Depletion of <i>SNORA33</i> Abolishes ψ of 28S-U4966 and Affects the Ribosome Translational Apparatus
Eukaryotic ribosomes are complex molecular nanomachines translating genetic information from mRNAs into proteins. There is natural heterogeneity in ribosome composition. The pseudouridylation (ψ) of ribosomal RNAs (rRNAs) is one of the key sources of ribosome heterogeneity. Nevertheless, the functio...
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MDPI AG
2023-08-01
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| Series: | International Journal of Molecular Sciences |
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| Online Access: | https://www.mdpi.com/1422-0067/24/16/12578 |
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| author | Alzbeta Chabronova Guus van den Akker Bas A. C. Housmans Marjolein M. J. Caron Andy Cremers Don A. M. Surtel Mandy J. Peffers Lodewijk W. van Rhijn Virginie Marchand Yuri Motorin Tim J. M. Welting |
| author_facet | Alzbeta Chabronova Guus van den Akker Bas A. C. Housmans Marjolein M. J. Caron Andy Cremers Don A. M. Surtel Mandy J. Peffers Lodewijk W. van Rhijn Virginie Marchand Yuri Motorin Tim J. M. Welting |
| author_sort | Alzbeta Chabronova |
| collection | DOAJ |
| description | Eukaryotic ribosomes are complex molecular nanomachines translating genetic information from mRNAs into proteins. There is natural heterogeneity in ribosome composition. The pseudouridylation (ψ) of ribosomal RNAs (rRNAs) is one of the key sources of ribosome heterogeneity. Nevertheless, the functional consequences of ψ-based ribosome heterogeneity and its relevance for human disease are yet to be understood. Using HydraPsiSeq and a chronic disease model of non-osteoarthritic primary human articular chondrocytes exposed to osteoarthritic synovial fluid, we demonstrated that the disease microenvironment is capable of instigating site-specific changes in rRNA ψ profiles. To investigate one of the identified differential rRNA ψ sites (28S-ψ4966), we generated <i>SNORA22</i> and <i>SNORA33</i> KO SW1353 cell pools using LentiCRISPRv2/Cas9 and evaluated the ribosome translational capacity by <sup>35</sup>S-Met/Cys incorporation, assessed the mode of translation initiation and ribosomal fidelity using dual luciferase reporters, and assessed cellular and ribosomal proteomes by LC-MS/MS. We uncovered that the depletion of <i>SNORA33</i>, but not <i>SNORA22</i>, reduced 28S-ψ4966 levels. The resulting loss of 28S-ψ4966 affected ribosomal protein composition and function and led to specific changes in the cellular proteome. Overall, our pioneering findings demonstrate that cells dynamically respond to disease-relevant changes in their environment by altering their rRNA pseudouridylation profiles, with consequences for ribosome function and the cellular proteome relevant to human disease. |
| first_indexed | 2024-03-10T23:53:54Z |
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| issn | 1661-6596 1422-0067 |
| language | English |
| last_indexed | 2024-03-10T23:53:54Z |
| publishDate | 2023-08-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | International Journal of Molecular Sciences |
| spelling | doaj.art-f79530137c2145ee8eaffd0161a54d5d2023-11-19T01:25:21ZengMDPI AGInternational Journal of Molecular Sciences1661-65961422-00672023-08-0124161257810.3390/ijms241612578Depletion of <i>SNORA33</i> Abolishes ψ of 28S-U4966 and Affects the Ribosome Translational ApparatusAlzbeta Chabronova0Guus van den Akker1Bas A. C. Housmans2Marjolein M. J. Caron3Andy Cremers4Don A. M. Surtel5Mandy J. Peffers6Lodewijk W. van Rhijn7Virginie Marchand8Yuri Motorin9Tim J. M. Welting10Laboratory for Experimental Orthopedics, Department of Orthopedic Surgery, Maastricht University, 6229 HX Maastricht, The NetherlandsLaboratory for Experimental Orthopedics, Department of Orthopedic Surgery, Maastricht University, 6229 HX Maastricht, The NetherlandsLaboratory for Experimental Orthopedics, Department of Orthopedic Surgery, Maastricht