One of the remarkable features of eukaryotes is the nucleus, delimited by the nuclear envelope (NE), a complex structure and home to the nuclear lamina and nuclear pore complex (NPC). For decades, these structures were believed to be mainly architectural elements and, in the case of the NPC, simply facilitating nucleocytoplasmic trafficking. More recently, the critical roles of the lamina, NPC and other NE constituents in genome organisation, maintaining chromosomal domains and regulating gene expression have been recognised. Importantly, mutations in genes encoding lamina and NPC components lead to pathogenesis in humans, while pathogenic protozoa disrupt the progression of normal development and expression of pathogenesis-related genes. Here, we review features of the lamina and NPC across eukaryotes and discuss how these elements are structured in trypanosomes, protozoa of high medical and veterinary importance, highlighting lineage-specific and conserved aspects of nuclear organisation.

• Klíčová slova
• evolutionary diversity, nuclear lamina, nuclear pore complex, nucleus, trypanosoma,
• MeSH
• aktivní transport - buněčné jádro fyziologie   MeSH
• jaderný obal MeSH
• jaderný pór genetika   metabolismus   MeSH
• komplex proteinů jaderného póru * metabolismus   MeSH
• lidé MeSH
• Trypanosoma * genetika   metabolismus   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH
• Názvy látek
• komplex proteinů jaderného póru * MeSH

Export of RNA from the nucleus is essential for all eukaryotic cells and has emerged as a major step in the control of gene expression. mRNA molecules are required to complete a complex series of processing events and pass a quality control system to protect the cytoplasm from the translation of aberrant proteins. Many of these events are highly conserved across eukaryotes, reflecting their ancient origin, but significant deviation from a canonical pathway as described from animals and fungi has emerged in the trypanosomatids. With significant implications for the mechanisms that control gene expression and hence differentiation, responses to altered environments and fitness as a parasite, these deviations may also reveal additional, previously unsuspected, mRNA export pathways.

• Klíčová slova
• eukaryogenesis, mRNA export, nuclear pore complex, polycistronic transcription, trans-splicing, trypanosomes,
• MeSH
• aktivní transport - buněčné jádro genetika   MeSH
• buněčné jádro genetika   metabolismus   MeSH
• messenger RNA genetika   MeSH
• RNA * genetika   metabolismus   MeSH
• Trypanosoma * genetika   metabolismus   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH
• Research Support, N.I.H., Extramural MeSH
• Research Support, U.S. Gov't, Non-P.H.S. MeSH
• Názvy látek
• messenger RNA MeSH
• RNA * MeSH

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.

• Klíčová slova
• evolution, gene expression, mRNA export, proteomics, trypanosomes,
• MeSH
• aktivní transport - buněčné jádro MeSH
• proteomika * MeSH
• RNA MeSH
• sestřih RNA MeSH
• transport RNA MeSH
• Trypanosoma cruzi * MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• RNA MeSH

Every type of nucleic acid in cells undergoes programmed chemical post-transcriptional modification. Generally, modification enzymes use substrates derived from intracellular metabolism, one exception is queuine (q)/queuosine (Q), which eukaryotes obtain from their environment; made by bacteria and ultimately taken into eukaryotic cells via currently unknown transport systems. Here, we use a combination of molecular, cell biology and biophysical approaches to show that in Trypanosoma brucei tRNA Q levels change dynamically in response to concentration variations of a sub-set of amino acids in the growth media. Most significant were variations in tyrosine, which at low levels lead to increased Q content for all the natural tRNAs substrates of tRNA-guanine transglycosylase (TGT). Such increase results from longer nuclear dwell time aided by retrograde transport following cytoplasmic splicing. In turn high tyrosine levels lead to rapid decrease in Q content. Importantly, the dynamic changes in Q content of tRNAs have negligible effects on global translation or growth rate but, at least, in the case of tRNATyr it affected codon choice. These observations have implications for the occurrence of other tunable modifications important for 'normal' growth, while connecting the intracellular localization of modification enzymes, metabolites and tRNAs to codon selection and implicitly translational output.

