Proteomics Uncovers Novel Components of an Interactive Protein Network Supporting RNA Export in Trypanosomes

• Published in: Molecular & cellular proteomics Vol. 21; no. 3; p. 100208
• Main Authors: Inoue, Alexandre Haruo, Domingues, Patricia Ferreira, Serpeloni, Mariana, Hiraiwa, Priscila Mazzocchi, Vidal, Newton Medeiros, Butterfield, Erin R., del Pino, Ricardo Canavate, Ludwig, Adriana, Boehm, Cordula, Field, Mark C., Ávila, Andréa Rodrigues
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 01.03.2022; American Society for Biochemistry and Molecular Biology
• Subjects: Active Transport, Cell Nucleus; evolution; gene expression; mRNA export; Proteomics; RNA; RNA Splicing; RNA Transport; Trypanosoma cruzi; trypanosomes; NPC; mRNA export; proteomics; NMD; NPGs; evolution; INT; UTR; mRNP; TREX; trypanosomes; gene expression; EJC
• DOI: 10.1016/j.mcpro.2022.100208
• ISSN: 1535-9476; 1535-9484

In trypanosomatids, transcription is polycistronic and all mRNAs are processed by trans-splicing, with export mediated by noncanonical mechanisms. Although mRNA export is central to gene regulation and expression, few orthologs of proteins involved in mRNA export in higher eukaryotes are detectable in trypanosome genomes, necessitating direct identification of protein components. We previously described conserved mRNA export pathway components in Trypanosoma cruzi, including orthologs of Sub2, a component of the TREX complex, and eIF4AIII (previously Hel45), a core component of the exon junction complex (EJC). Here, we searched for protein interactors of both proteins using cryomilling and mass spectrometry. Significant overlap between TcSub2 and TceIF4AIII-interacting protein cohorts suggests that both proteins associate with similar machinery. We identified several interactions with conserved core components of the EJC and multiple additional complexes, together with proteins specific to trypanosomatids. Additional immunoisolations of kinetoplastid-specific proteins both validated and extended the superinteractome, which is capable of supporting RNA processing from splicing through to nuclear export and cytoplasmic events. We also suggest that only proteomics is powerful enough to uncover the high connectivity between multiple aspects of mRNA metabolism and to uncover kinetoplastid-specific components that create a unique amalgam to support trypanosome mRNA maturation. [Display omitted] •Gene expression in trypanosomes is mediated by noncanonical mechanisms.•Trypanosome mRNA nuclear export system comprises unique proteins to kinetoplastids.•The present work highlights an amalgam of kinetoplastid-specific and conserved components.•Our data support a highly coupled mRNA maturation pathway. This work was executed under full ethical compliance, and authorship is limited to those who have made significant contributions. The manuscript is original work, and works of others have been appropriately cited. We provide raw data for appropriate datasets at public databases. This work was performed standard compliant with community-acceptable guidelines and parameters. No known hazard was caused, nor any involvement of human subjects. Animal use for production of antibodies was performed under the institutional ethical guidelines.