University, 6229 HX Maastricht, The NetherlandsLaboratory for Experimental Orthopedics, Department of Orthopedic Surgery, Maastricht University, 6229 HX Maastricht, The NetherlandsLaboratory for Experimental Orthopedics, Department of Orthopedic Surgery, Maastricht University, 6229 HX Maastricht, The NetherlandsLaboratory for Experimental Orthopedics, Department of Orthopedic Surgery, Maastricht University, 6229 HX Maastricht, The NetherlandsInstitute of Life Course and Medical Sciences, University of Liverpool, Liverpool L8 7TX, UKLaboratory for Experimental Orthopedics, Department of Orthopedic Surgery, Maastricht University, 6229 HX Maastricht, The NetherlandsUAR2008 IBSLor CNRS-INSERM-Université de Lorraine, F54000 Nancy, FranceUAR2008 IBSLor CNRS-INSERM-Université de Lorraine, F54000 Nancy, FranceLaboratory for Experimental Orthopedics, Department of Orthopedic Surgery, Maastricht University, 6229 HX Maastricht, The NetherlandsEukaryotic ribosomes are complex molecular nanomachines translating genetic information from mRNAs into proteins. There is natural heterogeneity in ribosome composition. The pseudouridylation (ψ) of ribosomal RNAs (rRNAs) is one of the key sources of ribosome heterogeneity. Nevertheless, the functional consequences of ψ-based ribosome heterogeneity and its relevance for human disease are yet to be understood. Using HydraPsiSeq and a chronic disease model of non-osteoarthritic primary human articular chondrocytes exposed to osteoarthritic synovial fluid, we demonstrated that the disease microenvironment is capable of instigating site-specific changes in rRNA ψ profiles. To investigate one of the identified differential rRNA ψ sites (28S-ψ4966), we generated <i>SNORA22</i> and <i>SNORA33</i> KO SW1353 cell pools using LentiCRISPRv2/Cas9 and evaluated the ribosome translational capacity by <sup>35</sup>S-Met/Cys incorporation, assessed the mode of translation initiation and ribosomal fidelity using dual luciferase reporters, and assessed cellular and ribosomal proteomes by LC-MS/MS. We uncovered that the depletion of <i>SNORA33</i>, but not <i>SNORA22</i>, reduced 28S-ψ4966 levels. The resulting loss of 28S-ψ4966 affected ribosomal protein composition and function and led to specific changes in the cellular proteome. Overall, our pioneering findings demonstrate that cells dynamically respond to disease-relevant changes in their environment by altering their rRNA pseudouridylation profiles, with consequences for ribosome function and the cellular proteome relevant to human disease.https://www.mdpi.com/1422-0067/24/16/12578ribosomal RNA28Sepitranscriptomeribosomechondrocytesosteoarthritis |
| spellingShingle | Alzbeta Chabronova Guus van den Akker Bas A. C. Housmans Marjolein M. J. Caron Andy Cremers Don A. M. Surtel Mandy J. Peffers Lodewijk W. van Rhijn Virginie Marchand Yuri Motorin Tim J. M. Welting Depletion of <i>SNORA33</i> Abolishes ψ of 28S-U4966 and Affects the Ribosome Translational Apparatus International Journal of Molecular Sciences ribosomal RNA 28S epitranscriptome ribosome chondrocytes osteoarthritis |
| title | Depletion of <i>SNORA33</i> Abolishes ψ of 28S-U4966 and Affects the Ribosome Translational Apparatus |
| title_full | Depletion of <i>SNORA33</i> Abolishes ψ of 28S-U4966 and Affects the Ribosome Translational Apparatus |
| title_fullStr | Depletion of <i>SNORA33</i> Abolishes ψ of 28S-U4966 and Affects the Ribosome Translational Apparatus |
| title_full_unstemmed | Depletion of <i>SNORA33</i> Abolishes ψ of 28S-U4966 and Affects the Ribosome Translational Apparatus |
| title_short | Depletion of <i>SNORA33</i> Abolishes ψ of 28S-U4966 and Affects the Ribosome Translational Apparatus |
| title_sort | depletion of i snora33 i abolishes ψ of 28s u4966 and affects the ribosome translational apparatus |
| topic | ribosomal RNA 28S epitranscriptome ribosome chondrocytes osteoarthritis |
| url | https://www.mdpi.com/1422-0067/24/16/12578 |
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