{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"institution":[{"name":"bioRxiv"}],"indexed":{"date-parts":[[2026,1,15]],"date-time":"2026-01-15T22:41:30Z","timestamp":1768516890516,"version":"3.49.0"},"posted":{"date-parts":[[2024,9,21]]},"group-title":"Biochemistry","reference-count":51,"publisher":"openRxiv","license":[{"start":{"date-parts":[[2024,9,21]],"date-time":"2024-09-21T00:00:00Z","timestamp":1726876800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/www.biorxiv.org\/about\/FAQ#license"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"accepted":{"date-parts":[[2024,9,21]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                <jats:p>\n                  Eukaryotic ribosomes are enriched with pseudouridine, particularly at the functional centers targeted by antibiotics. Here we investigated the roles of pseudouridine in aminoglycoside-mediated translation inhibition by comparing the structural and functional properties of the wild-type ribosomes and those lacking pseudouridine (\n                  <jats:italic>cbf5<\/jats:italic>\n                  -D95A). We showed that the\n                  <jats:italic>cbf5<\/jats:italic>\n                  -D95A ribosomes have decreased thermostability and high sensitivity to aminoglycosides. When presented with an internal ribosome entry site (IRES) RNA, elongation factor eEF2, GTP, sordarin, hygromycin B preferentially binds to the\n                  <jats:italic>cbf5<\/jats:italic>\n                  -D95A ribosomes during initiation by blocking eEF2 binding and stalls the ribosomes in a non-rotated conformation, further hindering translocation. Hygromycin B binds to the inter-subunit bridge B2a that is known to be sensitive to pseudouridine, revealing a functional link between pseudouridine and aminoglycoside inhibition. Our results suggest that pseudouridine enhances both thermostability and conformational fitness of the ribosomes, thereby influencing their susceptibility to aminoglycosides.\n                <\/jats:p>\n                <jats:sec>\n                  <jats:title>Highlights<\/jats:title>\n                  <jats:list list-type=\"bullet\">\n                    <jats:list-item>\n                      <jats:p>Loss of pseudouridine increases cell sensitivity to aminoglycosides<\/jats:p>\n                    <\/jats:list-item>\n                    <jats:list-item>\n                      <jats:p>Pseudouridine enhances ribosome thermostability<\/jats:p>\n                    <\/jats:list-item>\n                    <jats:list-item>\n                      <jats:p>Hygromycin B competes with eEF2 for the non-rotated ribosome<\/jats:p>\n                    <\/jats:list-item>\n                    <jats:list-item>\n                      <jats:p>Hygromycin B deforms the codon-anticodon duplex<\/jats:p>\n                    <\/jats:list-item>\n                  <\/jats:list>\n                <\/jats:sec>","DOI":"10.1101\/2024.09.20.614200","type":"posted-content","created":{"date-parts":[[2024,9,22]],"date-time":"2024-09-22T02:15:15Z","timestamp":1726971315000},"source":"Crossref","is-referenced-by-count":0,"title":["Unveil the Molecular Interplay between Aminoglycosides and Pseudouridine in IRES Translation"],"prefix":"10.64898","author":[{"ORCID":"https:\/\/orcid.org\/0000-0003-0485-2780","authenticated-orcid":false,"given":"Yu","family":"Zhao","sequence":"first","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Chong","family":"Xu","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Xin","family":"Chen","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Hong","family":"Jin","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-2046-9861","authenticated-orcid":false,"given":"Hong","family":"Li","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"54368","reference":[{"key":"2024092511351040000_2024.09.20.614200v1.1","doi-asserted-by":"crossref","first-page":"1138","DOI":"10.1080\/15476286.2016.1259781","article-title":"Tuning the ribosome: The influence of rRNA modification on eukaryotic ribosome biogenesis and function","volume":"14","year":"2017","journal-title":"RNA biology"},{"key":"2024092511351040000_2024.09.20.614200v1.2","doi-asserted-by":"publisher","DOI":"10.1016\/S1097-2765(03)00040-6"},{"key":"2024092511351040000_2024.09.20.614200v1.3","doi-asserted-by":"publisher","DOI":"10.1016\/j.molcel.2007.10.012"},{"key":"2024092511351040000_2024.09.20.614200v1.4","doi-asserted-by":"crossref","first-page":"eadg8190","DOI":"10.1126\/sciadv.adg8190","article-title":"Regulation of translation by ribosomal RNA pseudouridylation","volume":"9","year":"2023","journal-title":"Sci Adv"},{"key":"2024092511351040000_2024.09.20.614200v1.5","doi-asserted-by":"publisher","DOI":"10.1016\/j.str.2022.04.002"},{"key":"2024092511351040000_2024.09.20.614200v1.6","doi-asserted-by":"publisher","DOI":"10.1038\/nature24482"},{"key":"2024092511351040000_2024.09.20.614200v1.7","doi-asserted-by":"publisher","DOI":"10.1038\/nsmb.2992"},{"key":"2024092511351040000_2024.09.20.614200v1.8","doi-asserted-by":"publisher","DOI":"10.1038\/nsmb.2994"},{"key":"2024092511351040000_2024.09.20.614200v1.9","doi-asserted-by":"publisher","DOI":"10.1126\/science.abf3546"},{"key":"2024092511351040000_2024.09.20.614200v1.10","doi-asserted-by":"publisher","DOI":"10.1261\/rna.2320605"},{"key":"2024092511351040000_2024.09.20.614200v1.11","doi-asserted-by":"publisher","DOI":"10.1093\/nar\/gkt1330"},{"key":"2024092511351040000_2024.09.20.614200v1.12","doi-asserted-by":"publisher","DOI":"10.1093\/nar\/23.24.5020"},{"key":"2024092511351040000_2024.09.20.614200v1.13","doi-asserted-by":"crossref","unstructured":"M. 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