Category: Publications

A Practical Guide to Quantitative Interactor Screening with Next-Generation Sequencing (QIS-Seq)

Posted on by biota in Publications

Gong Y, Desveaux D, Guttman DS, Lewis JD

Methods Mol Biol. Published Aug 2017.
https://doi.org/10.1007/978-1-4939-7027-8_1

PMID: 28849555

Abstract

Yeast two-hybrid screens are a powerful approach to identify protein-protein interactions; however, they are typically limited in the number of interactions identified, and lack quantitative values to ascribe confidence scores to the interactions that are obtained. We have developed a high-throughput, quantitative, yeast two-hybrid screening approach coupled with next-generation sequencing. This strategy allows the identification of interacting proteins that are preferentially associated with a bait of interest, and helps eliminate nonspecific interacting proteins. The method is high-throughput, allowing many more baits to be tested and many more candidate interacting proteins to be identified. Quantitative data allows the interactors to be ascribed confidence scores based on their enrichment with particular baits, and can identify both common and rare interacting proteins.

Draft Genome Sequences of 10 Environmental Pseudomonas aeruginosa Strains Isolated from Soils, Sediments, and Waters

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Shrestha SD, Guttman DS, Perron GG

Genome Announc. Published Aug 24, 2017.
https://doi.org/10.1128/genomeA.00804-17

PMID: 28839021

Abstract

Pseudomonas aeruginosa is an important opportunistic pathogen that has the ability to grow in a range of environmental niches. Here, we report the draft genome sequences of 10 environmental strains of the bacterium isolated from soils, sediments, and waters in various locations in North America and South Africa.

Expression profiles of miRNAs from bovine mammary glands in response to Streptococcus agalactiae-induced mastitis

Posted on by biota in Publications

Pu J, Li R, Zhang C, Chen D, Liao X, Zhu Y, Geng X, Ji D, Mao Y, Gong Y, Yang Z

J Dairy Res. Published Aug 2017.
https://doi.org/10.1017/S0022029917000437

PMID: 28831974

Abstract

This study aimed to describe the expression profiles of microRNAs (miRNAs) from mammary gland tissues collected from dairy cows with Streptococcus agalactiae-induced mastitis and to identify differentially expressed miRNAs related to mastitis. The mammary glands of Chinese Holstein cows were challenged with Streptococcus agalactiae to induce mastitis. Small RNAs were isolated from the mammary tissues of the test and control groups and then sequenced using the Solexa sequencing technology to construct two small RNA libraries. Potential target genes of these differentially expressed miRNAs were predicted using the RNAhybrid software, and KEGG pathways associated with these genes were analysed. A total of 18 555 913 and 20 847 000 effective reads were obtained from the test and control groups, respectively. In total, 373 known and 399 novel miRNAs were detected in the test group, and 358 known and 232 novel miRNAs were uncovered in the control group. A total of 35 differentially expressed miRNAs were identified in the test group compared to the control group, including 10 up-regulated miRNAs and 25 down-regulated miRNAs. Of these miRNAs, miR-223 exhibited the highest degree of up-regulation with an approximately 3-fold increase in expression, whereas miR-26a exhibited the most decreased expression level (more than 2-fold). The RNAhybrid software predicted 18 801 genes as potential targets of these 35 miRNAs. Furthermore, several immune response and signal transduction pathways, including the RIG-I-like receptor signalling pathway, cytosolic DNA sensing pathway and Notch signal pathway, were enriched in these predicted targets. In summary, this study provided experimental evidence for the mechanism underlying the regulation of bovine mastitis by miRNAs and showed that miRNAs might be involved in signal pathways during S. agalactiae-induced mastitis.

ePlant: Visualizing and Exploring Multiple Levels of Data for Hypothesis Generation in Plant Biology

Posted on by biota in Publications

Waese J, Fan J, Pasha A, Yu H, Fucile G, Shi R, Cumming M, Kelley LA, Sternberg MJ, Krishnakumar V, Ferlanti E, Miller J, Town C, Stuerzlinger W, Provart NJ