• MeSH
• aminokyseliny metabolismus   MeSH
• chromatografie kapalinová metody   MeSH
• guanin analogy a deriváty   metabolismus   MeSH
• kodon genetika   metabolismus   MeSH
• nukleosid Q metabolismus   MeSH
• pentosyltransferasy genetika   metabolismus   MeSH
• protozoální proteiny genetika   metabolismus   MeSH
• RNA transferová Tyr genetika   metabolismus   MeSH
• RNA transferová genetika   metabolismus   MeSH
• sestřih RNA MeSH
• tandemová hmotnostní spektrometrie metody   MeSH
• Trypanosoma brucei brucei genetika   metabolismus   MeSH
• tyrosin metabolismus   MeSH
• živiny metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH
• Názvy látek
• aminokyseliny MeSH
• guanin MeSH
• kodon MeSH
• nukleosid Q MeSH
• pentosyltransferasy MeSH
• protozoální proteiny MeSH
• queuine tRNA-ribosyltransferase MeSH Prohlížeč
• queuine MeSH Prohlížeč
• RNA transferová Tyr MeSH
• RNA transferová MeSH
• tyrosin MeSH

Kinetoplastids, including Trypanosoma brucei, control gene expression primarily at the posttranscriptional level. Nuclear mRNA export is an important, but understudied, step in this process. The general heterodimeric export factors, Mex67/Mtr2, function in the export of mRNAs and tRNAs in T. brucei, but RNA binding proteins (RBPs) that regulate export processes by controlling the dynamics of Mex67/Mtr2 ribonucleoprotein formation or transport have not been identified. Here, we report that DRBD18, an essential and abundant T. brucei RBP, associates with Mex67/Mtr2 in vivo, likely through its direct interaction with Mtr2. DRBD18 downregulation results in partial accumulation of poly(A)+ mRNA in the nucleus, but has no effect on the localization of intron-containing or mature tRNAs. Comprehensive analysis of transcriptomes from whole-cell and cytosol in DRBD18 knockdown parasites demonstrates that depletion of DRBD18 leads to impairment of nuclear export of a subset of mRNAs. CLIP experiments reveal the association of DRBD18 with several of these mRNAs. Moreover, DRBD18 knockdown leads to a partial accumulation of the Mex67/Mtr2 export receptors in the nucleus. Taken together, the current study supports a model in which DRBD18 regulates the selective nuclear export of mRNAs by promoting the mobilization of export competent mRNPs to the cytosol through the nuclear pore complex.

• Klíčová slova
• FISH, RNA binding protein, RNAseq, mRNA export, nucleoporin, trypanosome,
• MeSH
• aktivní transport - buněčné jádro MeSH
• genový knockdown metody   MeSH
• membránové transportní proteiny metabolismus   MeSH
• messenger RNA metabolismus   MeSH
• nukleocytoplazmatické transportní proteiny metabolismus   MeSH
• proteiny vázající RNA genetika   metabolismus   MeSH
• protozoální proteiny genetika   metabolismus   MeSH
• regulace genové exprese MeSH
• RNA transferová metabolismus   MeSH
• transkriptom MeSH
• transport RNA MeSH
• Trypanosoma brucei brucei genetika   metabolismus   MeSH
• vazba proteinů MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH
• Názvy látek
• membránové transportní proteiny MeSH
• messenger RNA MeSH
• nukleocytoplazmatické transportní proteiny MeSH
• proteiny vázající RNA MeSH
• protozoální proteiny MeSH
• RNA transferová MeSH

Transfer RNAs play a key role in protein synthesis. Following transcription, tRNAs are extensively processed prior to their departure from the nucleus to become fully functional during translation. This includes removal of 5′ leaders and 3′ trailers by a specific endo- and/or exonuclease, 3′ CCA tail addition, posttranscriptional modifications and in some cases intron removal. In this minireview, the critical factors of nuclear tRNA trafficking are described based on studies in classical models such as yeast and human cell lines. In addition, recent findings and identification of novel regulatory loops of nuclear tRNA trafficking in trypanosomes are discussed with emphasis on tRNA modifications. The comparison between the representatives of opisthokonts and excavates serves here to understand the evolutionary conservation and diversity of nuclear tRNA export mechanisms.

• Klíčová slova
• Nuclear tRNA export, Trypanosoma brucei, tRNA modification,
• MeSH
• buněčné linie MeSH
• lidé MeSH
• RNA jaderná genetika   metabolismus   MeSH
• RNA transferová genetika   metabolismus   MeSH
• Saccharomyces cerevisiae genetika   metabolismus   MeSH
• Trypanosoma genetika   metabolismus   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH
• Názvy látek
• RNA jaderná MeSH
• RNA transferová MeSH