N 6-methyladenosine in poly(A) tails stabilize VSG transcripts

Idálio J. Viegas; Juan Pereira de Macedo; Lúcia Serra; Mariana De Niz; Adriana Temporão; Sara Silva Pereira; Aashiq H. Mirza; Ed Bergstrom; João A. Rodrigues; Francisco Aresta-Branco; Samie R. Jaffrey; Luisa M. Figueiredo

doi:10.1038/s41586-022-04544-0

N ⁶-methyladenosine in poly(A) tails stabilize VSG transcripts

Idálio J. Viegas, Juan Pereira de Macedo, Lúcia Serra, Mariana De Niz, Adriana Temporão, Sara Silva Pereira, Aashiq H. Mirza, Ed Bergstrom, João A. Rodrigues, Francisco Aresta-Branco, Samie R. Jaffrey, Luisa M. Figueiredo^*

^*Corresponding author for this work

Cell & Developmental Biology

Research output: Contribution to journal › Article › peer-review

42 Scopus citations

Abstract

RNA modifications are important regulators of gene expression¹. In Trypanosoma brucei, transcription is polycistronic and thus most regulation happens post-transcriptionally². N⁶-methyladenosine (m⁶A) has been detected in this parasite, but its function remains unknown³. Here we found that m⁶A is enriched in 342 transcripts using RNA immunoprecipitation, with an enrichment in transcripts encoding variant surface glycoproteins (VSGs). Approximately 50% of the m⁶A is located in the poly(A) tail of the actively expressed VSG transcripts. m⁶A residues are removed from the VSG poly(A) tail before deadenylation and mRNA degradation. Computational analysis revealed an association between m⁶A in the poly(A) tail and a 16-mer motif in the 3′ untranslated region of VSG genes. Using genetic tools, we show that the 16-mer motif acts as a cis-acting motif that is required for inclusion of m⁶A in the poly(A) tail. Removal of this motif from the 3′ untranslated region of VSG genes results in poly(A) tails lacking m⁶A, rapid deadenylation and mRNA degradation. To our knowledge, this is the first identification of an RNA modification in the poly(A) tail of any eukaryote, uncovering a post-transcriptional mechanism of gene regulation.

Original language	English (US)
Pages (from-to)	362-370
Number of pages	9
Journal	Nature
Volume	604
Issue number	7905
DOIs	https://doi.org/10.1038/s41586-022-04544-0
State	Published - Apr 14 2022

Funding

We are grateful to support from the Howard Hughes Medical Institute International Early Career Scientist Program (55007419), a European Molecular Biology Organization Installation grant (2151) and La Caixa Foundation (HR20-00361). This work was also partially supported by the ONEIDA project (LISBOA-01-0145-FEDER-016417) co-funded by Fundos Europeus Estruturais e de Investimento (FEEI) from ?Programa Operacional Regional Lisboa 2020? and by national funds from Funda??o para a Ci?ncia e a Tecnologia (FCT).?S.R.J. was supported by NIH (R35 NS111631). Researchers were funded by individual fellowships from FCT (PD/BD/105838/2014 to I.J.V., 2020.06827.BD to L.S., SFRH/BD/80718/2011 to F.A.-B.,?PD/BD/138891/2018 to A.T. and CEECIND/03322/2018 to L.M.F.); a Novartis Foundation for Biomedical-Biological research to J.P.d.M.; a Human Frontier Science Programme long-term postdoctoral fellowship to M.D.N. (LT000047/2019); a Marie Sk?odowska-Curie Individual Standard European Fellowship to S.S.P. (grant no. 839960); the GlycoPar Marie Curie Initial Training Network (GA 608295) to J.A.R. We thank J. Thomas-Oates (University of York, Centre of Excellence in Mass Spectrometry, Department of Chemistry) for the mass spectrometry analysis. The York Centre of Excellence in Mass Spectrometry was created thanks to a major capital investment through Science City York, supported by Yorkshire Forward with funds from the Northern Way Initiative, and subsequent support from EPSRC (EP/K039660/1 and EP/M028127/1). We also thank A. Temudo, A. Nascimento and A. Lima for bioimaging assistance; the laboratories of A. Tom?s and J. Kelly for providing RNA from Leishmania and T. cruzi , respectively; and A. Pena and members of the Figueiredo and Jaffrey laboratories for helpful discussions. We are grateful to support from the Howard Hughes Medical Institute International Early Career Scientist Program (55007419), a European Molecular Biology Organization Installation grant (2151) and La Caixa Foundation (HR20-00361). This work was also partially supported by the ONEIDA project (LISBOA-01-0145-FEDER-016417) co-funded by Fundos Europeus Estruturais e de Investimento (FEEI) from \u2018Programa Operacional Regional Lisboa 2020\u2019 and by national funds from Funda\u00E7\u00E3o para a Ci\u00EAncia e a Tecnologia (FCT). S.R.J. was supported by NIH (R35 NS111631). Researchers were funded by individual fellowships from FCT (PD/BD/105838/2014 to I.J.V., 2020.06827.BD to L.S., SFRH/BD/80718/2011 to F.A.-B., PD/BD/138891/2018 to A.T. and CEECIND/03322/2018 to L.M.F.); a Novartis Foundation for Biomedical-Biological research to J.P.d.M.; a Human Frontier Science Programme long-term postdoctoral fellowship to M.D.N. (LT000047/2019); a Marie Sk\u0142odowska-Curie Individual Standard European Fellowship to S.S.P. (grant no. 839960); the GlycoPar Marie Curie Initial Training Network (GA 608295) to J.A.R. We thank J. Thomas-Oates (University of York, Centre of Excellence in Mass Spectrometry, Department of Chemistry) for the mass spectrometry analysis. The York Centre of Excellence in Mass Spectrometry was created thanks to a major capital investment through Science City York, supported by Yorkshire Forward with funds from the Northern Way Initiative, and subsequent support from EPSRC (EP/K039660/1 and EP/M028127/1). We also thank A. Temudo, A. Nascimento and A. Lima for bioimaging assistance; the laboratories of A. Tom\u00E1s and J. Kelly for providing RNA from Leishmania and T. cruzi, respectively; and A. Pena and members of the Figueiredo and Jaffrey laboratories for helpful discussions.

