Publications

Chemical genetics and strigolactone perception

Posted on by biota in Publications

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.

Using GCaMP3 to Study Ca2+ Signaling in Nicotiana Species

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DeFalco TA, Toyota M, Phan V, Karia P, Moeder W, Gilroy S, Yoshioka K

Plant Cell Physiol. Published Jul 1, 2017.
https://doi.org/10.1093/pcp/pcx053

PMID: 28482045

Abstract

Ca2+ signaling is a central component of plant biology; however, direct analysis of in vivo Ca2+ levels is experimentally challenging. In recent years, the use of genetically encoded Ca2+ indicators has revolutionized the study of plant Ca2+ signaling, although such studies have been largely restricted to the model plant Arabidopsis. We have developed stable transgenic Nicotiana benthamiana and Nicotiana tabacum lines expressing the single-wavelength fluorescent Ca2+ indicator, GCaMP3. Ca2+ levels in these plants can be imaged in situ using fluorescence microscopy, and these plants can be used qualitatively and semi-quantitatively to evaluate Ca2+ signals in response to a broad array of abiotic or biotic stimuli, such as cold shock or pathogen-associated molecular patterns (PAMPs). Furthermore, these tools can be used in conjunction with well-established N. benthamiana techniques such as virus-induced gene silencing (VIGS) or transient heterologous expression to assay the effects of loss or gain of function on Ca2+ signaling, an approach which we validated via silencing or transient expression of the PAMP receptors FLS2 (Flagellin Sensing 2) or EFR (EF-Tu receptor), respectively. Using these techniques, along with chemical inhibitor treatments, we demonstrate how these plants can be used to elucidate the molecular components governing Ca2+ signaling in response to specific stimuli.

Cone-like rhodopsin expressed in the all-cone retina of the colubrid pine snake as a potential adaptation to diurnality

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Bhattacharyya N, Darren B, Schott RK, Tropepe V, Chang BSW

J Exp Biol. Published Jul 1, 2017.
https://doi.org/10.1242/jeb.156430

PMID: 28468872

Abstract

Colubridae is the largest and most diverse family of snakes, with visual systems that reflect this diversity, encompassing a variety of retinal photoreceptor organizations. The transmutation theory proposed by Walls postulates that photoreceptors could evolutionarily transition between cell types in squamates, but few studies have tested this theory. Recently, evidence for transmutation and rod-like machinery in an all-cone retina has been identified in a diurnal garter snake (Thamnophis), and it appears that the rhodopsin gene at least may be widespread among colubrid snakes. However, functional evidence supporting transmutation beyond the existence of the rhodopsin gene remains rare. We examined the all-cone retina of another colubrid, Pituophis melanoleucus, thought to be more secretive/burrowing than Thamnophis We found that P. melanoleucus expresses two cone opsins (SWS1, LWS) and rhodopsin (RH1) within the eye. Immunohistochemistry localized rhodopsin to the outer segment of photoreceptors in the all-cone retina of the snake and all opsin genes produced functional visual pigments when expressed in vitro Consistent with other studies, we found that P. melanoleucus rhodopsin is extremely blue-shifted. Surprisingly, P. melanoleucus rhodopsin reacted with hydroxylamine, a typical cone opsin characteristic. These results support the idea that the rhodopsin-containing photoreceptors of P. melanoleucus are the products of evolutionary transmutation from rod ancestors, and suggest that this phenomenon may be widespread in colubrid snakes. We hypothesize that transmutation may be an adaptation for diurnal, brighter-light vision, which could result in increased spectral sensitivity and chromatic discrimination with the potential for colour vision.

Topo-phylogeny: Visualizing evolutionary relationships on a topographic landscape

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Waese J, Provart NJ, Guttman DS

PLoS One. Published May 1, 2017
https://doi.org/10.1371/journal.pone.0175895

PMID: 28459802

Abstract

Phylogenetic trees are the de facto standard for visualizing evolutionary relationships, but large trees can be difficult to interpret because they require a high cognitive load to identify relationships between multiple operational taxonomic units (OTUs). We present a new tool for displaying phylogenetic relationships as a topographic map in which OTUs autonomously attract or repel one another based on their individual branch lengths and distance to a common ancestor. This data visualization paradigm makes it possible to preattentively identify the nature of the relationship between items without having to trace a complex network of branches back to the root. This tool was developed for exploring phylogenetic data, but the technique could be extended for visualizing other hierarchical structures as well.

Calmodulin as a Ca2+-Sensing Subunit of Arabidopsis Cyclic Nucleotide-Gated Channel Complexes

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Fischer C, DeFalco TA, Karia P, Snedden WA, Moeder W, Yoshioka K, Dietrich P

Plant Cell Physiol. Published Jul 1, 2017
https://doi.org/10.1093/pcp/pcx052

PMID: 28419310

Abstract

Ca2+ serves as a universal second messenger in eukaryotic signaling pathways, and the spatial and temporal patterns of Ca2+ concentration changes are determined by feedback and feed-forward regulation of the involved transport proteins. Cyclic nucleotide-gated channels (CNGCs) are Ca2+-permeable channels that interact with the ubiquitous Ca2+ sensor calmodulin (CaM). CNGCs interact with CaMs via diverse CaM-binding sites, including an IQ-motif, which has been identified in the C-termini of CNGC20 and CNGC12. Here we present a family-wide analysis of the IQ-motif from all 20 Arabidopsis CNGC isoforms. While most of their IQ-peptides interacted with conserved CaMs in yeast, some were unable to do so, despite high sequence conservation across the family. We showed that the CaM binding ability of the IQ-motif is highly dependent on its proximal and distal vicinity. We determined that two alanine residues positioned N-terminal to the core IQ-sequence play a significant role in CaM binding, and identified a polymorphism at this site that promoted or inhibited CaM binding in yeast. Through detailed biophysical analysis of the CNGC2 IQ-motif, we found that this polymorphism specifically affected the Ca2+-independent interactions with the C-lobe of CaM. This same polymorphism partially suppressed the induction of programmed cell death by CNGC11/12 in planta. Our work expands the model of CNGC regulation, and posits that the C-lobe of apo-CaM is permanently associated with the channel at the N-terminal part of the IQ-domain. This mode allows CaM to function as a Ca2+-sensing regulatory subunit of the channel complex, providing a mechanism by which Ca2+ signals may be fine-tuned.