Ribosomal RNA-based epitranscriptomic regulation of chondrocyte translation and proteome in osteoarthritis

Alzbeta Chabronova; Guus G H van den Akker; Bas A C Housmans; Marjolein M J Caron; Andy Cremers; Don A M Surtel; Kanin Wichapong; Mandy M J Peffers; Lodewijk van Rhijn; Virginie Marchand; Yuri Motorin; Tim J M Welting

doi:10.1016/j.joca.2022.12.010

Ribosomal RNA-based epitranscriptomic regulation of chondrocyte translation and proteome in osteoarthritis

Alzbeta Chabronova, Guus G H van den Akker, Bas A C Housmans, Marjolein M J Caron, Andy Cremers, Don A M Surtel, Kanin Wichapong, Mandy M J Peffers, van Rhijn, Virginie Marchand, Yuri Motorin, Tim J M Welting

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Abstract

OBJECTIVE: Osteoarthritis-related cartilage extracellular matrix remodeling is dependent on changes in chondrocyte protein expression. Yet, the role of ribosomes in chondrocyte translation regulation is unknown. In this exploratory study, we investigated ribosomal RNA (rRNA) epitranscriptomic-based ribosome heterogeneity in human articular chondrocytes and its relevance for osteoarthritis.

METHODS: Sequencing-based rRNA 2'-O-methylation profiling analysis (RiboMethSeq) was performed on non-OA primary human articular chondrocytes (n = 5) exposed for 14 days to osteoarthritic synovial fluid (14 donors, pooled, 20% v/v). The SW1353 SNORD71 KO cell pool was generated using LentiCRISPRv2/Cas9. The mode of translation initiation and fidelity were determined by dual-luciferase reporters. The cellular proteome was analyzed by LC-MS/MS and collagen type I protein expression was evaluated by immunoblotting. Loading of COL1A1 mRNA into polysomes was determined by sucrose gradient ultracentrifugation and fractionation.

RESULTS: We discovered that osteoarthritic synovial fluid instigates site-specific changes in the rRNA 2'-O-me profile of primary human articular chondrocytes. We identified five sites with differential 2'-O-me levels. The 2'-O-me status of 5.8S-U14 (one of identified differential 2'-O-me sites; decreased by 7.7%, 95% CI [0.9-14.5%]) was targeted by depleting the level of its guide snoRNA SNORD71 (50% decrease, 95% CI [33-64%]). This resulted in an altered ribosome translation modus (e.g., CrPV IRES, FC 3, 95% CI [2.2-4.1]) and promoted translation of COL1A1 mRNA which led to increased levels of COL1A1 protein (FC 1.7, 95% CI [1.3-2.0]).

CONCLUSIONS: Our data identify a novel concept suggesting that articular chondrocytes employ rRNA epitranscriptomic mechanisms in osteoarthritis development.

Original language	English
Pages (from-to)	374-385
Number of pages	12
Journal	Osteoarthritis and Cartilage
Volume	31
Issue number	3
Early online date	5 Jan 2023
DOIs	https://doi.org/10.1016/j.joca.2022.12.010
Publication status	Published - Mar 2023

Keywords

5.8S rRNA
Chondrocytes
Collagen type I
Preferential translation
Ribosome
rRNA epitranscriptomic regulation

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Chabronova, A., van den Akker, G. G. H., Housmans, B. A. C., Caron, M. M. J., Cremers, A., Surtel, D. A. M., Wichapong, K., Peffers, M. M. J., van Rhijn, Marchand, V., Motorin, Y., & Welting, T. J. M. (2023). Ribosomal RNA-based epitranscriptomic regulation of chondrocyte translation and proteome in osteoarthritis. Osteoarthritis and Cartilage, 31(3), 374-385. https://doi.org/10.1016/j.joca.2022.12.010

@article{934d20b01d6542eab3e79c93ec6a907e,

title = "Ribosomal RNA-based epitranscriptomic regulation of chondrocyte translation and proteome in osteoarthritis",

