A Python library for FAIRer access and deposition to the Metabolomics Workbench Data Repository

Introduction

The Metabolomics Workbench Data Repository is a public repository of mass spectrometry and nuclear magnetic resonance data and metadata derived from a wide variety of metabolomics studies. The data and metadata for each study is deposited, stored, and accessed via files in the domain-specific ‘mwTab’ flat file format.

Objectives

In order to improve the accessibility, reusability, and interoperability of the data and metadata stored in ‘mwTab’ formatted files, we implemented a Python library and package. This Python package, named ‘mwtab’, is a parser for the domain-specific ‘mwTab’ flat file format, which provides facilities for reading, accessing, and writing ‘mwTab’ formatted files. Furthermore, the package provides facilities to validate both the format and required metadata elements of a given ‘mwTab’ formatted file.

Methods

In order to develop the ‘mwtab’ package we used the official ‘mwTab’ format specification. We used Git version control along with Python unit-testing framework as well as continuous integration service to run those tests on multiple versions of Python. Package documentation was developed using sphinx documentation generator.

Results

The ‘mwtab’ package provides both Python programmatic library interfaces and command-line interfaces for reading, writing, and validating ‘mwTab’ formatted files. Data and associated metadata are stored within Python dictionary- and list-based data structures, enabling straightforward, ‘pythonic’ access and manipulation of data and metadata. Also, the package provides facilities to convert ‘mwTab’ files into a JSON formatted equivalent, enabling easy reusability of the data by all modern programming languages that implement JSON parsers. The ‘mwtab’ package implements its metadata validation functionality based on a pre-defined JSON schema that can be easily specialized for specific types of metabolomics studies. The library also provides a command-line interface for interconversion between ‘mwTab’ and JSONized formats in raw text and a variety of compressed binary file formats.

Conclusions

The ‘mwtab’ package is an easy-to-use Python package that provides FAIRer utilization of the Metabolomics Workbench Data Repository. The source code is freely available on GitHub and via the Python Package Index. Documentation includes a ‘User Guide’, ‘Tutorial’, and ‘API Reference’. The GitHub repository also provides ‘mwtab’ package unit-tests via a continuous integration service.

     

Abnormal Astrocytosis in the Basal Ganglia Pathway of Git1?/? Mice

Attention deficit/hyperactivity disorder (ADHD) is one of the most common neurodevelopmental disorders, affecting approximately 5% of children. However, the neural mechanisms underlying its development and treatment are yet to be elucidated. In this study, we report that an ADHD mouse model, which harbors a deletion in the Git1 locus, exhibits severe astrocytosis in the globus pallidus (GP) and thalamic reticular nucleus (TRN), which send modulatory GABAergic inputs to the thalamus. A moderate level of astrocytosis was displayed in other regions of the basal ganglia pathway, including the ventrobasal thalamus and cortex, but not in other brain regions, such as the caudate putamen, basolateral amygdala, and hippocampal CA1. This basal ganglia circuit-selective astrocytosis was detected in both in adult (2–3 months old) and juvenile (4 weeks old) Git1^−/− mice, suggesting a developmental origin. Astrocytes play an active role in the developing synaptic circuit; therefore, we performed an immunohistochemical analysis of synaptic markers. We detected increased and decreased levels of GABA and parvalbumin (PV), respectively, in the GP. This suggests that astrocytosis may alter synaptic transmission in the basal ganglia. Intriguingly, increased GABA expression colocalized with the astrocyte marker, GFAP, indicative of an astrocytic origin. Collectively, these results suggest that defects in basal ganglia circuitry, leading to impaired inhibitory modulation of the thalamus, are neural correlates for the ADHD-associated behavioral manifestations in Git1^−/− mice.     

