PXD027664 is an
original dataset announced via ProteomeXchange.
Dataset Summary
Title | N-Acetyl-β-D-hexosaminidases mediate the generation of paucimannosidic proteins via a putative non-canonical truncation pathway in human neutrophils |
Description | We recently discovered that human neutrophils express immunomodulatory glycoproteins carrying unusual and highly truncated paucimannosidic N-glycans (Man1-3GlcNAc2Fuc0-1), but their biosynthesis remains elusive. Guided by the well-characterised truncation pathway in invertebrates and plants in which the N-acetyl-β-D-hexosaminidase (Hex) isoenzymes catalyse paucimannosidic protein (PMP) formation, we here set out to test if the homologous human Hex α and β subunits encoded by HEXA and HEXB drive a similar truncation pathway in human neutrophils. To this end, we performed quantitative glycomics and glycoproteomics of several CRISPR-Cas9-edited Hex-disrupted neutrophil-like HL-60 mutants (HEXA-/- and HEXB-/-) and matching unedited cell lines. Hex disruption was validated using next-generation sequencing, ELISA, proteomics and Hex activity assays. Excitingly, all Hex-disrupted mutants displayed significantly reduced levels of paucimannosylation, particularly of Man2-3GlcNAc2Fuc1, relative to unedited HL-60 suggesting that both HEXA and HEXB contribute to PMP formation via a hitherto unexplored truncation pathway in neutrophils. Quantitative N-glycomics indeed demonstrated reduced utilisation of a putative non-canonical truncation pathway in favour of the canonical elongation pathway in all Hex-disrupted mutants relative to unedited controls. Quantitative glycoproteomics recapitulated the truncation-to-elongation switch in all Hex-deficient mutants and showed a pronounced switch for N-glycoproteins co-trafficking with Hex to the azurophilic granules of neutrophils such as myeloperoxidase. Finally, we supported the Hex-PMP relationship by documenting that primary neutrophils isolated from an early-onset Sandhoff disease patient (HEXB-/-) displayed dramatically reduced paucimannosylation relative to neutrophils from an age-matched unaffected donor. We conclude that both human Hex α and β mediate PMP formation via a putative non-canonical truncation pathway in neutrophils. |
HostingRepository | PRIDE |
AnnounceDate | 2023-11-14 |
AnnouncementXML | Submission_2023-11-14_08:40:50.733.xml |
DigitalObjectIdentifier | |
ReviewLevel | Peer-reviewed dataset |
DatasetOrigin | Original dataset |
RepositorySupport | Unsupported dataset by repository |
PrimarySubmitter | Julian Ugonotti |
SpeciesList | scientific name: Homo sapiens (Human); NCBI TaxID: 9606; |
ModificationList | TMT6plex-126 reporter+balance reagent acylated residue |
Instrument | Q Exactive HF |
Dataset History
Revision | Datetime | Status | ChangeLog Entry |
0 | 2021-07-30 03:44:49 | ID requested | |
1 | 2023-03-11 02:34:11 | announced | |
⏵ 2 | 2023-11-14 08:40:52 | announced | 2023-11-14: Updated project metadata. |
Publication List
Ugonotti J, Kawahara R, Loke I, Zhu Y, Chatterjee S, Tjondro HC, Sumer-Bayraktar Z, Neelamegham S, Thaysen-Andersen M, -D-hexosaminidases mediate the generation of paucimannosidic proteins via a putative noncanonical truncation pathway in human neutrophils. Glycobiology, 32(3):218-229(2022) [pubmed] |
Keyword List
ProteomeXchange project tag: Biology/Disease-Driven Human Proteome Project (B/D-HPP), Human Proteome Project, Glycoproteomics (B/D-HPP) |
submitter keyword: N-Acetyl-β-D-hexosaminidase, biosynthesis, paucimannose, truncation, neutrophil, mass spectrometry, N-glycosylation |
Contact List
Dr. Morten Thaysen-Andersen |
contact affiliation | Department of Molecular Sciences, Macquarie University, Sydney, NSW, Australia |
contact email | morten.andersen@mq.edu.au |
lab head | |
Julian Ugonotti |
contact affiliation | Macquarie University |
contact email | julian.ugonotti@hdr.mq.edu.au |
dataset submitter | |
Full Dataset Link List
Dataset FTP location
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PRIDE project URI |
Repository Record List
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[ - ]
- PRIDE
- PXD027664
- Label: PRIDE project
- Name: N-Acetyl-β-D-hexosaminidases mediate the generation of paucimannosidic proteins via a putative non-canonical truncation pathway in human neutrophils