abstract = "OBJECTIVE: Osteoarthritis-related cartilage extracellular matrix remodeling is dependent on changes in chondrocyte protein expression. Yet, the role of ribosomes in chondrocyte translation regulation is unknown. In this exploratory study, we investigated ribosomal RNA (rRNA) epitranscriptomic-based ribosome heterogeneity in human articular chondrocytes and its relevance for osteoarthritis.METHODS: Sequencing-based rRNA 2'-O-methylation profiling analysis (RiboMethSeq) was performed on non-OA primary human articular chondrocytes (n = 5) exposed for 14 days to osteoarthritic synovial fluid (14 donors, pooled, 20% v/v). The SW1353 SNORD71 KO cell pool was generated using LentiCRISPRv2/Cas9. The mode of translation initiation and fidelity were determined by dual-luciferase reporters. The cellular proteome was analyzed by LC-MS/MS and collagen type I protein expression was evaluated by immunoblotting. Loading of COL1A1 mRNA into polysomes was determined by sucrose gradient ultracentrifugation and fractionation.RESULTS: We discovered that osteoarthritic synovial fluid instigates site-specific changes in the rRNA 2'-O-me profile of primary human articular chondrocytes. We identified five sites with differential 2'-O-me levels. The 2'-O-me status of 5.8S-U14 (one of identified differential 2'-O-me sites; decreased by 7.7%, 95% CI [0.9-14.5%]) was targeted by depleting the level of its guide snoRNA SNORD71 (50% decrease, 95% CI [33-64%]). This resulted in an altered ribosome translation modus (e.g., CrPV IRES, FC 3, 95% CI [2.2-4.1]) and promoted translation of COL1A1 mRNA which led to increased levels of COL1A1 protein (FC 1.7, 95% CI [1.3-2.0]).CONCLUSIONS: Our data identify a novel concept suggesting that articular chondrocytes employ rRNA epitranscriptomic mechanisms in osteoarthritis development.",

keywords = "5.8S rRNA, Chondrocytes, Collagen type I, Preferential translation, Ribosome, rRNA epitranscriptomic regulation",

author = "Alzbeta Chabronova and {van den Akker}, {Guus G H} and Housmans, {Bas A C} and Caron, {Marjolein M J} and Andy Cremers and Surtel, {Don A M} and Kanin Wichapong and Peffers, {Mandy M J} and {van Rhijn} and Virginie Marchand and Yuri Motorin and Welting, {Tim J M}",

note = "Funding Information: This work was supported by a grant from Stichting de Weijerhorst (Bewegen zonder Pijn), and grants from the Dutch Arthritis Foundation (grants 17-2-401 and LLP14 ). Mandy Peffers is funded through a Wellcome Trust Clinical Intermediate Fellowship (grant 107471/Z/15/Z ) and supported by Versus Arthritis as part of the MRC Versus Arthritis Centre for Integrated research into Musculoskeletal Ageing (CIMA). Publisher Copyright: {\textcopyright} 2023 The Authors",

year = "2023",

month = mar,

doi = "10.1016/j.joca.2022.12.010",

language = "English",

volume = "31",

pages = "374--385",

journal = "Osteoarthritis and Cartilage",

issn = "1063-4584",

publisher = "W.B. Saunders Ltd",

number = "3",

}

Chabronova, A, van den Akker, GGH, Housmans, BAC, Caron, MMJ, Cremers, A, Surtel, DAM, Wichapong, K, Peffers, MMJ, van Rhijn, Marchand, V, Motorin, Y & Welting, TJM 2023, 'Ribosomal RNA-based epitranscriptomic regulation of chondrocyte translation and proteome in osteoarthritis', Osteoarthritis and Cartilage, vol. 31, no. 3, pp. 374-385. https://doi.org/10.1016/j.joca.2022.12.010

TY - JOUR

T1 - Ribosomal RNA-based epitranscriptomic regulation of chondrocyte translation and proteome in osteoarthritis

AU - Chabronova, Alzbeta

AU - van den Akker, Guus G H

AU - Housmans, Bas A C

AU - Caron, Marjolein M J

AU - Cremers, Andy

AU - Surtel, Don A M

AU - Wichapong, Kanin

AU - Peffers, Mandy M J

AU - van Rhijn, null

AU - Marchand, Virginie

AU - Motorin, Yuri

AU - Welting, Tim J M

N1 - Funding Information: This work was supported by a grant from Stichting de Weijerhorst (Bewegen zonder Pijn), and grants from the Dutch Arthritis Foundation (grants 17-2-401 and LLP14 ). Mandy Peffers is funded through a Wellcome Trust Clinical Intermediate Fellowship (grant 107471/Z/15/Z ) and supported by Versus Arthritis as part of the MRC Versus Arthritis Centre for Integrated research into Musculoskeletal Ageing (CIMA). Publisher Copyright: © 2023 The Authors