Specific Dephosphorylation at Tyr-554 of Git1 by Ptprz Promotes Its Association with Paxillin and Hic-5

G protein-coupled receptor kinase-interactor 1 (Git1) is involved in cell motility control by serving as an adaptor that links signaling proteins such as Pix and PAK to focal adhesion proteins. We previously demonstrated that Git1 was a multiply tyrosine-phosphorylated protein, its primary phosphorylation site was Tyr-554 in the vicinity of the focal adhesion targeting-homology (FAH) domain, and this site was selectively dephosphorylated by protein tyrosine phosphatase receptor type Z (Ptprz). In the present study, we showed that Tyr-554 phosphorylation reduced the association of Git1 with the FAH-domain-binding proteins, paxillin and Hic-5, based on immunoprecipitation experiments using the Tyr-554 mutants of Git1. The Tyr-554 phosphorylation of Git1 was higher, and its binding to paxillin was consistently lower in the brains of Ptprz-deficient mice than in those of wild-type mice. We then investigated the role of Tyr-554 phosphorylation in cell motility control using three different methods: random cell motility, wound healing, and Boyden chamber assays. The shRNA-mediated knockdown of endogenous Git1 impaired cell motility in A7r5 smooth muscle cells. The motility defect was rescued by the exogenous expression of wild-type Git1 and a Git1 mutant, which only retained Tyr-554 among the multiple potential tyrosine phosphorylation sites, but not by the Tyr-554 phosphorylation-defective or phosphorylation-state mimic Git1 mutant. Our results suggested that cyclic phosphorylation-dephosphorylation at Tyr-554 of Git1 was crucial for dynamic interactions between Git1 and paxillin/Hic-5 in order to ensure coordinated cell motility.     

msatcommander: detection of microsatellite repeat arrays and automated, locus-specific primer design

msatcommander is a platform-independent program designed to search for microsatellite arrays, design primers, and tag primers using an automated routine. msatcommander accepts as input DNA sequence data in single-sequence or concatenated, fasta-formatted files. Search data and locus-specific primers are written to comma-separated value files for subsequent use in spreadsheet or database programs. Binary versions of the graphical interface for msatcommander are available for Apple OS X and Windows XP. Users of other operating systems may run the graphical interface version using the available source code, provided their environment supports at least Python 2.4, Biopython 1.43, and wxPython 2.8. msatcommander is available from http://code.google.com/p/msatcommander/.     

GUI-BioPASED: A program for molecular dynamics simulations of biopolymers with a graphical user interface

There are several software packages for simulating the main types of biopolymers with molecular dynamics methods. However, work with these packages is rather complicated, especially for experimenters, who are nonexperts in computational structural biology. We have developed GUI-BioPASED, a web-based GUI for the molecular dynamics software BioPASED. This version provides for formulating the task as a single executable file, exporting this file to user’s PC, ensuring its execution, and, thereby, partially automating the task part responsible for the majority of errors. A user-friendly cross-platform interface is created, which performs data check, reports errors, and hints at possible ways to correct them. BioPASED, a general purpose molecular dynamics program, and GUI-BioPASED are described along with a typical example of their usage.     

MacSyFinder: A Program to Mine Genomes for Molecular Systems with an Application to CRISPR-Cas Systems

Motivation

Biologists often wish to use their knowledge on a few experimental models of a given molecular system to identify homologs in genomic data. We developed a generic tool for this purpose.

Results

Macromolecular System Finder (MacSyFinder) provides a flexible framework to model the properties of molecular systems (cellular machinery or pathway) including their components, evolutionary associations with other systems and genetic architecture. Modelled features also include functional analogs, and the multiple uses of a same component by different systems. Models are used to search for molecular systems in complete genomes or in unstructured data like metagenomes. The components of the systems are searched by sequence similarity using Hidden Markov model (HMM) protein profiles. The assignment of hits to a given system is decided based on compliance with the content and organization of the system model. A graphical interface, MacSyView, facilitates the analysis of the results by showing overviews of component content and genomic context. To exemplify the use of MacSyFinder we built models to detect and class CRISPR-Cas systems following a previously established classification. We show that MacSyFinder allows to easily define an accurate “Cas-finder” using publicly available protein profiles.

Availability and Implementation

MacSyFinder is a standalone application implemented in Python. It requires Python 2.7, Hmmer and makeblastdb (version 2.2.28 or higher). It is freely available with its source code under a GPLv3 license at https://github.com/gem-pasteur/macsyfinder. It is compatible with all platforms supporting Python and Hmmer/makeblastdb. The “Cas-finder” (models and HMM profiles) is distributed as a compressed tarball archive as Supporting Information.     