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10.1038/s41586-022-04544-0

Cite this

@article{cbe8b23601b744b98603c4e875071bae,

title = "N 6-methyladenosine in poly(A) tails stabilize VSG transcripts",

abstract = "RNA modifications are important regulators of gene expression1. In Trypanosoma brucei, transcription is polycistronic and thus most regulation happens post-transcriptionally2. N6-methyladenosine (m6A) has been detected in this parasite, but its function remains unknown3. Here we found that m6A is enriched in 342 transcripts using RNA immunoprecipitation, with an enrichment in transcripts encoding variant surface glycoproteins (VSGs). Approximately 50% of the m6A is located in the poly(A) tail of the actively expressed VSG transcripts. m6A residues are removed from the VSG poly(A) tail before deadenylation and mRNA degradation. Computational analysis revealed an association between m6A in the poly(A) tail and a 16-mer motif in the 3′ untranslated region of VSG genes. Using genetic tools, we show that the 16-mer motif acts as a cis-acting motif that is required for inclusion of m6A in the poly(A) tail. Removal of this motif from the 3′ untranslated region of VSG genes results in poly(A) tails lacking m6A, rapid deadenylation and mRNA degradation. To our knowledge, this is the first identification of an RNA modification in the poly(A) tail of any eukaryote, uncovering a post-transcriptional mechanism of gene regulation.",

author = "Viegas, {Id{\'a}lio J.} and {de Macedo}, {Juan Pereira} and L{\'u}cia Serra and {De Niz}, Mariana and Adriana Tempor{\~a}o and {Silva Pereira}, Sara and Mirza, {Aashiq H.} and Ed Bergstrom and Rodrigues, {Jo{\~a}o A.} and Francisco Aresta-Branco and Jaffrey, {Samie R.} and Figueiredo, {Luisa M.}",