PY - 2023/3

Y1 - 2023/3

N2 - OBJECTIVE: Osteoarthritis-related cartilage extracellular matrix remodeling is dependent on changes in chondrocyte protein expression. Yet, the role of ribosomes in chondrocyte translation regulation is unknown. In this exploratory study, we investigated ribosomal RNA (rRNA) epitranscriptomic-based ribosome heterogeneity in human articular chondrocytes and its relevance for osteoarthritis.METHODS: Sequencing-based rRNA 2'-O-methylation profiling analysis (RiboMethSeq) was performed on non-OA primary human articular chondrocytes (n = 5) exposed for 14 days to osteoarthritic synovial fluid (14 donors, pooled, 20% v/v). The SW1353 SNORD71 KO cell pool was generated using LentiCRISPRv2/Cas9. The mode of translation initiation and fidelity were determined by dual-luciferase reporters. The cellular proteome was analyzed by LC-MS/MS and collagen type I protein expression was evaluated by immunoblotting. Loading of COL1A1 mRNA into polysomes was determined by sucrose gradient ultracentrifugation and fractionation.RESULTS: We discovered that osteoarthritic synovial fluid instigates site-specific changes in the rRNA 2'-O-me profile of primary human articular chondrocytes. We identified five sites with differential 2'-O-me levels. The 2'-O-me status of 5.8S-U14 (one of identified differential 2'-O-me sites; decreased by 7.7%, 95% CI [0.9-14.5%]) was targeted by depleting the level of its guide snoRNA SNORD71 (50% decrease, 95% CI [33-64%]). This resulted in an altered ribosome translation modus (e.g., CrPV IRES, FC 3, 95% CI [2.2-4.1]) and promoted translation of COL1A1 mRNA which led to increased levels of COL1A1 protein (FC 1.7, 95% CI [1.3-2.0]).CONCLUSIONS: Our data identify a novel concept suggesting that articular chondrocytes employ rRNA epitranscriptomic mechanisms in osteoarthritis development.

AB - OBJECTIVE: Osteoarthritis-related cartilage extracellular matrix remodeling is dependent on changes in chondrocyte protein expression. Yet, the role of ribosomes in chondrocyte translation regulation is unknown. In this exploratory study, we investigated ribosomal RNA (rRNA) epitranscriptomic-based ribosome heterogeneity in human articular chondrocytes and its relevance for osteoarthritis.METHODS: Sequencing-based rRNA 2'-O-methylation profiling analysis (RiboMethSeq) was performed on non-OA primary human articular chondrocytes (n = 5) exposed for 14 days to osteoarthritic synovial fluid (14 donors, pooled, 20% v/v). The SW1353 SNORD71 KO cell pool was generated using LentiCRISPRv2/Cas9. The mode of translation initiation and fidelity were determined by dual-luciferase reporters. The cellular proteome was analyzed by LC-MS/MS and collagen type I protein expression was evaluated by immunoblotting. Loading of COL1A1 mRNA into polysomes was determined by sucrose gradient ultracentrifugation and fractionation.RESULTS: We discovered that osteoarthritic synovial fluid instigates site-specific changes in the rRNA 2'-O-me profile of primary human articular chondrocytes. We identified five sites with differential 2'-O-me levels. The 2'-O-me status of 5.8S-U14 (one of identified differential 2'-O-me sites; decreased by 7.7%, 95% CI [0.9-14.5%]) was targeted by depleting the level of its guide snoRNA SNORD71 (50% decrease, 95% CI [33-64%]). This resulted in an altered ribosome translation modus (e.g., CrPV IRES, FC 3, 95% CI [2.2-4.1]) and promoted translation of COL1A1 mRNA which led to increased levels of COL1A1 protein (FC 1.7, 95% CI [1.3-2.0]).CONCLUSIONS: Our data identify a novel concept suggesting that articular chondrocytes employ rRNA epitranscriptomic mechanisms in osteoarthritis development.

KW - 5.8S rRNA

KW - Chondrocytes

KW - Collagen type I

KW - Preferential translation

KW - Ribosome

KW - rRNA epitranscriptomic regulation

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U2 - 10.1016/j.joca.2022.12.010

DO - 10.1016/j.joca.2022.12.010

M3 - Article

C2 - 36621590

SN - 1063-4584

VL - 31

SP - 374

EP - 385

JO - Osteoarthritis and Cartilage

JF - Osteoarthritis and Cartilage

IS - 3

ER -

Ribosomal RNA-based epitranscriptomic regulation of chondrocyte translation and proteome in osteoarthritis

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