YTHDF proteins bind the N6-methyladenosine (m6A)-modified mRNAs, influencing their processing, stability, and translation. Therefore, the members of this protein family play crucial roles in gene regulation and several physiological and pathophysiological conditions. YTHDF proteins contain a hydrophobic pocket that accommodates the m6A embedded in the RRACH consensus sequence on mRNAs. We exploited the presence of this cage to set up an m6A-competitive assay and performed a high-throughput screen aimed at identifying ligands binding in the m6A pocket. We report the organoselenium compound ebselen as the first-in-class inhibitor of the YTHDF m6A-binding domain. Ebselen, whose interaction with YTHDF proteins was validated via orthogonal assays, cannot discriminate between the binding domains of the three YTHDF paralogs but can disrupt the interaction of the YTHDF m6A domain with the m6A-decorated mRNA targets. X-ray, mass spectrometry, and NMR studies indicate that in YTHDF1 ebselen binds close to the m6A cage, covalently to the Cys412 cysteine, or interacts reversibly depending on the reducing environment. We also showed that ebselen engages YTHDF proteins within cells, interfering with their mRNA binding. Finally, we produced a series of ebselen structural analogs that can interact with the YTHDF m6A domain, proving that ebselen expansion is amenable for developing new inhibitors. Our work demonstrates the feasibility of drugging the YTH domain in YTHDF proteins and opens new avenues for the development of disruptors of m6A recognition.Superscript/Subscript Available

Small-Molecule Ebselen Binds to YTHDF Proteins Interfering with the Recognition of N 6-Methyladenosine-Modified RNAs / Micaelli, Mariachiara; Dalle Vedove, Andrea; Cerofolini, Linda; Vigna, Jacopo; Sighel, Denise; Zaccara, Sara; Bonomo, Isabelle; Poulentzas, Georgios; Rosatti, Emanuele Filiberto; Cazzanelli, Giulia; Alunno, Laura; Belli, Romina; Peroni, Daniele; Dassi, Erik; Murakami, Shino; Jaffrey, Samie R; Fragai, Marco; Mancini, Ines; Lolli, Graziano; Quattrone, Alessandro; Provenzani, Alessandro. - In: ACS PHARMACOLOGY & TRANSLATIONAL SCIENCE. - ISSN 2575-9108. - 5:10(2022), pp. 872-891. [10.1021/acsptsci.2c00008]

Small-Molecule Ebselen Binds to YTHDF Proteins Interfering with the Recognition of N 6-Methyladenosine-Modified RNAs

Micaelli, Mariachiara;Dalle Vedove, Andrea;Vigna, Jacopo;Sighel, Denise;Zaccara, Sara;Bonomo, Isabelle;Poulentzas, Georgios;Rosatti, Emanuele Filiberto;Cazzanelli, Giulia;Alunno, Laura;Belli, Romina;Peroni, Daniele;Dassi, Erik;Mancini, Ines;Lolli, Graziano;Quattrone, Alessandro;Provenzani, Alessandro
2022-01-01

Abstract

YTHDF proteins bind the N6-methyladenosine (m6A)-modified mRNAs, influencing their processing, stability, and translation. Therefore, the members of this protein family play crucial roles in gene regulation and several physiological and pathophysiological conditions. YTHDF proteins contain a hydrophobic pocket that accommodates the m6A embedded in the RRACH consensus sequence on mRNAs. We exploited the presence of this cage to set up an m6A-competitive assay and performed a high-throughput screen aimed at identifying ligands binding in the m6A pocket. We report the organoselenium compound ebselen as the first-in-class inhibitor of the YTHDF m6A-binding domain. Ebselen, whose interaction with YTHDF proteins was validated via orthogonal assays, cannot discriminate between the binding domains of the three YTHDF paralogs but can disrupt the interaction of the YTHDF m6A domain with the m6A-decorated mRNA targets. X-ray, mass spectrometry, and NMR studies indicate that in YTHDF1 ebselen binds close to the m6A cage, covalently to the Cys412 cysteine, or interacts reversibly depending on the reducing environment. We also showed that ebselen engages YTHDF proteins within cells, interfering with their mRNA binding. Finally, we produced a series of ebselen structural analogs that can interact with the YTHDF m6A domain, proving that ebselen expansion is amenable for developing new inhibitors. Our work demonstrates the feasibility of drugging the YTH domain in YTHDF proteins and opens new avenues for the development of disruptors of m6A recognition.Superscript/Subscript Available
2022
10
Micaelli, Mariachiara; Dalle Vedove, Andrea; Cerofolini, Linda; Vigna, Jacopo; Sighel, Denise; Zaccara, Sara; Bonomo, Isabelle; Poulentzas, Georgios; Rosatti, Emanuele Filiberto; Cazzanelli, Giulia; Alunno, Laura; Belli, Romina; Peroni, Daniele; Dassi, Erik; Murakami, Shino; Jaffrey, Samie R; Fragai, Marco; Mancini, Ines; Lolli, Graziano; Quattrone, Alessandro; Provenzani, Alessandro
Small-Molecule Ebselen Binds to YTHDF Proteins Interfering with the Recognition of N 6-Methyladenosine-Modified RNAs / Micaelli, Mariachiara; Dalle Vedove, Andrea; Cerofolini, Linda; Vigna, Jacopo; Sighel, Denise; Zaccara, Sara; Bonomo, Isabelle; Poulentzas, Georgios; Rosatti, Emanuele Filiberto; Cazzanelli, Giulia; Alunno, Laura; Belli, Romina; Peroni, Daniele; Dassi, Erik; Murakami, Shino; Jaffrey, Samie R; Fragai, Marco; Mancini, Ines; Lolli, Graziano; Quattrone, Alessandro; Provenzani, Alessandro. - In: ACS PHARMACOLOGY & TRANSLATIONAL SCIENCE. - ISSN 2575-9108. - 5:10(2022), pp. 872-891. [10.1021/acsptsci.2c00008]
File in questo prodotto:
File Dimensione Formato  
acsptsci.2c00008.pdf

accesso aperto

Tipologia: Versione editoriale (Publisher’s layout)
Licenza: Creative commons
Dimensione 9.84 MB
Formato Adobe PDF
9.84 MB Adobe PDF Visualizza/Apri

I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione

Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/11572/359881
Citazioni
  • ???jsp.display-item.citation.pmc??? 0
  • Scopus 23
  • ???jsp.display-item.citation.isi??? 21
social impact