Pseudouridine increases ribosome stability in a thermophilic eukaryote Chaetomium thermophilum
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PROVIDER: PRJEB86624 | ENA |
REPOSITORIES: ENA
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Project description:A correct genome annotation is fundamental for research in the field of molecular and structural biology. The annotation of the reference genome of Chaetomium thermophilum has been reported previously, but it is limited to open reading frames (ORFs) of genes and contains only a few noncoding transcripts. In this study, we identified and annotated by deep RNA sequencing full-length transcripts of C.thermophilum. We identified 7044 coding genes and a large number of noncoding genes (n=4567). Astonishingly, 23% of the coding genes are alternatively spliced. We identified 679 novel coding genes and corrected the structural organization of more than 50% of the previously annotated genes. Furthermore, we substantially extended the Gene Ontology (GO) and Enzyme Commission (EC) lists, which provide comprehensive search tools for potential industrial applications and basic research. The identified novel transcripts and improved annotation will help understanding the gene regulatory landscape in C.thermophilum. The analysis pipeline developed here can be used to build transcriptome assemblies and identify coding and noncoding RNAs of other species. The new genome annotation of the GTF file can be found here.
2019-12-31 | GSE116834 | GEO
Project description:Mitochondrial complex I is a redox-driven proton pump that generates most of the proton-motive force powering oxidative phosphorylation and ATP synthesis in eukaryotes. We report the structure of complex I from the thermophilic eukaryote Chaetomium thermophilum, determined by electron cryo-microscopy to 2.4 A resolution in the open and closed conformation. Complex I has two arms, the peripheral and membrane arm, forming an L-shape. The two conformations differ in the relative position of the two arms. The open-to-closed transition is accompanied by substantial conformational changes in the Q-binding cavity and the E-channel, and by the formation of an aqueous connection between the E-channel and an extensive aqueous passage inside the membrane arm. The observed similarities provide strong support for a conserved, common mechanism that applies across all species from fungi to mammals. Furthermore, the complex is inhibited by the detergent DDM, which binds reversibly to two sites in the Q-binding cavity.
2022-11-29 | PXD033234 | Pride
Project description:The thermophilic fungus Chaetomium thermophilum has been successfully used in the past for biochemical and high resolution structural studies of protein complexes, but subsequent functional analysis of these assemblies were hindered due to the lack of genetic tools in this thermophile, which are typically amenable in several other mesophilic eukaryotic model organisms, in particular the yeast Saccharomycers cerevisiae. Hence, we aimed to develop a regulatable gene-expression system in C. thermophilum, which might facilitate such in vivo studies, based on what we know about the galactose-inducible GAL promoter in yeast. To identify sugar-regulatable promoters in C. thermophilum, we performed comparative xylose- versus glucose-dependent gene expression studies, which uncovered a number of enzymes induced by xylose but repressed by glucose. Subsequently, we cloned the promoters of the two most stringently regulated genes, the xylosidase-like gene (XYL) and xylitol dehydrogenase (XDH), obtained from this genome-wide analysis in front of the thermostable YFP (yellow fluorescent protein) reporter. In this way, we could demonstrate xylose-dependent YFP expression by either western blotting or life cell imaging fluorescence microscopy. Prompted by these results, we finally expressed a well-characterized dominant-negative ribosome assembly factor mutant, rsa4 E117>D, under the control of the XDH promoter, which allowed us to induce a nuclear export defect of the pre-60S subunit when C. thermophilum cells were grown in xylose but not glucose containing medium. Altogether, our study recognized xylose-regulatable promoters in Chaetomium thermophilum, which may foster functional studies of genes of interest in this thermophilic eukaryotic model organism.
2022-09-26 | GSE214043 | GEO
Project description:Global Transcriptome Characterization and Assembly of thermophilic ascomycete Chaetomium thermophilum
| PRJNA480310 | ENA
Project description:TMT-labeled LC-MSMS was performed to identify and quantify proteins from three different TAP-purification pull-outs from Chaetomium thermophilum. The aim was to identify interaction partners and to study, whether the two tagged proteins (Naa50 and Naa15) are likely to interact, as they do in other organisms. CtNaa50 is a special homolog of known Naa50 proteins and in this case, it does not interact with Naa15, but other identified proteins.
