scholarly journals Large-scale prediction and analysis of protein sub-mitochondrial localization with DeepMito

2020 ◽  
Vol 21 (S8) ◽  
Author(s):  
Castrense Savojardo ◽  
Pier Luigi Martelli ◽  
Giacomo Tartari ◽  
Rita Casadio
Keyword(s):  
Subcellular Localization ◽  
Cellular Localization ◽  
Large Scale ◽  
Protein Localization ◽  
Functional Characterization ◽  
Experimental Information ◽  
Complete Characterization ◽  
Mitochondrial Proteins ◽  
Mitochondrial Localization

Abstract Background The prediction of protein subcellular localization is a key step of the big effort towards protein functional annotation. Many computational methods exist to identify high-level protein subcellular compartments such as nucleus, cytoplasm or organelles. However, many organelles, like mitochondria, have their own internal compartmentalization. Knowing the precise location of a protein inside mitochondria is crucial for its accurate functional characterization. We recently developed DeepMito, a new method based on a 1-Dimensional Convolutional Neural Network (1D-CNN) architecture outperforming other similar approaches available in literature. Results Here, we explore the adoption of DeepMito for the large-scale annotation of four sub-mitochondrial localizations on mitochondrial proteomes of five different species, including human, mouse, fly, yeast and Arabidopsis thaliana. A significant fraction of the proteins from these organisms lacked experimental information about sub-mitochondrial localization. We adopted DeepMito to fill the gap, providing complete characterization of protein localization at sub-mitochondrial level for each protein of the five proteomes. Moreover, we identified novel mitochondrial proteins fishing on the set of proteins lacking any subcellular localization annotation using available state-of-the-art subcellular localization predictors. We finally performed additional functional characterization of proteins predicted by DeepMito as localized into the four different sub-mitochondrial compartments using both available experimental and predicted GO terms. All data generated in this study were collected into a database called DeepMitoDB (available at http://busca.biocomp.unibo.it/deepmitodb), providing complete functional characterization of 4307 mitochondrial proteins from the five species. Conclusions DeepMitoDB offers a comprehensive view of mitochondrial proteins, including experimental and predicted fine-grain sub-cellular localization and annotated and predicted functional annotations. The database complements other similar resources providing characterization of new proteins. Furthermore, it is also unique in including localization information at the sub-mitochondrial level. For this reason, we believe that DeepMitoDB can be a valuable resource for mitochondrial research.

scholarly journals Functional characterization of 3D protein structures informed by human genetic diversity

2019 ◽  
Vol 116 (18) ◽  
pp. 8960-8965 ◽  
Author(s):  
Michael Hicks ◽  
Istvan Bartha ◽  
Julia di Iulio ◽  
J. Craig Venter ◽  
Amalio Telenti
Keyword(s):  
Large Scale ◽  
Functional Characterization ◽  
Protein Structures ◽  
Large Scale Data ◽  
Allosteric Sites ◽  
Missense Variation ◽  
Binding Pockets ◽  
Definition Of ◽  
Scale Data

Sequence variation data of the human proteome can be used to analyze 3D protein structures to derive functional insights. We used genetic variant data from nearly 140,000 individuals to analyze 3D positional conservation in 4,715 proteins and 3,951 homology models using 860,292 missense and 465,886 synonymous variants. Sixty percent of protein structures harbor at least one intolerant 3D site as defined by significant depletion of observed over expected missense variation. Structural intolerance data correlated with deep mutational scanning functional readouts for PPARG, MAPK1/ERK2, UBE2I, SUMO1, PTEN, CALM1, CALM2, and TPK1 and with shallow mutagenesis data for 1,026 proteins. The 3D structural intolerance analysis revealed different features for ligand binding pockets and orthosteric and allosteric sites. Large-scale data on human genetic variation support a definition of functional 3D sites proteome-wide.

scholarly journals Crowdsourced RNA design discovers diverse, reversible, efficient, self-contained molecular sensors

2019 ◽  
Author(s):  
Johan O. L. Andreasson ◽  
Michael R. Gotrik ◽  
Michelle J. Wu ◽  
Hannah K. Wayment-Steele ◽  
Wipapat Kladwang ◽  
...  
Keyword(s):  
High Throughput ◽  
Online Community ◽  
Large Scale ◽  
Functional Characterization ◽  
New Paradigm ◽  
Molecular Sensors ◽  
Rna Switches ◽  
Rna Sensors ◽  
Rna Design