note = "Funding Information: We are grateful to support from the Howard Hughes Medical Institute International Early Career Scientist Program (55007419), a European Molecular Biology Organization Installation grant (2151) and La Caixa Foundation (HR20-00361). This work was also partially supported by the ONEIDA project (LISBOA-01-0145-FEDER-016417) co-funded by Fundos Europeus Estruturais e de Investimento (FEEI) from {\textquoteleft}Programa Operacional Regional Lisboa 2020{\textquoteright} and by national funds from Funda{\c c}{\~a}o para a Ci{\^e}ncia e a Tecnologia (FCT). S.R.J. was supported by NIH (R35 NS111631). Researchers were funded by individual fellowships from FCT (PD/BD/105838/2014 to I.J.V., 2020.06827.BD to L.S., SFRH/BD/80718/2011 to F.A.-B., PD/BD/138891/2018 to A.T. and CEECIND/03322/2018 to L.M.F.); a Novartis Foundation for Biomedical-Biological research to J.P.d.M.; a Human Frontier Science Programme long-term postdoctoral fellowship to M.D.N. (LT000047/2019); a Marie Sk{\l}odowska-Curie Individual Standard European Fellowship to S.S.P. (grant no. 839960); the GlycoPar Marie Curie Initial Training Network (GA 608295) to J.A.R. We thank J. Thomas-Oates (University of York, Centre of Excellence in Mass Spectrometry, Department of Chemistry) for the mass spectrometry analysis. The York Centre of Excellence in Mass Spectrometry was created thanks to a major capital investment through Science City York, supported by Yorkshire Forward with funds from the Northern Way Initiative, and subsequent support from EPSRC (EP/K039660/1 and EP/M028127/1). We also thank A. Temudo, A. Nascimento and A. Lima for bioimaging assistance; the laboratories of A. Tom{\'a}s and J. Kelly for providing RNA from Leishmania and T. cruzi, respectively; and A. Pena and members of the Figueiredo and Jaffrey laboratories for helpful discussions. Funding Information: We are grateful to support from the Howard Hughes Medical Institute International Early Career Scientist Program (55007419), a European Molecular Biology Organization Installation grant (2151) and La Caixa Foundation (HR20-00361). This work was also partially supported by the ONEIDA project (LISBOA-01-0145-FEDER-016417) co-funded by Fundos Europeus Estruturais e de Investimento (FEEI) from ?Programa Operacional Regional Lisboa 2020? and by national funds from Funda??o para a Ci?ncia e a Tecnologia (FCT).?S.R.J. was supported by NIH (R35 NS111631). Researchers were funded by individual fellowships from FCT (PD/BD/105838/2014 to I.J.V., 2020.06827.BD to L.S., SFRH/BD/80718/2011 to F.A.-B.,?PD/BD/138891/2018 to A.T. and CEECIND/03322/2018 to L.M.F.); a Novartis Foundation for Biomedical-Biological research to J.P.d.M.; a Human Frontier Science Programme long-term postdoctoral fellowship to M.D.N. (LT000047/2019); a Marie Sk?odowska-Curie Individual Standard European Fellowship to S.S.P. (grant no. 839960); the GlycoPar Marie Curie Initial Training Network (GA 608295) to J.A.R. We thank J. Thomas-Oates (University of York, Centre of Excellence in Mass Spectrometry, Department of Chemistry) for the mass spectrometry analysis. The York Centre of Excellence in Mass Spectrometry was created thanks to a major capital investment through Science City York, supported by Yorkshire Forward with funds from the Northern Way Initiative, and subsequent support from EPSRC (EP/K039660/1 and EP/M028127/1). We also thank A. Temudo, A. Nascimento and A. Lima for bioimaging assistance; the laboratories of A. Tom?s and J. Kelly for providing RNA from Leishmania and T. cruzi , respectively; and A. Pena and members of the Figueiredo and Jaffrey laboratories for helpful discussions. Publisher Copyright: {\textcopyright} 2022, The Author(s), under exclusive licence to Springer Nature Limited.",

year = "2022",

month = apr,

day = "14",

doi = "10.1038/s41586-022-04544-0",

language = "English (US)",

volume = "604",

pages = "362--370",

journal = "Nature",

issn = "0028-0836",

publisher = "Nature Research",

number = "7905",

}

TY - JOUR

T1 - N 6-methyladenosine in poly(A) tails stabilize VSG transcripts

AU - Viegas, Idálio J.

AU - de Macedo, Juan Pereira

AU - Serra, Lúcia

AU - De Niz, Mariana

AU - Temporão, Adriana

AU - Silva Pereira, Sara

AU - Mirza, Aashiq H.

AU - Bergstrom, Ed

AU - Rodrigues, João A.

AU - Aresta-Branco, Francisco

AU - Jaffrey, Samie R.

AU - Figueiredo, Luisa M.

