Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Crosslinks in Soluble Extract of DSSO Crosslinked Intact Tetrahymena thermophila Cilia

ABSTRACT: Intact cilia were isolated from dibucaine-treated Tetrahymena thermophila and subjected to DSSO crosslinking. Following quenching of the DSSO, a soluble "membrane and matrix" extract was generated by the addition of buffer containing 1% NP-40. The insoluble axonemes were removed by centrifugation and the supernatant extract was digested with trypsin, and crosslinked peptides were enriched using size exclusion chromatography. Crosslinks were identified using an MS2-MS3 data collection method and the Proteome Discoverer XlinkX node. Crosslink searching used a smaller fasta generated from analysis of the .RAW files and the Basic Sequest workflow.

INSTRUMENT(S): Orbitrap Fusion Lumos

ORGANISM(S): Tetrahymena Thermophila (ncbitaxon:5911)

SUBMITTER: Edward Marcotte

PROVIDER: MSV000090056 | MassIVE | Mon Aug 01 09:54:00 BST 2022

SECONDARY ACCESSION(S): PXD035702

REPOSITORIES: MassIVE

ACCESS DATA

Json Xml

Dataset's files

Source:

			Action	DRS
		Other

Items per page:

1 - 1 of 1

Similar Datasets

DSSO Crosslinking of Intact Tetrahymena Thermophila Cilia

Project description:The MS-cleavable crosslinker, DSSO, was added to cilia isolated from Tetrahymena thermophila. After digestion crosslinked peptides were enriched by size exclusion chromatography. Data were collected using an MS2/MS3 method. Analysis was done using the XlinkX node of PD2.3 and a fasta file generated from standard MS1/MS2 analysis of the crosslinked samples.

2022-07-19 | MSV000089917 | MassIVE

DSSO Crosslinking of IEX column fractions from Tetrahymena cilia extract fractionation.

Project description:Procedure details are reported in McCafferty et al. eLife2022;11e81977. Briefly cilia extract from Tetrahymena thermophila SB715 was fractionated on a mixed bed IEX HPLC column and all resulting fractions were crosslinked by addition of DSSO to 0.5mM. Following reduction, alkylation, trypsin digestion, and desalting, mass spectra were collected on an Orbitrap Fusion Lumos using a DDA MS2-MS3 method and spectra were processed in ProteomeDiscoverer 2.3 using the Xlinkx node to identify crosslinks. The look-up database for crosslink identification was generated from the same .RAW files using a standard Basic workflow to identify proteins present in the fractions.

2023-04-20 | MSV000091761 | MassIVE

DSSO Crosslinking of Partially Purified IFT-A from Tetrahymena thermophila

Project description:DSSO crosslinking data from two Tetrahymena thermophila samples: IEX fractions from chromatography of a cilia extract, and IFT-A-enriched fractions from preparative SEC of cilia extract.

2022-03-25 | MSV000089131 | MassIVE

Protein Identification in Tetrahymena thermophila cilia IEX fractions used for XL-MS

Project description:Tetrahymena thermophila SB715 were deciliated by pH shock substituting HEPES for Tris in all subsequent buffers. Ciliry proteins were solubilized with buffer containing 1%NP40, and fractionated by HPLC on a mixed-bed IEX column. DSSO was added to 0.5 mM to all collected fractions. Crosslinking was quenched with Tris pH 8.0 at 28 mM and fractions were reduced, alkylated, and digested with trypsin for mass spectrometry. Spectra were collected on an Orbitrap Fusion Lumos using a DDA MS2-MS3 method for both protein and crosslink identification. Data in this deposition include the RAW files and MSBlender pipeline files used for protein identification. Crosslink analysis will be deposited separately.

2023-04-13 | MSV000091710 | MassIVE

WT and FAP256-KO Cilia Proteome from Tetrahymena thermophila

Project description:Cilia are essential organelles that protrude from the cell body. Cilia are made of a microtubule-based structure called the axoneme. In most types of cilia, the ciliary tip is distinct from the rest of the cilium. By analysing Tetrahymena thermophila cells lacking a ciliary tip-enriched protein CEP104/FAP256 by mass spectrometry, we discovered candidates for the central pair cap complex and explained the potential functions of CEP104/FAP256. These data provide new insights into the function of the ciliary tip and the mechanisms of ciliary assembly and length regulation.