Plant Cell. Published Aug 2017.
https://doi.org/10.1105/tpc.17.00073

PMID: 28808136

Abstract

A big challenge in current systems biology research arises when different types of data must be accessed from separate sources and visualized using separate tools. The high cognitive load required to navigate such a workflow is detrimental to hypothesis generation. Accordingly, there is a need for a robust research platform that incorporates all data and provides integrated search, analysis, and visualization features through a single portal. Here, we present ePlant (http://bar.utoronto.ca/eplant), a visual analytic tool for exploring multiple levels of Arabidopsis thaliana data through a zoomable user interface. ePlant connects to several publicly available web services to download genome, proteome, interactome, transcriptome, and 3D molecular structure data for one or more genes or gene products of interest. Data are displayed with a set of visualization tools that are presented using a conceptual hierarchy from big to small, and many of the tools combine information from more than one data type. We describe the development of ePlant in this article and present several examples illustrating its integrative features for hypothesis generation. We also describe the process of deploying ePlant as an “app” on Araport. Building on readily available web services, the code for ePlant is freely available for any other biological species research.

Chemical genetics and strigolactone perception

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Lumba S, Bunsick M, McCourt P

F1000Res. Published Jun 22, 2017.
https://doi.org/10.12688/f1000research.11379.1

PMID: 28690842

Abstract

Strigolactones (SLs) are a collection of related small molecules that act as hormones in plant growth and development. Intriguingly, SLs also act as ecological communicators between plants and mycorrhizal fungi and between host plants and a collection of parasitic plant species. In the case of mycorrhizal fungi, SLs exude into the soil from host roots to attract fungal hyphae for a beneficial interaction. In the case of parasitic plants, however, root-exuded SLs cause dormant parasitic plant seeds to germinate, thereby allowing the resulting seedling to infect the host and withdraw nutrients. Because a laboratory-friendly model does not exist for parasitic plants, researchers are currently using information gleaned from model plants like Arabidopsis in combination with the chemical probes developed through chemical genetics to understand SL perception of parasitic plants. This work first shows that understanding SL signaling is useful in developing chemical probes that perturb SL perception. Second, it indicates that the chemical space available to probe SL signaling in both model and parasitic plants is sizeable. Because these parasitic pests represent a major concern for food insecurity in the developing world, there is great need for chemical approaches to uncover novel lead compounds that perturb parasitic plant infections.

Evolution of nonspectral rhodopsin function at high altitudes

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Castiglione GM, Hauser FE, Liao BS, Lujan NK, Van Nynatten A, Morrow JM, Schott RK, Bhattacharyya N, Dungan SZ, Chang BSW

Proc Natl Acad Sci USA. Published Jul 11, 2017.
https://doi.org/10.1073/pnas.1705765114

PMID: 28642345

Abstract

High-altitude environments present a range of biochemical and physiological challenges for organisms through decreases in oxygen, pressure, and temperature relative to lowland habitats. Protein-level adaptations to hypoxic high-altitude conditions have been identified in multiple terrestrial endotherms; however, comparable adaptations in aquatic ectotherms, such as fishes, have not been as extensively characterized. In enzyme proteins, cold adaptation is attained through functional trade-offs between stability and activity, often mediated by substitutions outside the active site. Little is known whether signaling proteins [e.g., G protein-coupled receptors (GPCRs)] exhibit natural variation in response to cold temperatures. Rhodopsin (RH1), the temperature-sensitive visual pigment mediating dim-light vision, offers an opportunity to enhance our understanding of thermal adaptation in a model GPCR. Here, we investigate the evolution of rhodopsin function in an Andean mountain catfish system spanning a range of elevations. Using molecular evolutionary analyses and site-directed mutagenesis experiments, we provide evidence for cold adaptation in RH1. We find that unique amino acid substitutions occur at sites under positive selection in high-altitude catfishes, located at opposite ends of the RH1 intramolecular hydrogen-bonding network. Natural high-altitude variants introduced into these sites via mutagenesis have limited effects on spectral tuning, yet decrease the stability of dark-state and light-activated rhodopsin, accelerating the decay of ligand-bound forms. As found in cold-adapted enzymes, this phenotype likely compensates for a cold-induced decrease in kinetic rates-properties of rhodopsin that mediate rod sensitivity and visual performance. Our results support a role for natural variation in enhancing the performance of GPCRs in response to cold temperatures.