2022-10-14 | PXD035320 | Pride
Project description:Prp28 (pre-mRNA-splicing ATP-dependent RNA helicase 28) is a spliceosomal DEAD-box helicase which is involved in two steps of spliceosome assembly. It is required for the formation of commitment complex 2 in an ATP-independent manner as well as for the formation of the pre-catalytic spliceosome, which in contrast is ATP-dependent. During the latter step, Prp28 is crucial for the integration of the U4/U6·U5 tri-snRNP since it displaces the U1 snRNP and allows the U6 snRNP to base-pair with the 5'-splice site. Here, the crystal structure of Prp28 from the thermophilic fungus Chaetomium thermophilum is reported at 3.2 Å resolution and is compared with the available structures of homologues.
| S-EPMC4854570 | biostudies-literature
Project description:Mitochondrial complex I is a redox-driven proton pump that generates proton-motive force across the inner mitochondrial membrane, powering oxidative phosphorylation and ATP synthesis in eukaryotes. We report the structure of complex I from the thermophilic fungus Chaetomium thermophilum, determined by cryoEM up to 2.4-Å resolution. We show that the complex undergoes a transition between two conformations, which we refer to as state 1 and state 2. The conformational switch is manifest in a twisting movement of the peripheral arm relative to the membrane arm, but most notably in substantial rearrangements of the Q-binding cavity and the E-channel, resulting in a continuous aqueous passage from the E-channel to subunit ND5 at the far end of the membrane arm. The conformational changes in the complex interior resemble those reported for mammalian complex I, suggesting a highly conserved, universal mechanism of coupling electron transport to proton pumping.
| S-EPMC9699679 | biostudies-literature
Project description:RNA helicases are indispensable for all organisms in each domain of life and have implications in numerous cellular processes. The DEAH-box RNA helicase Prp43 is involved in pre-mRNA splicing as well as rRNA maturation. Here, the crystal structure of Chaetomium thermophilum Prp43 at 2.9 Å resolution is revealed. Furthermore, it is demonstrated that Prp43 from C. thermophilum is capable of functionally replacing its orthologue from Saccharomyces cerevisiae in spliceosomal disassembly assays.
| S-EPMC4741191 | biostudies-literature
Project description:The arrangement of proteins into complexes is a key organizational principle for many cellular functions. Although the topology of many complexes has been systematically analyzed in isolation, their molecular sociology in situ remains elusive. Here, we show that crude cellular extracts of a eukaryotic thermophile, Chaetomium thermophilum, retain basic principles of cellular organization. Using a structural proteomics approach, we simultaneously characterized the abundance, interactions and structure of a third of the C. thermophilum proteome within these extracts. We identified 27 distinct protein communities that include 108 interconnected complexes, which dynamically associate with each other and functionally benefit from being in close proximity in the cell.
2017-08-07 | PXD006660 | Pride
Project description:The arrangement of proteins into complexes is a key organizational principle for many cellular functions. Although the topology of many complexes has been systematically analyzed in isolation, their molecular sociology in situ remains elusive. Here, we show that crude cellular extracts of a eukaryotic thermophile, Chaetomium thermophilum, retain basic principles of cellular organization. Using a structural proteomics approach, we simultaneously characterized the abundance, interactions and structure of a third of the C. thermophilum proteome within these extracts. We identified 27 distinct protein communities that include 108 interconnected complexes, which dynamically associate with each other and functionally benefit from being in close proximity in the cell. Furthermore, we investigated the structure of fatty acid synthase within these extracts by cryoEM and this revealed multiple, flexible states of the enzyme in adaptation to its association with other complexes, thus exemplifying the need for in situ studies. As the components of the captured protein communities are known – at both the protein and complex level – this study constitutes another step forward towards a molecular understanding ofsubcellular organization.
2017-07-26 | PXD006626 | Pride