2023-09-01 | PXD042582 | Pride

Constitutive secretion in Tetrahymena thermophila.

Project description:The growth, survival, and life cycle progression of the freshwater ciliated protozoan Tetrahymena thermophila are responsive to protein signals thought to be released by constitutive secretion. In addition to providing insights about ciliate communication, studies of constitutive secretion are of interest for evaluating the utility of T. thermophila as a platform for the expression of secreted protein therapeutics. For these reasons, we undertook an unbiased investigation of T. thermophila secreted proteins using wild-type and secretion mutant strains. Extensive tandem mass spectrometry analyses of secretome samples were performed. We identified a total of 207 secretome proteins, most of which were not detected in a set of abundant whole-cell protein identifications. Numerous proteases and other hydrolases were secreted from cells grown in rich medium but not cells transferred to a nutrient starvation condition. On the other hand, we detected the starvation-enhanced secretion of a small number of cytosolic proteins, suggestive of an exosome-like pathway in T. thermophila. Subsets of proteins from the T. thermophila regulated secretion pathway were detected with differential representation across strains and culture conditions. Finally, many secretome proteins had a predicted N-terminal signal sequence but no other annotated characteristic or functional classification. Our work provides the first comprehensive analysis of secreted proteins in T. thermophila and establishes the groundwork for future studies of constitutive protein secretion biology and biotechnology in ciliates.

| S-EPMC2863955 | biostudies-literature

Multifunctional acyltransferases from Tetrahymena thermophila.

Project description:Multifunctional acyltransferases are able to catalyze the esterification of various acyl-acceptors with activated fatty acids. Here we describe the identification of four proteins from Tetrahymena thermophila that share certain properties with mammalian acyltransferases regarding their predicted transmembrane structure, their molecular mass and the typical acyltransferase motif. Expression of the Tetrahymena sequences results in production of triacylglycerols and wax esters in recombinant yeast when appropriate substrates are provided. The in vitro characterization shows, that these enzymes are capable of esterifying different acyl-acceptors including fatty alcohols, diols, diacylglycerols and isoprenols with acyl-CoA thioesters. Based on these catalytic activities and the sequence similarities of the Tetrahymena proteins with acyl-CoA:diacylglycerol acyltransferase 2 (DGAT2) family members, we conclude that we identified a new group of DGAT2-related multifunctional acyltransferases from protozoan organisms.

| S-EPMC3311841 | biostudies-literature

RNA-seq analysis of Tetrahymena thermophila

Project description:We present a comprehensive transcriptome of ciliate T. thermophila using the Illumina RNA-seq platform. The data was generated from the six mRNA samples of growth, starvation and conjugation of Tetrahymena. Despite an AT rich genome, there are about 124.6 million reads mapped to T. thermophila genome. Using these mapped reads, we have significantly improved the previous genome annotation and investigated the gene expression. Besides, our result also provided a comprehensive understanding of the alternative splicing in T. thermophila, and suggested the existence of the regulated unproductive splicing and translation (RUST) in the single-celled eukaryote.

2012-03-15 | GSE27971 | GEO

Septins stabilize mitochondria in Tetrahymena thermophila.

Project description:We describe phylogenetic and functional studies of three septins in the free-living ciliate Tetrahymena thermophila. Both deletion and overproduction of septins led to vacuolization of mitochondria, destabilization of the nuclear envelope, and increased autophagy. All three green fluorescent protein-tagged septins localized to mitochondria. Specific septins localized to the outer mitochondrial membrane, to septa formed during mitochondrial scission, or to the mitochondrion-associated endoplasmic reticulum. The only other septins known to localize to mitochondria are human ARTS and murine M-septin, both alternatively spliced forms of Sep4 (S. Larisch, Cell Cycle 3:1021-1023, 2004; S. Takahashi, R. Inatome, H. Yamamura, and S. Yanagi, Genes Cells 8:81-93, 2003). It therefore appears that septins have been recruited to mitochondrial functions independently in at least two eukaryotic lineages and in both cases are involved in apoptotic events.

| S-EPMC2519767 | biostudies-literature

RNA-seq analysis of Tetrahymena thermophila

2012-03-15 | E-GEOD-27971 | biostudies-arrayexpress