Expression atlas and comparative coexpression network analyses reveal important genes involved in the formation of lignified cell wall in Brachypodium distachyon

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Sibout R, Proost S, Hansen BO, Vaid N, Giorgi FM, Ho-Yue-Kuang S, Legée F, Cézart L, Bouchabké-Coussa O, Soulhat C, Provart N, Pasha A, Le Bris P, Roujol D, Hofte H, Jamet E, Lapierre C, Persson S, Mutwil M

New Phytol. Published Aug, 2017.
https://doi.org/10.1111/nph.14635

PMID: 28617955

Abstract

While Brachypodium distachyon (Brachypodium) is an emerging model for grasses, no expression atlas or gene coexpression network is available. Such tools are of high importance to provide insights into the function of Brachypodium genes. We present a detailed Brachypodium expression atlas, capturing gene expression in its major organs at different developmental stages. The data were integrated into a large-scale coexpression database ( www.gene2function.de), enabling identification of duplicated pathways and conserved processes across 10 plant species, thus allowing genome-wide inference of gene function. We highlight the importance of the atlas and the platform through the identification of duplicated cell wall modules, and show that a lignin biosynthesis module is conserved across angiosperms. We identified and functionally characterised a putative ferulate 5-hydroxylase gene through overexpression of it in Brachypodium, which resulted in an increase in lignin syringyl units and reduced lignin content of mature stems, and led to improved saccharification of the stem biomass. Our Brachypodium expression atlas thus provides a powerful resource to reveal functionally related genes, which may advance our understanding of important biological processes in grasses.

The expanding role of the Ehmt2/G9a complex in neurodevelopment

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Deimling SJ, Olsen JB, Tropepe V

Neurogenesis (Austin). Published May 2, 2017.
https://doi.org/10.1080/23262133.2017.1316888

PMID: 28596979

Abstract

Epigenetic regulators play a crucial role in neurodevelopment. One such epigenetic complex, Ehmt1/2 (G9a/GLP), is essential for repressing gene transcription by methylating H3K9 in a highly tissue- and temporal-specific manner. Recently, data has emerged suggesting that this complex plays additional roles in regulating the activity of numerous other non-histone proteins. While much is known about the downstream effects of Ehmt1/2 function, evidence is only beginning to come to light suggesting the control of Ehmt1/2 function may be, at least in part, due to context-dependent binding partners. Here we review emerging roles for the Ehmt1/2 complex suggesting that it may play a much larger role than previously recognized, and discuss binding partners that we and others have recently characterized which act to coordinate its activity during early neurodevelopment.

The perception of strigolactones in vascular plants

Posted on by biota in Publications

Lumba S, Holbrook-Smith D, McCourt P

Nat Chem Biol. Published May 17, 2017.
https://doi.org/10.1038/nchembio.2340

PMID: 28514432

Abstract

Small-molecule hormones play central roles in plant development, ranging from cellular differentiation and organ formation to developmental response instruction in changing environments. A recently discovered collection of related small molecules collectively called strigolactones are of particular interest, as these hormones also function as ecological communicators between plants and fungi and between parasitic plants and their hosts. Advances from model plant systems have begun to unravel how, as a hormone, strigolactone is perceived and transduced. In this Review, we summarize this information and examine how understanding strigolactone hormone signaling is leading to insights into parasitic plant infections. We specifically focus on how the development of chemical probes can be used in combination with model plant systems to dissect strigolactone’s perception in the parasitic plant Striga hermonthica. This information is particularly relevant since Striga is considered one of the largest impediments to food security in sub-Saharan Africa.

Found in Translation: Applying Lessons from Model Systems to Strigolactone Signaling in Parasitic Plants

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Lumba S, Subha A, McCourt P

Trends Biochem Sci. Published July, 2017.
https://doi.org/10.1016/j.tibs.2017.04.006

PMID: 28495334

Abstract

Strigolactones (SLs) are small molecules that act as endogenous hormones to regulate plant development as well as exogenous cues that help parasitic plants to infect their hosts. Given that parasitic plants are experimentally challenging systems, researchers are using two approaches to understand how they respond to host-derived SLs. The first involves extrapolating information on SLs from model genetic systems to dissect their roles in parasitic plants. The second uses chemicals to probe SL signaling directly in the parasite Striga hermonthica. These approaches indicate that parasitic plants have co-opted a family of α/β hydrolases to perceive SLs. The importance of this genetic and chemical information cannot be overstated since parasitic plant infestations are major obstacles to food security in the developing world.