Analysis of endogenous NOTCH1 from POFUT1 S162L patient fibroblasts reveals the importance of the O-fucose modification on EGF12 in human development.

Item request has been placed! ×
Item request cannot be made. ×
loading   Processing Request
Read More Add to Saved list
  • Additional Information
    • Source:
      Publisher: IRL Press at Oxford University Press Country of Publication: England NLM ID: 9104124 Publication Model: Print Cited Medium: Internet ISSN: 1460-2423 (Electronic) Linking ISSN: 09596658 NLM ISO Abbreviation: Glycobiology Subsets: MEDLINE
    • Publication Information:
      Original Publication: Oxford ; New York : IRL Press at Oxford University Press, c1990-
    • Subject Terms:
      Fibroblasts*/metabolism ; Fucose*/metabolism ; Fucosyltransferases*/metabolism ; Fucosyltransferases*/genetics ; Receptor, Notch1*/metabolism ; Receptor, Notch1*/chemistry; Humans ; EGF Family of Proteins/metabolism
    • Abstract:
      NOTCH1 is a transmembrane receptor interacting with membrane-tethered ligands on opposing cells that mediate the direct cell-cell interaction necessary for many cell fate decisions. Protein O-fucosyltransferase 1 (POFUT1) adds O-fucose to Epidermal Growth Factor (EGF)-like repeats in the NOTCH1 extracellular domain, which is required for trafficking and signaling activation. We previously showed that POFUT1 S162L caused a 90% loss of POFUT1 activity and global developmental defects in a patient; however, the mechanism by which POFUT1 contributes to these symptoms is still unclear. Compared to controls, POFUT1 S162L patient fibroblast cells had an equivalent amount of NOTCH1 on the cell surface but showed a 60% reduction of DLL1 ligand binding and a 70% reduction in JAG1 ligand binding. To determine if the reduction of O-fucose on NOTCH1 in POFUT1 S162L patient fibroblasts was the cause of these effects, we immunopurified endogenous NOTCH1 from control and patient fibroblasts and analyzed O-fucosylation using mass spectral glycoproteomics methods. NOTCH1 EGF8 to EGF12 comprise the ligand binding domain, and O-fucose on EGF8 and EGF12 physically interact with ligands to enhance affinity. Glycoproteomics of NOTCH1 from POFUT1 S162L patient fibroblasts showed WT fucosylation levels at all sites analyzed except for a large decrease at EGF9 and the complete absence of O-fucose at EGF12. Since the loss of O-fucose on EGF12 is known to have significant effects on NOTCH1 activity, this may explain the symptoms observed in the POFUT1 S162L patient.
      (© The Author(s) 2024. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: [email protected].)
    • Comments:
      Update of: bioRxiv. 2024 Apr 09:2024.04.09.588484. doi: 10.1101/2024.04.09.588484. (PMID: 38645096)
    • References:
      Nature. 2020 Jun;582(7811):259-264. (PMID: 32499639)
      Sci Rep. 2020 Sep 30;10(1):16163. (PMID: 32999360)
      Science. 2005 Mar 11;307(5715):1599-603. (PMID: 15692013)
      Science. 2015 Feb 20;347(6224):847-53. (PMID: 25700513)
      J Biol Chem. 2016 Jul 29;291(31):16348-60. (PMID: 27268051)
      Pharmacol Res. 2016 Jun;108:57-64. (PMID: 27107790)
      Science. 2017 Mar 24;355(6331):1320-1324. (PMID: 28254785)
      Nat Commun. 2017 Aug 4;8(1):185. (PMID: 28775322)
      Nucleic Acids Res. 2022 Jan 7;50(D1):D543-D552. (PMID: 34723319)
      Nat Rev Mol Cell Biol. 2012 Jun 22;13(7):448-62. (PMID: 22722607)
      J Biol Chem. 2022 Jul;298(7):102064. (PMID: 35623385)
      Proc Natl Acad Sci U S A. 1994 Feb 1;91(3):913-7. (PMID: 8302866)
      J Cell Biol. 2009 Mar 9;184(5):621-9. (PMID: 19255248)
      Dev Cell. 2017 Jan 23;40(2):193-201. (PMID: 28089369)
      Glycobiology. 2019 Aug 20;29(9):620-624. (PMID: 31184695)
      Cell. 2002 Dec 13;111(6):893-904. (PMID: 12526814)
      J Biol Chem. 2012 Oct 5;287(41):33934-44. (PMID: 22872643)
      Nat Chem Biol. 2017 Jul;13(7):757-763. (PMID: 28530709)
      Life Sci Alliance. 2020 Oct 27;3(12):. (PMID: 33109684)
      J Biol Chem. 2017 Sep 22;292(38):15964-15973. (PMID: 28729422)
      J Biol Chem. 2022 Dec;298(12):102616. (PMID: 36265581)
      J Biol Chem. 2020 Oct 23;295(43):14710-14722. (PMID: 32820046)
      Proc Natl Acad Sci U S A. 2008 Feb 5;105(5):1539-44. (PMID: 18227520)
      Cell. 2009 Apr 17;137(2):216-33. (PMID: 19379690)
      Proc Natl Acad Sci U S A. 2014 May 20;111(20):7290-5. (PMID: 24803430)
      Glycobiology. 2018 May 1;28(5):276-283. (PMID: 29452367)
      Curr Top Dev Biol. 2010;92:1-29. (PMID: 20816391)
      BMC Dev Biol. 2019 Oct 7;19(1):19. (PMID: 31590629)
      Cell Rep. 2019 Nov 12;29(7):2054-2066.e6. (PMID: 31722217)
      Ann Rheum Dis. 2011 Jul;70(7):1304-10. (PMID: 21450749)
      Proc Natl Acad Sci U S A. 2003 Apr 29;100(9):5234-9. (PMID: 12697902)
      Cell. 2022 Jan 6;185(1):218. (PMID: 34995515)
      Mol Aspects Med. 2021 Jun;79:100938. (PMID: 33341260)
      Nat Rev Mol Cell Biol. 2006 Sep;7(9):678-89. (PMID: 16921404)
      J Biol Chem. 2001 Oct 26;276(43):40338-45. (PMID: 11524432)
    • Grant Information:
      R01 GM061126 United States GM NIGMS NIH HHS; R35 GM148433 United States GM NIGMS NIH HHS; Georgia Research Alliance
    • Contributed Indexing:
      Keywords: NOTCH1; O-fucose; POFUT1; mass spectrometry
    • Accession Number:
      28RYY2IV3F (Fucose)
      EC 2.4.1.- (Fucosyltransferases)
      0 (Receptor, Notch1)
      0 (NOTCH1 protein, human)
      EC 2.4.1.221 (polypeptide fucosyltransferase)
      0 (EGF Family of Proteins)
    • Publication Date:
      Date Created: 20240708 Date Completed: 20240716 Latest Revision: 20240929
    • Publication Date:
      20240929
    • Accession Number:
      PMC11249915
    • Accession Number:
      10.1093/glycob/cwae047
    • Accession Number:
      38976017