AbstractInternet-based scientific communities promise a means to apply distributed, diverse human intelligence towards previously intractable scientific problems. However, current implementations have not allowed communities to propose experiments to test all emerging hypotheses at scale or to modify hypotheses in response to experiments. We report high-throughput methods for molecular characterization of nucleic acids that enable the large-scale videogame-based crowdsourcing of functional RNA sensor design, followed by high-throughput functional characterization. Iterative design testing of thousands of crowdsourced RNA sensor designs produced near-thermodynamically optimal and reversible RNA switches that act as self-contained molecular sensors and couple five distinct small molecule inputs to three distinct protein binding and fluorogenic outputs—results that surpass computational and expert-based design. This work represents a new paradigm for widely distributed experimental bioscience.One Sentence SummaryOnline community discovers standalone RNA sensors.

scholarly journals A Mycobacterial Systems Resource for the Research Community

mBio ◽  
2021 ◽  
Vol 12 (2) ◽  
Author(s):  
J. A. Judd ◽  
J. Canestrari ◽  
R. Clark ◽  
A. Joseph ◽  
P. Lapierre ◽  
...  
Keyword(s):  
Mycobacterium Smegmatis ◽  
Protein Localization ◽  
Functional Characterization ◽  
Model Organism ◽  
Research Community ◽  
Mycobacterial Species ◽  
Biological Resource ◽  
Content Type ◽  
Core Genes

ABSTRACT Functional characterization of bacterial proteins lags far behind the identification of new protein families. This is especially true for bacterial species that are more difficult to grow and genetically manipulate than model systems such as Escherichia coli and Bacillus subtilis. To facilitate functional characterization of mycobacterial proteins, we have established a Mycobacterial Systems Resource (MSR) using the model organism Mycobacterium smegmatis. This resource focuses specifically on 1,153 highly conserved core genes that are common to many mycobacterial species, including Mycobacterium tuberculosis, in order to provide the most relevant information and resources for the mycobacterial research community. The MSR includes both biological and bioinformatic resources. The biological resource includes (i) an expression plasmid library of 1,116 genes fused to a fluorescent protein for determining protein localization; (ii) a library of 569 precise deletions of nonessential genes; and (iii) a set of 843 CRISPR-interference (CRISPRi) plasmids specifically targeted to silence expression of essential core genes and genes for which a precise deletion was not obtained. The bioinformatic resource includes information about individual genes and a detailed assessment of protein localization. We anticipate that integration of these initial functional analyses and the availability of the biological resource will facilitate studies of these core proteins in many Mycobacterium species, including the less experimentally tractable pathogens M. abscessus, M. avium, M. kansasii, M. leprae, M. marinum, M. tuberculosis, and M. ulcerans. IMPORTANCE Diseases caused by mycobacterial species result in millions of deaths per year globally, and present a substantial health and economic burden, especially in immunocompromised patients. Difficulties inherent in working with mycobacterial pathogens have hampered the development and application of high-throughput genetics that can inform genome annotations and subsequent functional assays. To facilitate mycobacterial research, we have created a biological and bioinformatic resource (https://msrdb.org/) using Mycobacterium smegmatis as a model organism. The resource focuses specifically on 1,153 proteins that are highly conserved across the mycobacterial genus and, therefore, likely perform conserved mycobacterial core functions. Thus, functional insights from the MSR will apply to all mycobacterial species. We believe that the availability of this mycobacterial systems resource will accelerate research throughout the mycobacterial research community.

scholarly journals A robust metaproteomics pipeline for a holistic taxonomic and functional characterization of microbial communities from marine particles

2019 ◽  
Author(s):  
Doreen Schultz ◽  
Daniela Zühlke ◽  
Jörg Bernhardt ◽  
Thomas Ben Francis ◽  
Dirk Albrecht ◽  
...  
Keyword(s):  
Microbial Communities ◽  
Bacterial Communities ◽  
Large Scale ◽  
Functional Characterization ◽  
Phosphorus Uptake ◽  
Free Living ◽  
Technical Problems ◽  
Eukaryotic Microorganisms ◽  
Marine Particles

