UAP56 is a conserved crucial component of a divergent mRNA export pathway in <i>Toxoplasma gondii</i>

Serpeloni, Mariana; Jiménez-Ruiz, Elena; Vidal, Newton Medeiros; Kroeber, Constanze; Andenmatten, Nicole; Lemgruber, Leandro; Mörking, Patrícia Alves; Pall, Gurman S.; Meissner, Markus; Ávila, Andréa Rodrigues

doi:10.1111/mmi.13485

Cited by 22 publications

(22 citation statements)

References 78 publications

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“…For transient gene disruption, each gene-specific gRNA (sequences in Table S4) vector, synthesised by GenScript using backbone vector pU6-SAG1gRNA-DHFR (Serpeloni et al, 2016), was co-transfected with the CAS9-FLAG expressing vector (pU6-universal) (Sidik et al, 2016).…”

Section: Plasmid Constructmentioning

confidence: 99%

Identification of the Toxoplasma gondii mitochondrial ribosome, and characterisation of a protein essential for mitochondrial translation

Lacombe

Maclean

Ovciarikova

et al. 2019

Molecular Microbiology

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Summary Apicomplexan parasites cause diseases such as malaria and toxoplasmosis. The apicomplexan mitochondrion shows striking differences from common model organisms, including fundamental processes such as mitochondrial translation. Despite evidence that mitochondrial translation is essential for parasite survival, it is largely understudied. Progress has been restricted by the absence of functional assays to detect apicomplexan mitochondrial translation, a lack of knowledge of proteins involved in the process and the inability to identify and detect mitoribosomes. We report the localization of 12 new mitochondrial proteins, including 6 putative mitoribosomal proteins. We demonstrate the integration of three mitoribosomal proteins in macromolecular complexes, and provide evidence suggesting these are apicomplexan mitoribosomal subunits, detected here for the first time. Finally, a new analytical pipeline detected defects in mitochondrial translation upon depletion of the small subunit protein 35 (TgmS35), while other mitochondrial functions remain unaffected. Our work lays a foundation for the study of apicomplexan mitochondrial translation.

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Section: Plasmid Constructmentioning

confidence: 99%

Identification of the Toxoplasma gondii mitochondrial ribosome, and characterisation of a protein essential for mitochondrial translation

Lacombe

Maclean

Ovciarikova

et al. 2019

Molecular Microbiology

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“…Perhaps investigations of the nuclear transport machineries in other divergent eukaryotes will help answer this question. Indeed, a recent study has shown that mRNA export pathways in Apicomplexa also appears to be divergent 86 ; in addition, so far investigators have been unable to identify any Mex67 orthologs in Toxoplasma gondii, indicating that this organism may be even more divergent Figure 3. Evolution of the NPC.…”

Section: Tms and Alps: Multiple Ways To Tether A Leviathanmentioning

confidence: 99%

“…86 In fact, very little is known about mRNA processing and export in most protists. Thus our study highlights and reinforces the need to sample and study a broad distribution of eukaryotic taxa to gain insight into evolutionary origins of function and mechanism at the nuclear envelope -as well as of course in many other cellular processes.…”

Section: Tms and Alps: Multiple Ways To Tether A Leviathanmentioning

confidence: 99%

Comparative interactomics provides evidence for functional specialization of the nuclear pore complex

Obado

Field

Rout

2017

Nucleus

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The core architecture of the eukaryotic cell was established well over one billion years ago, and is largely retained in all extant lineages. However, eukaryotic cells also possess lineagespecific features, frequently keyed to specific functional requirements. One quintessential core eukaryotic structure is the nuclear pore complex (NPC), responsible for regulating exchange of macromolecules between the nucleus and cytoplasm as well as acting as a nuclear organizational hub. NPC architecture has been best documented in one eukaryotic supergroup, the Opisthokonts (e.g. Saccharomyces cerevisiae and Homo sapiens), which although compositionally similar, have significant variations in certain NPC subcomplex structures. The variation of NPC structure across other taxa in the eukaryotic kingdom however, remains poorly understood. We explored trypanosomes, highly divergent organisms, and mapped and assigned their NPC proteins to specific substructures to reveal their NPC architecture. We showed that the NPC central structural scaffold is conserved, likely across all eukaryotes, but more peripheral elements can exhibit very significant lineage-specific losses, duplications or other alterations in their components. Amazingly, trypanosomes lack the major components of the mRNA export platform that are asymmetrically localized within yeast and vertebrate NPCs. Concomitant with this, the trypanosome NPC is ALMOST completely symmetric with the nuclear basket being the only major source of asymmetry. We suggest these features point toward a stepwise evolution of the NPC in which a coating scaffold first stabilized the pore after which selective gating emerged and expanded, leading to the addition of peripheral remodeling machineries on the nucleoplasmic and cytoplasmic sides of the pore.

