open access
Avagliano Trezza, Rossella, Punt, A. Mattijs, Mientjes, Edwin, van den Berg, Marlene, Zampeta, F. Isabella, de Graaf, Ilona J., van der Weegen, Yana, Demmers, Jeroen A. A. +2 more·2021·Scientific Reports
DOI 10.1038/s41598-021-82319-9PMC PMC7862399

Published version of a preprint

Open this paper in CoChat — free accountOpen PDF

Abstract

Angelman syndrome (AS) is a severe neurodevelopmental disorder caused by brain-specific loss of UBE3A, an E3 ubiquitin protein ligase. A substantial number of possible ubiquitination targets of UBE3A have been identified, although evidence of being direct UBE3A substrates is often lacking. Here we identified the synaptic protein Rabphilin-3a (RPH3A), an effector of the RAB3A small GTPase involved in axonal vesicle priming and docking, as a ubiquitination target of UBE3A. We found that the UBE3A and RAB3A binding sites on RPH3A partially overlap, and that RAB3A binding to RPH3A interferes with UBE3A binding. We confirmed previous observations that RPH3A levels are critically dependent on RAB3A binding but, rather surprisingly, we found that the reduced RPH3A levels in the absence of RAB3A are not mediated by UBE3A. Indeed, while we found that RPH3A is ubiquitinated in a UBE3A-dependent manner in mouse brain, UBE3A mono-ubiquitinates RPH3A and does not facilitate RPH3A degradation. Moreover, we found that an AS-linked UBE3A missense mutation in the UBE3A region that interacts with RPH3A, abrogates the interaction with RPH3A. In conclusion, our results identify RPH3A as a novel target of UBE3A and suggest that UBE3A-dependent ubiquitination of RPH3A serves a non-degradative function.

The claims and statistics of this paper haven’t been checked yet.

Run the check — free, no account needed. It reads the open-access text and checks the sources it cites.

A few free checks a day · papers already verified open instantly · results are public

Integrity signals

Registry

Crossref · 10.1038/s41598-021-82319-9

Cites

2 of 2 references verified0 citation errors · 0 not checkable

Statistics

Claims the paper makes

The claims and statistics of this paper haven’t been checked yet.

Run the check — free, no account needed. It reads the open-access text and checks the sources it cites.

A few free checks a day · papers already verified open instantly · results are public

Claims other papers make about this one

Each is checked against this paper’s own text: does it say what the citing paper says it does?

  • Mutations in UBE3A and USP9X disrupt protein homeostasis by altering substrate protein half-life, abundance, and functional state.

    “Mutations in these genes disrupt protein homeostasis by altering substrate protein half-life, abundance, and functional state (Avagliano Trezza et al., 2021).”

    From Biological hierarchy from genotype to phenotype in syndromic and non-syndromic autism spectrum disorder (2026) · cites this paper as Avagliano Trezza et al., 2021

    The Results report that an Angelman syndrome-linked UBE3A variant reduces binding to RPH3A. But the Results also find no significant change in RPH3A abundance, and the Discussion says its ubiquitination does not affect protein stability; a possible effect on activity remains a hypothesis. The paper does not address USP9X or establish the claimed changes in substrate half-life and abundance. (LLM, conf=0.88)

    Experimental — accuracy not yet measured

Self-check

Percentages, sums and reported differences re-computed from the paper’s own numbers.

The self-check needs the paper’s full text.