Setup of a scientific computing environment for computational biology: Simulation of a genome-scale metabolic model of Escherichia coli as an example

Computational analysis of biological data is becoming increasingly important, especially in this era of big data. Computational analysis of biological data allows efficiently deriving biological insights for given data, and sometimes even counterintuitive ones that may challenge the existing knowledge. Among experimental researchers without any prior exposure to computer programming, computational analysis of biological data has often been considered to be a task reserved for computational biologists. However, thanks to the increasing availability of user-friendly computational resources, experimental researchers can now easily access computational resources, including a scientific computing environment and packages necessary for data analysis. In this regard, we here describe the process of accessing Jupyter Notebook, the most popular Python coding environment, to conduct computational biology. Python is currently a mainstream programming language for biology and biotechnology. In particular, Anaconda and Google Colaboratory are introduced as two representative options to easily launch Jupyter Notebook. Finally, a Python package COBRApy is demonstrated as an example to simulate 1) specific growth rate of Escherichia coli as well as compounds consumed or generated under a minimal medium with glucose as a sole carbon source, and 2) theoretical production yield of succinic acid, an industrially important chemical, using E. coli. This protocol should serve as a guide for further extended computational analyses of biological data for experimental researchers without computational background.     

Algamica: revisions to a key-based computerized counting program for free-living,attached, and benthic algae

A computerized counting program for algae and other microscopic bodies, named Algamica, is presented here. This program is a revised version of the original computer counting program of Hamilton published in 1990. This DOS-based software can enumerate all types of microscopic algae (i.e. phytoplankton, periphyton, diatoms), for which adequate expression of results are provided. Automated calculations of densities, biovolumes, surface areas and carbon biomass are available at the termination of each sample count. A simple counter for other micro-organisms is also available. A comprehensive guide manual file has been added to allow for a friendly first contact with the program and its options. This software conforms to current enumeration methodology. This version is available for PC computers, from website Algamica.ibelgique.com. Minimum required memory is 200 KB.     

Deriving quantitative dynamics information for proteins and RNAs using ROTDIF with a graphical user interface

To facilitate rigorous analysis of molecular motions in proteins, DNA, and RNA, we present a new version of ROTDIF, a program for determining the overall rotational diffusion tensor from single- or multiple-field nuclear magnetic resonance relaxation data. We introduce four major features that expand the program’s versatility and usability. The first feature is the ability to analyze, separately or together, ¹³C and/or ¹⁵N relaxation data collected at a single or multiple fields. A significant improvement in the accuracy compared to direct analysis of R ₂/R ₁ ratios, especially critical for analysis of ¹³C relaxation data, is achieved by subtracting high-frequency contributions to relaxation rates. The second new feature is an improved method for computing the rotational diffusion tensor in the presence of biased errors, such as large conformational exchange contributions, that significantly enhances the accuracy of the computation. The third new feature is the integration of the domain alignment and docking module for relaxation-based structure determination of multi-domain systems. Finally, to improve accessibility to all the program features, we introduced a graphical user interface that simplifies and speeds up the analysis of the data. Written in Java, the new ROTDIF can run on virtually any computer platform. In addition, the new ROTDIF achieves an order of magnitude speedup over the previous version by implementing a more efficient deterministic minimization algorithm. We not only demonstrate the improvement in accuracy and speed of the new algorithm for synthetic and experimental ¹³C and ¹⁵N relaxation data for several proteins and nucleic acids, but also show that careful analysis required especially for characterizing RNA dynamics allowed us to uncover subtle conformational changes in RNA as a function of temperature that were opaque to previous analysis.     

multiplierz v2.0: A Python‐based ecosystem for shared access and analysis of native mass spectrometry data

The continued evolution of modern mass spectrometry instrumentation and associated methods represents a critical component in efforts to decipher the molecular mechanisms which underlie normal physiology and understand how dysregulation of biological pathways contributes to human disease. The increasing scale of these experiments combined with the technological diversity of mass spectrometers presents several challenges for community‐wide data access, analysis, and distribution. Here we detail a redesigned version of multiplierz, our Python software library which leverages our common application programming interface (mzAPI) for analysis and distribution of proteomic data. New features include support for a wider range of native mass spectrometry file types, interfaces to additional database search engines, compatibility with new reporting formats, and high‐level tools to perform post‐search proteomic analyses. A GUI desktop environment, mzDesktop, provides access to multiplierz functionality through a user friendly interface. multiplierz is available for download from: https://github.com/BlaisProteomics/multiplierz ; and mzDesktop is available for download from: https://sourceforge.net/projects/multiplierz/     

