{"doi":"10.1101/2023.05.30.542711","title":"Rescue of secretion of a rare-disease associated mis-folded mutant glycoprotein in <i>UGGT1</i> knock-out mammalian cells","abstract":"Abstract Endoplasmic reticulum (ER) retention of mis-folded glycoproteins is mediated by the ER- localised eukaryotic glycoprotein secretion checkpoint, UDP-glucose glycoprotein glucosyl-transferase (UGGT). The enzyme recognises a mis-folded glycoprotein and flags it for ER retention by reglucosylating one of its N-linked glycans. In the background of a congenital mutation in a secreted glycoprotein gene, UGGT-mediated ER retention can cause rare disease even if the mutant glycoprotein retains activity (“responsive mutant”). Here, we investigated the subcellular localisation of the human Trop-2 Q118E variant, which causes gelatinous drop- like corneal dystrophy (GDLD). Compared with the wild type Trop-2, which is correctly localised at the plasma membrane, the Trop-2-Q118E variant is found to be heavily retained in the ER. Using Trop-2-Q118E, we tested UGGT modulation as a rescue-of-secretion therapeutic strategy for congenital rare disease caused by responsive mutations in genes encoding secreted glycoproteins. We investigated secretion of a EYFP-fusion of Trop-2-Q118E by confocal laser scanning microscopy. As a limiting case of UGGT inhibition, mammalian cells harbouring CRISPR/Cas9-mediated inhibition of the UGGT1 and/or UGGT2 gene expressions were used. The membrane localisation of the Trop-2-Q118E-EYFP mutant was successfully rescued in UGGT1 -/- and UGGT1/2 -/- cells. UGGT1 also efficiently reglucosylated Trop-2-Q118E-EYFP in cellula . The study supports the hypothesis that UGGT1 modulation constitutes a novel therapeutic strategy for the treatment of Trop-2-Q118E associated GDLD, and it encourages the testing of modulators of ER glycoprotein folding Quality Control (ERQC) as broad-spectrum rescue- of-secretion drugs in rare diseases caused by responsive secreted glycoprotein mutants. Synopsis Deletion of the UGGT1 and UGGT1/2 genes in HEK 293T cells rescues secretion of an EYFP-fusion of the human Trop-2-Q118E glycoprotein mutant. The mutant is retained in the secretory pathway in wild type cells and it localises to the cell membrane in UGGT1 -/- single and UGGT1/2 -/- double knock-out cells. The Trop-2-Q118E glycoprotein disease mutant is efficiently glucosylated by UGGT1 in human cells demonstrating that it is a bona fide cellular UGGT1 substrate.","journal":"bioRxiv (Cold Spring Harbor Laboratory)","year":2023,"id":399513,"datarank":0.0,"base_score":0.0,"endowment":0.0,"self_citation_contribution":0.0,"citation_network_contribution":0.0,"self_endowment_contribution":0.0,"citer_contribution":0.0,"corpus_percentile":null,"corpus_rank":null,"citation_count":1,"citer_count":0,"citers_with_citation_signal":0,"citers_with_endowment":0,"datacite_reuse_total":0,"is_dataset":false,"is_dataset_confidence":0.9503,"is_data_producer":false,"deposit_databanks":null,"is_oa":true,"file_count":0,"downloads":0,"has_version_chain":false,"published_date":"2023-01-01","fair_score":null,"fair_percentile":null,"algorithm_id":"datarank_citation_only_1hop_v6","ranking_scope":"data_only","authors":[{"id":320511,"name":"Kevin P. Guay","orcid":"0000-0002-3468-942X","position":1,"is_corresponding":false},{"id":1175953,"name":"Tatiana Soldà","orcid":null,"position":2,"is_corresponding":false},{"id":1055136,"name":"Charlie J. Hitchman","orcid":"0009-0003-2762-7057","position":3,"is_corresponding":false},{"id":989146,"name":"Johan C. Hill","orcid":"0000-0002-8816-7360","position":4,"is_corresponding":false},{"id":321587,"name":"Snežana Vasiljević","orcid":"0000-0002-8886-6225","position":5,"is_corresponding":false},{"id":989148,"name":"Andrea Lia","orcid":"0000-0003-4146-0647","position":6,"is_corresponding":false},{"id":743633,"name":"Carlos P. Modenutti","orcid":"0000-0001-5352-7234","position":7,"is_corresponding":false},{"id":719593,"name":"Kees R. Straatman","orcid":"0000-0002-9812-492X","position":8,"is_corresponding":false},{"id":989150,"name":"Angelo Santino","orcid":"0000-0002-3348-0747","position":9,"is_corresponding":false},{"id":1175635,"name":"Maurizio Molinari","orcid":"0000-0002-7636-5829","position":10,"is_corresponding":false},{"id":989151,"name":"Nicole Zitzmann","orcid":"0000-0003-1969-4949","position":11,"is_corresponding":false},{"id":320512,"name":"Daniel N. Hebert","orcid":"0000-0003-1537-4446","position":12,"is_corresponding":false},{"id":989152,"name":"Pietro Roversi","orcid":"0000-0001-9280-9437","position":13,"is_corresponding":false},{"id":1175636,"name":"Marco Trerotola","orcid":"0000-0003-1855-7002","position":14,"is_corresponding":false},{"id":1175634,"name":"Gábor Tax","orcid":"0000-0003-4530-0798","position":0,"is_corresponding":true}],"reference_count":71,"raw_metadata":null,"created_at":"2026-07-19T01:19:56.084411Z","pmid":"37398215","pmcid":null,"fwci":null,"citation_percentile":null,"influential_citations":0,"oa_status":null,"license":null,"views":0,"total_file_size_bytes":0,"version_count":0,"fair_f":null,"fair_a":null,"fair_i":null,"fair_r":null,"fair_zscore":null,"fair_rationale":null,"fair_model":null,"fair_agent_version":null,"fair_fulltext_source":null,"fair_has_llm":null,"fair_computed_at":null,"clinical_trials":[],"software_tools":[],"db_accessions":[],"linked_datasets":[],"topics":[]}