Cites

Flagged first, then by citations

Research works printed but not linked: 47

  1. [1]Buiting K Williams C Horsthemke B Angelman syndrome—Insights into a rare neurogenetic disorder Nat. Rev. Neurol. 2016 12 584 593 10.1038/nrneurol.2016.133 27615419 · DOI
  2. [2]Scheffner M Huibregtse JM Vierstra RD Howley PM The HPV-16 E6 and E6-AP complex functions as a ubiquitin-protein ligase in the ubiquitination of p53 Cell 1993 75 495 505 10.1016/0092-8674(93)90384-3 8221889 · DOI
  3. [4]Komander D Rape M The ubiquitin code Annu. Rev. Biochem. 2012 81 203 229 10.1146/annurev-biochem-060310-170328 22524316 · DOI · Paper page
  4. [5]Swatek KN Komander D Ubiquitin modifications Cell Res. 2016 26 399 422 10.1038/cr.2016.39 27012465 PMC4822133 · DOI
  5. [6]Kobayashi F Nishiuchi T Takaki K Konno H Ubiquitin chain specificities of E6AP E3 ligase and its HECT domain Biochem. Biophys. Res. Commun. 2018 496 686 692 10.1016/j.bbrc.2017.12.076 29288669 · DOI
  6. [7]Lasalle JM Reiter LT Chamberlain SJ Epigenetic regulation of UBE3Aand roles in human neurodevelopmental disorders Epigenomics 2015 7 1213 1228 10.2217/epi.15.70 26585570 PMC4709177 · DOI
  7. [8]Südhof TC Rizo J Synaptic vesicle exocytosis CSH Perspect. Biol. 2011 3 a005637 a005637 PMC3225952 22026965
  8. [9]Cortès-Saladelafont E Diseases of the synaptic vesicle: A potential new group of neurometabolic disorders affecting neurotransmission Semin. Pediatr. Neurol. 2016 23 306 320 10.1016/j.spen.2016.11.005 28284392 · DOI
  9. [10]Wallace ML Burette AC Weinberg RJ Philpot BD Maternal loss of Ube3a produces an excitatory/inhibitory imbalance through neuron type-specific synaptic defects Neuron 2012 74 793 800 10.1016/j.neuron.2012.03.036 22681684 PMC3372864 · DOI · Paper page
  10. [11]Fukuda M Distinct Rab binding specificity of Rim1, Rim2, Rabphilin, and Noc2 J. Biol. Chem. 2003 278 15373 15380 10.1074/jbc.M212341200 12578829 · DOI
  11. [12]Stenmark H Rab GTPases as coordinators of vesicle traffic Nat. Rev. Mol. Cell Biol. 2009 10 513 525 10.1038/nrm2728 19603039 · DOI · Paper page
  12. [13]Schlüter OM Schmitz F Jahn R Rosenmund C Südhof TC A complete genetic analysis of neuronal Rab3 function J. Neurosci. 2004 24 6629 6637 10.1523/JNEUROSCI.1610-04.2004 15269275 PMC6729882 · DOI
  13. [14]Geppert M The role of Rab3A in neurotransmitter release Nature 1994 369 493 497 10.1038/369493a0 7911226 · DOI
  14. [15]Johannes L Evidence for a functional link between Rab3 and the SNARE complex J. Cell Sci. 1996 109 Pt 12 2875 2884 9013335
  15. [16]Araki K Purification and characterization of Rab GDIβ from rat brain Biochem. Biophys. Res. Comm. 1995 211 296 305 10.1006/bbrc.1995.1810 7779099 · DOI
  16. [17]Ostermeier C Brunger AT Structural basis of Rab effector specificity: crystal structure of the small G protein Rab3A complexed with the effector domain of rabphilin-3A Cell 1999 96 363 374 10.1016/S0092-8674(00)80549-8 10025402 · DOI
  17. [18]Schlüter OM Rabphilin knock-out mice reveal that rabphilin is not required for rab3 function in regulating neurotransmitter release J. Neurosci. 1999 19 5834 5846 10.1523/JNEUROSCI.19-14-05834.1999 10407024 PMC6783077 · DOI
  18. [19]Avagliano Trezza, R. et al. Loss of nuclear UBE3A causes electrophysiological and behavioral deficits in mice and is associated with Angelman syndrome. Nat. Neurosci.22, 1235–1247 (2019). 31235931
  19. [20]Clabecq A Henry J-P Darchen F Biochemical characterization of Rab3-GTPase-activating protein reveals a mechanism similar to that of Ras-GAP J. Biol. Chem. 2000 275 31786 31791 10.1074/jbc.M003705200 10859313 · DOI
  20. [21]Handley MTW Burgoyne RD The Rab27 effector Rabphilin, unlike Granuphilin and Noc2, rapidly exchanges between secretory granules and cytosol in PC12 cells Biochem. Biophys. Res. Comm. 2008 373 275 281 10.1016/j.bbrc.2008.06.043 18573236 · DOI
  21. [22]Wang T Van Woerden GM Elgersma Y Borst JGG Enhanced transmission at the calyx of Held synapse in a mouse model for Angelman syndrome Front. Cell. Neurosci. 2018 11 741 10.3389/fnins.2017.00741 PMC5758499 29354033 · DOI
  22. [23]van der Wal L Improvement of ubiquitylation site detection by Orbitrap mass spectrometry J. Proteomics 2018 172 49 56 10.1016/j.jprot.2017.10.014 29122726 · DOI
  23. [24]Keren-Kaplan T Synthetic biology approach to reconstituting the ubiquitylation cascade in bacteria EMBO J. 2011 31 378 390 10.1038/emboj.2011.397 22081111 PMC3261559 · DOI
  24. [25]Zaaroor-Regev D Regulation of the polycomb protein Ring1B by self-ubiquitination or by E6-AP may have implications to the pathogenesis of Angelman syndrome Proc. Natl. Acad. Sci. USA 2010 107 6788 6793 10.1073/pnas.1003108107 20351251 PMC2872415 · DOI