An ADP-ribosylation factor GTPase-activating protein Git2-short/KIAA0148 is involved in subcellular localization of paxillin and actin cytoskeletal organization

Paxillin acts as an adaptor protein in integrin signaling. We have shown that paxillin exists in a relatively large cytoplasmic pool, including perinuclear areas, in addition to focal complexes formed at the cell periphery and focal adhesions formed underneath the cell. Several ADP-ribosylation factor (ARF) GTPase-activating proteins (GAPs; ARFGAPs) have been shown to associate with paxillin. We report here that Git2-short/KIAA0148 exhibits properties of a paxillin-associated ARFGAP and appears to be colocalized with paxillin, primarily at perinuclear areas. A fraction of Git2-short was also localized to actin-rich structures at the cell periphery. Unlike paxillin, however, Git2-short did not accumulate at focal adhesions underneath the cell. Git2-short is a short isoform of Git2, which is highly homologous to p95PKL, another paxillin-binding protein, and showed a weaker binding affinity toward paxillin than that of Git2. The ARFGAP activities of Git2 and Git2-short have been previously demonstrated in vitro, and we provided evidence that at least one ARF isoform, ARF1, is an intracellular substrate for the GAP activity of Git2-short. We also showed that Git2-short could antagonize several known ARF1-mediated phenotypes: overexpression of Git2-short, but not its GAP-inactive mutant, caused the redistribution of Golgi protein beta-COP and reduced the amounts of paxillin-containing focal adhesions and actin stress fibers. Perinuclear localization of paxillin, which was sensitive to ARF inactivation, was also affected by Git2-short overexpression. On the other hand, paxillin localization to focal complexes at the cell periphery was unaffected or even augmented by Git2-short overexpression. Therefore, an ARFGAP protein weakly interacting with paxillin, Git2-short, exhibits pleiotropic functions involving the regulation of Golgi organization, actin cytoskeletal organization, and subcellular localization of paxillin, all of which need to be coordinately regulated during integrin-mediated cell adhesion and intracellular signaling.     

The Quality of Methods Reporting in Parasitology Experiments

There is a growing concern both inside and outside the scientific community over the lack of reproducibility of experiments. The depth and detail of reported methods are critical to the reproducibility of findings, but also for making it possible to compare and integrate data from different studies. In this study, we evaluated in detail the methods reporting in a comprehensive set of trypanosomiasis experiments that should enable valid reproduction, integration and comparison of research findings. We evaluated a subset of other parasitic (Leishmania, Toxoplasma, Plasmodium, Trichuris and Schistosoma) and non-parasitic (Mycobacterium) experimental infections in order to compare the quality of method reporting more generally. A systematic review using PubMed (2000–2012) of all publications describing gene expression in cells and animals infected with Trypanosoma spp was undertaken based on PRISMA guidelines; 23 papers were identified and included. We defined a checklist of essential parameters that should be reported and have scored the number of those parameters that are reported for each publication. Bibliometric parameters (impact factor, citations and h-index) were used to look for association between Journal and Author status and the quality of method reporting. Trichuriasis experiments achieved the highest scores and included the only paper to score 100% in all criteria. The mean of scores achieved by Trypanosoma articles through the checklist was 65.5% (range 32–90%). Bibliometric parameters were not correlated with the quality of method reporting (Spearman''s rank correlation coefficient <−0.5; p>0.05). Our results indicate that the quality of methods reporting in experimental parasitology is a cause for concern and it has not improved over time, despite there being evidence that most of the assessed parameters do influence the results. We propose that our set of parameters be used as guidelines to improve the quality of the reporting of experimental infection models as a pre-requisite for integrating and comparing sets of data.     

