{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,15]],"date-time":"2026-10-15T21:32:16Z","timestamp":1792099936196,"version":"4.4.0"},"reference-count":56,"publisher":"Oxford University Press (OUP)","issue":"3","license":[{"start":{"date-parts":[[2021,10,18]],"date-time":"2021-10-18T00:00:00Z","timestamp":1634515200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/academic.oup.com\/journals\/pages\/open_access\/funder_policies\/chorus\/standard_publication_model"}],"funder":[{"DOI":"10.13039\/100000002","name":"National Institutes of Health","doi-asserted-by":"publisher","award":["HL103411"],"award-info":[{"award-number":["HL103411"]}],"id":[{"id":"10.13039\/100000002","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"published-print":{"date-parts":[[2022,3,30]]},"abstract":"<jats:title>Abstract<\/jats:title>\n               <jats:p>We recently discovered that human neutrophils express immunomodulatory glycoproteins carrying unusual and highly truncated paucimannosidic N-glycans (Man1-3GlcNAc2Fuc0\u20131), but their biosynthesis remains elusive. Guided by the well-characterized truncation pathway in invertebrates and plants in which the N-acetyl-\u03b2-D-hexosaminidase (Hex) isoenzymes catalyze paucimannosidic protein (PMP) formation, we here set out to test if the homologous human Hex \u03b1 and \u03b2 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-KO and HEXB-KO) and matching unedited cell lines. Hex disruption was validated using next-generation sequencing, enzyme-linked immunosorbent assay (ELISA), quantitative proteomics and Hex activity assays. Excitingly, all Hex-disrupted mutants displayed significantly reduced levels of paucimannosylation, particularly 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 utilization of a putative noncanonical truncation pathway in favor 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-disrupted mutants and showed a greater switch for N-glycoproteins cotrafficking 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\u2212\/\u2212) displayed dramatically reduced paucimannosylation relative to neutrophils from an age-matched unaffected donor. We conclude that both human Hex \u03b1 and \u03b2 mediate PMP formation via a putative noncanonical truncation pathway in neutrophils.<\/jats:p>","DOI":"10.1093\/glycob\/cwab108","type":"journal-article","created":{"date-parts":[[2021,10,16]],"date-time":"2021-10-16T01:15:42Z","timestamp":1634346942000},"page":"218-229","source":"Crossref","is-referenced-by-count":28,"title":["<i>N<\/i>-acetyl-\u03b2-D-hexosaminidases mediate the generation of paucimannosidic proteins via a putative noncanonical truncation pathway in human neutrophils"],"prefix":"10.1093","volume":"32","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-3455-6766","authenticated-orcid":false,"given":"Julian","family":"Ugonotti","sequence":"first","affiliation":[{"name":"Department of Molecular Sciences, Macquarie University, Balaclava Road, Macquarie Park, Sydney, NSW 2109, Australia"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-0292-2637","authenticated-orcid":false,"given":"Rebeca","family":"Kawahara","sequence":"additional","affiliation":[{"name":"Department of Molecular Sciences, Macquarie University, Balaclava Road, Macquarie Park, Sydney, NSW 2109, Australia"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2558-0975","authenticated-orcid":false,"given":"Ian","family":"Loke","sequence":"additional","affiliation":[{"name":"Cordlife Group Limited, 1 Yishun Industrial Street, Singapore 768160, Singapore"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yuqi","family":"Zhu","sequence":"additional","affiliation":[{"name":"Department of Chemical and Biological Engineering, University at Buffalo, State University of New York, 906 Furnas Hall, Buffalo, NY 14260, 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Sydney, NSW 2109, Australia"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-1371-8500","authenticated-orcid":false,"given":"Sriram","family":"Neelamegham","sequence":"additional","affiliation":[{"name":"Department of Chemical and Biological Engineering, University at Buffalo, State University of New York, 906 Furnas Hall, Buffalo, NY 14260, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-8327-6843","authenticated-orcid":false,"given":"Morten","family":"Thaysen-Andersen","sequence":"additional","affiliation":[{"name":"Department of Molecular Sciences, Macquarie University, Balaclava Road, Macquarie Park, Sydney, NSW 2109, Australia"},{"name":"Biomolecular Discovery Research Centre, Macquarie University, Balaclava Road, Macquarie Park, Sydney, NSW 2109, 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