  25. [26]Greer PL The Angelman syndrome protein Ube3A regulates synapse development by ubiquitinating arc Cell 2010 140 704 716 10.1016/j.cell.2010.01.026 20211139 PMC2843143 · DOI · Paper page
  26. [27]Kühnle S Mothes B Matentzoglu K Scheffner M Role of the ubiquitin ligase E6AP/UBE3A in controlling levels of the synaptic protein Arc Proc. Natl. Acad. Sci. USA 2013 110 8888 8893 10.1073/pnas.1302792110 23671107 PMC3670309 · DOI
  27. [28]Mortensen F Role of ubiquitin and the HPV E6 oncoprotein in E6AP-mediated ubiquitination Proc. Natl. Acad. Sci. USA 2015 112 9872 9877 10.1073/pnas.1505923112 26216987 PMC4538620 · DOI
  28. [29]Kao WH Beaudenon SL Talis AL Huibregtse JM Howley PM Human papillomavirus type 16 E6 induces self-ubiquitination of the E6AP ubiquitin-protein ligase J. Virol. 2000 74 6408 6417 10.1128/JVI.74.14.6408-6417.2000 10864652 PMC112148 · DOI
  29. [30]Martinez-Noël G Network analysis of UBE3A/E6AP-associated proteins provides connections to several distinct cellular processes J. Mol. Biol. 2018 430 1024 1050 10.1016/j.jmb.2018.01.021 29426014 PMC5866790 · DOI
  30. [31]Huibregtse JM Scheffner M Howley PM Localization of the E6-AP regions that direct human papillomavirus E6 binding, association with p53, and ubiquitination of associated proteins Mol. Cell. Biol. 1993 13 4918 4927 10.1128/MCB.13.8.4918 8393140 PMC360130 · DOI
  31. [32]Burns ME Sasaki T Takai Y Augustine GJ Rabphilin-3A: A multifunctional regulator of synaptic vesicle traffic J. Gen. Physiol. 1998 111 243 255 10.1085/jgp.111.2.243 9450942 PMC2222762 · DOI
  32. [33]Coppola T Rabphilin dissociated from Rab3 promotes endocytosis through interaction with Rabaptin-5 J. Cell Sci. 2001 114 1757 1764 11309205
  33. [34]Ohya T Sasaki T Kato M Takai Y Involvement of Rabphilin3 in endocytosis through interaction with Rabaptin5 J. Biol. Chem. 1998 273 613 617 10.1074/jbc.273.1.613 9417123 · DOI
  34. [35]Stahl B Chou JH Li C Südhof TC Jahn R Rab3 reversibly recruits rabphilin to synaptic vesicles by a mechanism analogous to raf recruitment by ras EMBO J. 1996 15 1799 1809 10.1002/j.1460-2075.1996.tb00529.x 8617225 PMC450096 · DOI
  35. [36]Stenmark H Vitale G Ullrich O Zerial M Rabaptin-5 is a direct effector of the small GTPase Rab5 in endocytic membrane-fusion Cell 1995 83 423 432 10.1016/0092-8674(95)90120-5 8521472 · DOI
  36. [37]Judson MC GABAergic neuron-specific loss of Ube3a causes Angelman syndrome-like EEG abnormalities and enhances seizure susceptibility Neuron 2016 90 56 69 10.1016/j.neuron.2016.02.040 27021170 PMC4824651 · DOI
  37. [38]Stanic J Rabphilin 3A retains NMDA receptors at synapticsites through interaction with GluN2A/PSD-95complex Nat. Commun. 2015 6 1 16 10.1038/ncomms10181 PMC4703873 26679993 · DOI
  38. [39]Gray JA Distinct modes of AMPA receptor suppression at developing synapses by GluN2A and GluN2B: Single-cell NMDA receptor subunit deletion in vivo Neuron 2011 71 1085 1101 10.1016/j.neuron.2011.08.007 21943605 PMC3183990 · DOI
  39. [40]Rauner C Köhr G Triheteromeric NR1/NR2A/NR2B receptors constitute the major N-methyl-d-aspartate receptor population in adult hippocampal synapses J. Biol. Chem. 2011 286 7558 7566 10.1074/jbc.M110.182600 21190942 PMC3045010 · DOI
  40. [41]Burette AC Subcellular organization of UBE3A in neurons J. Comp. Neurol. 2016 525 233 251 10.1002/cne.24063 27339004 PMC5138109 · DOI
  41. [42]Dindot SV Antalffy BA Bhattacharjee MB Beaudet AL The Angelman syndrome ubiquitin ligase localizes to the synapse and nucleus, and maternal deficiency results in abnormal dendritic spine morphology Hum. Mol. Genet. 2008 17 111 118 10.1093/hmg/ddm288 17940072 · DOI · Paper page
  42. [43]Burette AC Subcellular organization of UBE3A in human cerebral cortex Mol. Autism 2018 9 1 14 10.1186/s13229-018-0238-0 30364390 PMC6194692 · DOI
  43. [44]Yashiro K Ube3a is required for experience-dependent maturation of the neocortex Nat. Neurosci. 2009 12 777 783 10.1038/nn.2327 19430469 PMC2741303 · DOI · Paper page
  44. [46]Avagliano Trezza, R., van den Burg, J., van den Oever, N. & Distel, B. A versatile plasmid system for reconstitution and analysis of mammalian ubiquitination cascades in yeast. Microb. Cell5, 150–157 (2018). PMC5826702 29487861
  45. [47]Gkourtsa, A., van den Burg, J., Avula, T., Hochstenbach, F. & Ben Distel. Microbiological Research. Microbiol. Res.190, 27–36 (2016). 27393996
  46. [48]Nelson JK The deubiquitylase USP2 regulates the LDLR pathway by counteracting the E3-ubiquitin ligase IDOL Circ. Res. 2016 118 410 419 10.1161/CIRCRESAHA.115.307298 26666640 · DOI
  47. [49]Jiang Y-H Mutation of the Angelman ubiquitin ligase in mice causes increased cytoplasmic p53 and deficits of contextual learning and long-term potentiation Neuron 1998 21 799 811 10.1016/S0896-6273(00)80596-6 9808466 · DOI · Paper page