EphA2 Engages Git1 to Suppress Arf6 Activity Modulating Epithelial Cell–Cell Contacts

ADP-ribosylation factor (Arf) 6 activity is crucially involved in the regulation of E-cadherin–based cell–cell adhesions. Erythropoietin-producing hepatocellular carcinoma (Eph)-family receptors recognize ligands, namely, ephrins, anchored to the membrane of apposing cells, and they mediate cell–cell contact-dependent events. Here, we found that Arf6 activity is down-regulated in Madin-Darby canine kidney cells, which is dependent on cell density and calcium ion concentration, and we provide evidence of a novel signaling pathway by which ligand-activated EphA2 suppresses Arf6 activity. This EphA2-mediated suppression of Arf6 activity was linked to the induction of cell compaction and polarization, but it was independent of the down-regulation of extracellular signal-regulated kinase 1/2 kinase activity. We show that G protein-coupled receptor kinase-interacting protein (Git) 1 and noncatalytic region of tyrosine kinase (Nck) 1 are involved in this pathway, in which ligand-activated EphA2, via its phosphorylated Tyr594, binds to the Src homology 2 domain of Nck1, and then via its Src homology 3 domain binds to the synaptic localizing domain of Git1 to suppress Arf6 activity. We propose a positive feedback loop in which E-cadherin–based cell–cell contacts enhance EphA-ephrinA signaling, which in turn down-regulates Arf6 activity to enhance E-cadherin–based cell–cell contacts as well as the apical-basal polarization of epithelial cells.     

The cell adhesion-associated protein Git2 regulates morphogenetic movements during zebrafish embryonic development

Signaling through cell adhesion complexes plays a critical role in coordinating cytoskeletal remodeling necessary for efficient cell migration. During embryonic development, normal morphogenesis depends on a series of concerted cell movements; but the roles of cell adhesion signaling during these movements are poorly understood. The transparent zebrafish embryo provides an excellent system to study cell migration during development. Here, we have identified zebrafish git2a and git2b, two new members of the GIT family of genes that encode ArfGAP proteins associated with cell adhesions. Loss-of-function studies revealed an essential role for Git2a in zebrafish cell movements during gastrulation. Time-lapse microscopy analysis demonstrated that antisense depletion of Git2a greatly reduced or arrested cell migration towards the vegetal pole of the embryo. These defects were rescued by expression of chicken GIT2, indicating a specific and conserved role for Git2 in controlling embryonic cell movements. Git2a knockdown embryos showed defects in cell morphology that were associated with reduced cell contractility. We show that Git2a is required for phosphorylation of myosin light chain (MLC), which regulates myosin II-mediated cell contractility. Consistent with this, embryos treated with Blebbistatin-a small molecule inhibitor for myosin II activity-exhibited cell movement defects similar to git2a knockdown embryos. These observations provide in vivo evidence of a physiologic role for Git2a in regulating cell morphogenesis and directed cell migration via myosin II activation during zebrafish embryonic development.     

Assessment of Femoral Version Should be Assessed Independently of Conventional Measures in Patellofemoral Instability

BackgroundThe purpose of the present study is to determine the association between femoral version and traditional pathologic bony factors commonly used to measure and define patellofemoral alignment.MethodsWe performed a retrospective review of patients treated for patellofemoral instability (PFI) at a single institution. Patients included underwent magnetic resonance imaging (MRI) of the lower extremity using a rotational protocol prior to medial patellofemoral ligament reconstruction with or without tibial tubercle osteotomy. Those with a history of ipsilateral lower extremity surgery were excluded. Two independent reviewers measured femoral version, tibial tubercle-trochlear groove (TT-TG) distance, tibial tubercle-posterior cruciate ligament (TT-PCL) distance, and tibial torsion (TT). Pearson correlation coefficients were used to describe the relationships between all radiographic measures.ResultsA total of 51 knees (43 patients) were included. The average age and body mass index were 23.7 ± 9.33 years and 29.23 ± 8.04 kg/ m2, respectively. The mean femoral version was 15.61 ± 11.57°. The degree of femoral version did not significantly correlate with TT-TG (r=0.103, p=0.474), TT-PCL (-0.086, p=0.550), or TT (r=0.111, p=0.438). Increased TT-TG distance was strongly associated with increased TT-PCL (r=0.470, p=0.001). In females, increased femoral version significantly correlated with increased TT (r=0.381, p=0.029).ConclusionNeither increased nor decreased amounts of femoral anteversion significantly correlated with TT-TG, TT-PCL, or TT. Therefore, assessment of femoral version should be measured independently of conventional measures when considering osteotomies to correct PFI.Level of Evidence: IV