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“…For transient gene disruption, each gene specific gRNA (sequences in Table S2) vector, synthesised by GenScript using backbone vector pU6-SAG1gRNA-DHFR (Serpeloni et al, 2016), was co-transfected with the CAS9-FLAG expressing vector (pU6-universal) (Sidik et al, 2016).…”

Section: Methodsmentioning

confidence: 99%

“…CRISPR/Cas9 transient expression - Parasites transiently co-expressing CAS9-FLAG expressing vector (pU6-universal) (Sidik et al, 2016) and gene specific sgRNA containing vector (Serpeloni et al, 2016), for genes TGME49_214790 / 263680 and 226280 , were fixed 24 hours post-transfection. The CAS9-FLAG and mitochondria were visualised by immunofluorescence as described above.…”

Section: Methodsmentioning

confidence: 99%

In silico screen identifies a new Toxoplasma gondii mitochondrial ribosomal protein essential for mitochondrial translation

Lacombe

Maclean

Ovciarikova

et al. 2019

Preprint

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22 23 24 2 Summary 25 26Apicomplexan parasites cause diseases such as malaria and toxoplasmosis. The 27 apicomplexan mitochondrion shows striking differences from common model organisms, 28including in fundamental processes such as mitochondrial translation. Despite evidence 29 that mitochondrial translation is essential for parasites survival, it is largely understudied. 30Progress has been restricted by the absence of functional assays to detect 31 apicomplexan mitochondrial translation, a lack of knowledge of proteins involved in the 32 process and the inability to identify and detect mitoribosomes. 33 Using mRNA expression patterns, 279 candidate mitochondrial housekeeping 34 components were identified in Toxoplasma. 11 were validated, including the 35 mitoribosomal small subunit protein 35 (TgmS35). TgmS35 tagging enabled the 36 detection of a macromolecular complex corresponding to the mitoribosomal small 37 subunit for the first time in apicomplexans. A new analytical pipeline detected defects in 38 mitochondrial translation upon TgmS35 depletion, while other mitochondrial functions 39 remain unaffected. Our work lays a foundation for the study of apicomplexan 40 mitochondrial translation. 41 42 43 Abbreviated summary 44 45 46The apicomplexan mitochondrion is divergent and essential yet poorly studied. 47Mitochondrial translation is predicted to utilize ribosomes assembled from fragmented 48 rRNA but this was never shown. Knowing the mitochondrial protein content is critical for 49 these studies. We identified 11 new mitochondrial proteins via in-silico searches. 50 Tagging and depletion of a mitoribosomal small subunit protein enabled the first 51 detection of a macromolecular ribosomal complex, and provided proof of principle for our 52 new mitochondrial translation analytic pipeline. 53 54 55 3 Introduction 56 57 Mitochondria are organelles of central importance to eukaryotic cells, providing key 58 nutrients and metabolites. Mitochondria are present in virtually all eukaryotic cells, 59 excluding a rare case of secondary loss (Karnkowska et al., 2016; Karnkowska and 60 Hampl, 2016). The recent interest in evolutionary cell biology has resulted in an 61 increasing appreciation of the diverse features of mitochondria found in divergent 62 organisms, which includes differences in fundamental pathways such as protein 63 translation (e.g. Ramrath et al., 2018). However, most of our knowledge of mitochondrial 64 biology is still based on studies in organisms that represent a small proportion of the 65 range of eukaryotic diversity, thus limiting our understanding. One of the challenges to 66 the study of mitochondrial biology in diverse organisms is the lack of tools to identify and 67 functionally characterise genes encoding species or phylum specific mitochondrial 68 proteins. 70Apicomplexa is a phylum of parasitic protists with high impact on human health globally, 71 which includes the malaria causing Plasmodium spp and the causative agent of 72 toxoplasmosis, Toxoplasma gondii. Apicomplexans represent o...

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UAP56 is a conserved crucial component of a divergent mRNA export pathway in Toxoplasma gondii

Cited by 22 publications

References 78 publications

Identification of the Toxoplasma gondii mitochondrial ribosome, and characterisation of a protein essential for mitochondrial translation

Identification of the Toxoplasma gondii mitochondrial ribosome, and characterisation of a protein essential for mitochondrial translation

Comparative interactomics provides evidence for functional specialization of the nuclear pore complex

In silico screen identifies a new Toxoplasma gondii mitochondrial ribosomal protein essential for mitochondrial translation

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