N1 - Funding Information: We are grateful to support from the Howard Hughes Medical Institute International Early Career Scientist Program (55007419), a European Molecular Biology Organization Installation grant (2151) and La Caixa Foundation (HR20-00361). This work was also partially supported by the ONEIDA project (LISBOA-01-0145-FEDER-016417) co-funded by Fundos Europeus Estruturais e de Investimento (FEEI) from ‘Programa Operacional Regional Lisboa 2020’ and by national funds from Fundação para a Ciência e a Tecnologia (FCT). S.R.J. was supported by NIH (R35 NS111631). Researchers were funded by individual fellowships from FCT (PD/BD/105838/2014 to I.J.V., 2020.06827.BD to L.S., SFRH/BD/80718/2011 to F.A.-B., PD/BD/138891/2018 to A.T. and CEECIND/03322/2018 to L.M.F.); a Novartis Foundation for Biomedical-Biological research to J.P.d.M.; a Human Frontier Science Programme long-term postdoctoral fellowship to M.D.N. (LT000047/2019); a Marie Skłodowska-Curie Individual Standard European Fellowship to S.S.P. (grant no. 839960); the GlycoPar Marie Curie Initial Training Network (GA 608295) to J.A.R. We thank J. Thomas-Oates (University of York, Centre of Excellence in Mass Spectrometry, Department of Chemistry) for the mass spectrometry analysis. The York Centre of Excellence in Mass Spectrometry was created thanks to a major capital investment through Science City York, supported by Yorkshire Forward with funds from the Northern Way Initiative, and subsequent support from EPSRC (EP/K039660/1 and EP/M028127/1). We also thank A. Temudo, A. Nascimento and A. Lima for bioimaging assistance; the laboratories of A. Tomás and J. Kelly for providing RNA from Leishmania and T. cruzi, respectively; and A. Pena and members of the Figueiredo and Jaffrey laboratories for helpful discussions. Funding Information: We are grateful to support from the Howard Hughes Medical Institute International Early Career Scientist Program (55007419), a European Molecular Biology Organization Installation grant (2151) and La Caixa Foundation (HR20-00361). This work was also partially supported by the ONEIDA project (LISBOA-01-0145-FEDER-016417) co-funded by Fundos Europeus Estruturais e de Investimento (FEEI) from ?Programa Operacional Regional Lisboa 2020? and by national funds from Funda??o para a Ci?ncia e a Tecnologia (FCT).?S.R.J. was supported by NIH (R35 NS111631). Researchers were funded by individual fellowships from FCT (PD/BD/105838/2014 to I.J.V., 2020.06827.BD to L.S., SFRH/BD/80718/2011 to F.A.-B.,?PD/BD/138891/2018 to A.T. and CEECIND/03322/2018 to L.M.F.); a Novartis Foundation for Biomedical-Biological research to J.P.d.M.; a Human Frontier Science Programme long-term postdoctoral fellowship to M.D.N. (LT000047/2019); a Marie Sk?odowska-Curie Individual Standard European Fellowship to S.S.P. (grant no. 839960); the GlycoPar Marie Curie Initial Training Network (GA 608295) to J.A.R. We thank J. Thomas-Oates (University of York, Centre of Excellence in Mass Spectrometry, Department of Chemistry) for the mass spectrometry analysis. The York Centre of Excellence in Mass Spectrometry was created thanks to a major capital investment through Science City York, supported by Yorkshire Forward with funds from the Northern Way Initiative, and subsequent support from EPSRC (EP/K039660/1 and EP/M028127/1). We also thank A. Temudo, A. Nascimento and A. Lima for bioimaging assistance; the laboratories of A. Tom?s and J. Kelly for providing RNA from Leishmania and T. cruzi , respectively; and A. Pena and members of the Figueiredo and Jaffrey laboratories for helpful discussions. Publisher Copyright: © 2022, The Author(s), under exclusive licence to Springer Nature Limited.

PY - 2022/4/14

Y1 - 2022/4/14

N2 - RNA modifications are important regulators of gene expression1. In Trypanosoma brucei, transcription is polycistronic and thus most regulation happens post-transcriptionally2. N6-methyladenosine (m6A) has been detected in this parasite, but its function remains unknown3. Here we found that m6A is enriched in 342 transcripts using RNA immunoprecipitation, with an enrichment in transcripts encoding variant surface glycoproteins (VSGs). Approximately 50% of the m6A is located in the poly(A) tail of the actively expressed VSG transcripts. m6A residues are removed from the VSG poly(A) tail before deadenylation and mRNA degradation. Computational analysis revealed an association between m6A in the poly(A) tail and a 16-mer motif in the 3′ untranslated region of VSG genes. Using genetic tools, we show that the 16-mer motif acts as a cis-acting motif that is required for inclusion of m6A in the poly(A) tail. Removal of this motif from the 3′ untranslated region of VSG genes results in poly(A) tails lacking m6A, rapid deadenylation and mRNA degradation. To our knowledge, this is the first identification of an RNA modification in the poly(A) tail of any eukaryote, uncovering a post-transcriptional mechanism of gene regulation.

AB - RNA modifications are important regulators of gene expression1. In Trypanosoma brucei, transcription is polycistronic and thus most regulation happens post-transcriptionally2. N6-methyladenosine (m6A) has been detected in this parasite, but its function remains unknown3. Here we found that m6A is enriched in 342 transcripts using RNA immunoprecipitation, with an enrichment in transcripts encoding variant surface glycoproteins (VSGs). Approximately 50% of the m6A is located in the poly(A) tail of the actively expressed VSG transcripts. m6A residues are removed from the VSG poly(A) tail before deadenylation and mRNA degradation. Computational analysis revealed an association between m6A in the poly(A) tail and a 16-mer motif in the 3′ untranslated region of VSG genes. Using genetic tools, we show that the 16-mer motif acts as a cis-acting motif that is required for inclusion of m6A in the poly(A) tail. Removal of this motif from the 3′ untranslated region of VSG genes results in poly(A) tails lacking m6A, rapid deadenylation and mRNA degradation. To our knowledge, this is the first identification of an RNA modification in the poly(A) tail of any eukaryote, uncovering a post-transcriptional mechanism of gene regulation.

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