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.

Instituto Carlos Chagas FIOCRUZ Curitiba Paraná Brazil

National Center for Biotechnology Information National Library of Medicine National Institutes of Health Bethesda Maryland USA

School of Life Sciences University of Dundee Dundee Scotland UK

School of Life Sciences University of Dundee Dundee Scotland UK; Biology Centre University of South Bohemia České Budějovice Czech Republic

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Trypanosomes lack a canonical EJC but possess an UPF1 dependent NMD-like pathway

A lineage-specific protein network at the trypanosome nuclear envelope

Distinct mRNA and protein interactomes highlight functional differentiation of major eIF4F-like complexes from Trypanosoma brucei

Sending the message: specialized RNA export mechanisms in trypanosomes