Cited by

Sorted by citations

Authors
Avagliano Trezza, R., Punt, A. M., Mientjes, E., van den Berg, M., Zampeta, F. I., de Graaf, I. J., van der Weegen, Y., Demmers, J. A. A., Elgersma, Y., & Distel, B.
Affiliations
Erasmus MC (NL)
Corresponding author
  • Elgersma, Ype
  • Distel, Ben
ORCID
Venue
Scientific Reports
Publisher
Springer Science and Business Media LLC
Published
2021-02-04
Volume
11
Issue
1
Pages
3007-3007
ISSN
2045-2322 (ISSN-L)
DOI
10.1038/s41598-021-82319-9
PMC
PMC7862399
OpenAlex
W3127239973
Cited by
16
References
2
Citations by year
2026: 5 · 2025: 4 · 2024: 3 · 2023: 1 · 2022: 1 · 2021: 2
FWCI
0.55
Type
journal article
Language
en
Open access
Licence
Locations
  • Scientific Reports (publishedVersion, cc-by)
  • bioRxiv (Cold Spring Harbor Laboratory) (acceptedVersion, cc-by-nc-nd)
  • Europe PMC (PubMed Central) (publishedVersion, other-oa)
  • PubMed Central (publishedVersion, cc-by)
  • PubMed (publishedVersion)
  • EUR Research Repository (Erasmus University Rotterdam) (publishedVersion, cc-by)
  • Data Archiving and Networked Services (DANS) (submittedVersion, other-oa)
  • EUR Research Repository (Erasmus University Rotterdam) (submittedVersion, cc-by)
  • RePub (Erasmus University Rotterdam) (submittedVersion, other-oa)
  • DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion, cc-by-sa)
  • Pure Amsterdam UMC (submittedVersion, cc-by)
Funders
  • Nederlandse Organisatie voor Wetenschappelijk Onderzoek (91209046)
  • Angelman Syndrome Alliance
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Ubiquitin and proteasome pathways
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Cell Biology › Cellular transport and secretion
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Genetics › Genetics and Neurodevelopmental Disorders
Keywords
Rabphilin-3A, UBE3A, monoubiquitination, Angelman syndrome, Rab3A, synaptic vesicle docking, E3 ubiquitin ligase, protein-protein interaction, mouse brain

About these checks

Registry, retraction, Cites and statistics results are computed, not judged, with one exception: a printed reference that no index lists is read once by a model before it counts as a citation error. Claims are extracted by a model and kept only when their quote appears verbatim in the text; each cited source is then checked, with a model’s help, for whether it says what the claim relies on. Those verdicts can be wrong — read the claim’s quote and the cited source before relying on one.

Statistics and arithmetic are recomputed, not judged.

Confidence bands are shown only for fields calibrated on our annotated test set. None is calibrated yet, so no band is shown.

What counts as a claim ›How we measure confidence ›How we check this ›

Are you an author, or do you see a problem with this page? Request a review