SummaryThis study aimed to establish a robust, reproducible and reliable metaproteomic pipeline for an in-depth characterization of marine particle-associated (PA) bacteria. To this end, we compared six well-established protein extraction protocols together with different MS-sample preparation techniques using particles sampled during a North Sea spring algae bloom in 2009. In this optimized workflow, proteins are extracted using a combination of SDS-containing lysis buffer and cell disruption by bead-beating, separated by SDS-PAGE, in-gel digested and analysed by LC-MS/MS, before MASCOT search against a metagenome-based database and data processing/visualization with the in-house-developed bioinformatics tools Prophane and Paver.As proof of principle, free-living (FL) and particulate communities sampled in April 2009 were analysed, resulting in an as yet unprecedented number of 9,354 and 5,034 identified protein groups for FL and PA bacteria, respectively. Our data revealed that FL and PA communities appeared similar in their taxonomic distribution, with notable exceptions: eukaryotic proteins and proteins assigned to Flavobacteriia, Cyanobacteria, and some proteobacterial genera were found more abundant on particles, whilst overall proteins belonging to Proteobacteria were more dominant in the FL fraction. In contrast, significant functional differences including proteins involved in polysaccharide degradation, sugar- and phosphorus uptake, adhesion, motility, and stress response were detected.Originality-Significance StatementMarine particles consist of organic particulate matter (e.g. phyto- or zooplankton) and particle-associated (PA) microbial communities, which are often embedded in a sugary matrix. A significant fraction of the decaying algal biomass in marine ecosystems is expected to be mineralized by PA heterotrophic communities, which are thus greatly contributing to large-scale carbon fluxes. Whilst numerous studies have investigated the succession of planktonic marine bacteria along phytoplankton blooms, the community structure and functionality of PA bacterial communities remained largely unexplored and knowledge on specific contributions of these microorganisms to carbon cycling is still surprisingly limited. This has been mostly been due to technical problems, i.e. to the difficulty to retrieve genomic DNA and proteins from these polysaccharide-rich entities, their enormous complexity and the high abundance of eukaryotic microorganisms.Our study presents an innovative, robust, reproducible, and reliable metaproteomics pipeline for marine particles, which will help to address and fill the above-described knowledge gap. Employing the here established workflow enabled us to identify more than 5,000 PA proteins, which is, at least to our knowledge, the largest number of protein groups ever assigned to marine particles. Notably, the novel pipeline has been validated by a first, comparative metaproteome analysis of free-living and PA bacterial communities indicating a significant functional shift enabling surface-associated bacteria to adapt to particle-specific living conditions. In conclusion, our novel metaproteomics pipeline presents a solid and promising methodological groundwork for future culture-independent analyses of seasonal taxonomic and functional successions of PA microbial communities in aquatic habitats.

scholarly journals Characterization of the phenotype with cognitive impairment and protein mislocalization in SCA34

2020 ◽  
Vol 6 (2) ◽  
pp. e403
Author(s):  
Marie Beaudin ◽  
Leila Sellami ◽  
Christian Martel ◽  
Lydia Touzel-Deschênes ◽  
Gabrielle Houle ◽  
...  
Keyword(s):  
Cellular Localization ◽  
Late Onset ◽  
Protein Localization ◽  
Age At Onset ◽  
Cerebellar Atrophy ◽  
Dermal Fibroblasts ◽  
Dominant Negative ◽  
Dominant Negative Effect ◽  
Negative Effect

ObjectiveTo better characterize the neurologic and cognitive profile of patients with spinocerebellar ataxia 34 (SCA34) caused by ELOVL4 mutations and to demonstrate the presence of ELOVL4 cellular localization and distribution abnormalities in skin-derived fibroblasts.MethodsWe investigated a 5-generation French-Canadian kindred presenting with a late-onset cerebellar ataxia and recruited age- and education-matched controls to evaluate the presence of neurocognitive impairment. Immunohistochemistry of dermal fibroblasts derived from a patient's skin biopsy was performed.ResultsPatients had a late-onset slowly progressive cerebellar syndrome (mean age at onset 47 years; range 32–60 years) characterized by truncal and limb ataxia, dysarthria, hypometric saccades, and saccadic pursuits. No patient had past or current signs of erythrokeratodermia variabilis, which had previously been reported. MRI revealed cerebellar atrophy, with pontine atrophy (4 of 6 patients), and cruciform hypersignal in the pons (2 of 6 patients). Fluorodeoxyglucose-PET showed diffuse cerebellar hypometabolism in all 5 tested patients with subtle parietal hypometabolism in 3. Significant cognitive deficits were found in executive functioning, along with apparent visuospatial, attention, and psychiatric involvement. Immunohistochemistry of dermal fibroblasts showed mislocalization of the ELOVL4 protein, which appeared punctate and aggregated, supporting a dominant negative effect of the mutation on protein localization.ConclusionsOur findings support the pathogenicity of ELOVL4 mutations in cerebellar dysfunction and provide a detailed characterization of the SCA34 phenotype, with neurocognitive changes typical of the cerebellar cognitive-affective syndrome.

scholarly journals Cellular localization and functional characterization of the equilibrative nucleoside transporters of antitumor nucleosides

2007 ◽  
Vol 98 (10) ◽  
pp. 1633-1637 ◽  
Author(s):  
Yoshio Endo ◽  
Tohru Obata ◽  
Daigo Murata ◽  
Mariho Ito ◽  
Kazuki Sakamoto ◽  
...  
Keyword(s):  
Cellular Localization ◽  
Functional Characterization ◽  
Nucleoside Transporters ◽  
Equilibrative Nucleoside Transporters
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