The next generation of training for arabidopsis researchers: Bioinformatics and Quantitative Biology

It has been more than 50 years since Arabidopsis (Arabidopsis thaliana) was first introduced as a model organism to understand basic processes in plant biology. A well-organized scientific community has used this small reference plant species to make numerous fundamental plant biology discoveries (Provart et al., 2016). Due to an extremely well-annotated genome and advances in high-throughput sequencing, our understanding of this organism and other plant species has become even more intricate and complex. Computational resources, including CyVerse,3 Araport,4 The Arabidopsis Information Resource (TAIR),5 and BAR,6 have further facilitated novel findings with just the click of a mouse. As we move toward understanding biological systems, Arabidopsis researchers will need to use more quantitative and computational approaches to extract novel biological findings from these data. Here, we discuss guidelines, skill sets, and core competencies that should be considered when developing curricula or training undergraduate or graduate students, postdoctoral researchers, and faculty. A selected case study provides more specificity as to the concrete issues plant biologists face and how best to address such challenges

Environmental Control of Root System Biology.

The plant root system traverses one of the most complex environments on earth. Understanding how roots support plant life on land requires knowing how soil properties affect the availability of nutrients and water and how roots manipulate the soil environment to optimize acquisition of these resources. Imaging of roots in soil allows the integrated analysis and modeling of environmental interactions occurring at micro- to macroscales. Advances in phenotyping of root systems is driving innovation in cross-platformcompatible methods for data analysis. Root systems acclimate to the environment through architectural changes that act at the root-type level as well as through tissue-specific changes that affect the metabolic needs of the root and the efficiency of nutrient uptake. A molecular understanding of the signaling mechanisms that guide local and systemic signaling is providing insight into the regulatory logic of environmental responses and has identified points where crosstalk between pathways occurs. Expected final online publication date for the Annual Review of Plant Biology Volume 67 is April 29, 2016. Please see http://www.annualreviews.org/catalog/pubdates.aspx for revised estimates

Live Imaging of Root Hairs

Root hairs are single cells specialized in the absorption of water and nutrients. Growing root hairs requires intensive cell wall changes to accommodate cell expansion at the apical end by a process known as tip growth. The cell wall of plants is a very rigid structure comprised largely of polysaccharides and hydroxyproline-rich O-glycoproteins. The importance of root hairs stems from their capacity to expand the surface of interaction between the root and the environment, in search for the necessary nutrients and water to allow plant growth. Therefore, it becomes crucial to deepen our knowledge of them, particularly in the light of the applicability in agriculture by allowing the expansion of croplands. Root hair growth is an extremely fast process, reaching growth rates of up to 1 μm/min and it also is a dynamic process; there can be situations in which the final length might not be affected but the growth rate is. Consequently, in this chapter we focus on a method for studying growth dynamics and rates during a time course. This method is versatile allowing for it to be used in other plant organs such as lateral root, hypocotyl, etc., and also in various conditions.Fil: Velásquez, Silvia Melina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Fisiología, Biología Molecular y Neurociencias. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Fisiología, Biología Molecular y Neurociencias; ArgentinaFil: Dinneny, Jose R.. University of Stanford; Estados UnidosFil: Estevez, Jose Manuel. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Fisiología, Biología Molecular y Neurociencias. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Fisiología, Biología Molecular y Neurociencias; Argentin

Methods to Promote Germination of Dormant <i>Setaria viridis</i> Seeds

<div><p><i>Setaria viridis</i> has recently emerged as a promising genetic model system to study diverse aspects of monocot biology. While the post-germination life cycle of <i>S</i>. <i>viridis</i> is approximately 8 weeks long, the prolonged dormancy of freshly harvested seeds can more than double the total time required between successive generations. Here we describe methods that promote seed germination in <i>S</i>. <i>viridis</i>. Our results demonstrate that treating <i>S</i>. <i>viridis</i> seeds with liquid smoke or a GA₃ and KNO₃ solution improves germination rates to 90% or higher even in seeds that are 6 days post-harvest with similar results obtained whether seeds are planted in soil or on gel-based media. Importantly, we show that these treatments have no significant effect on the growth of the adult plant. We have tested these treatments on diverse <i>S</i>. <i>viridis</i> accessions and show variation in their response. The methods described here will help advance research using this model grass species by increasing the pace at which successive generations of plants can be analyzed.</p></div

Effect of GA₃ and liquid smoke on dormancy in <i>S. viridis</i> seeds.

<p>(<b>A</b>) Dose dependent effect of GA₃ on promoting <i>S. viridis</i> seed germination (seed age 30 dph). (<b>B</b>) Effect of GA₃ and liquid smoke on stimulating germination of <i>S. viridis</i> seeds of different ages (dph) (n = 50 seeds per replicate). (<b>C</b>) An apparent concentration effect of GA₃ in promoting <i>S. viridis</i> seed germination (n = 25 to 30 seeds per replicate, seed age 30 dph). Error bars represent the standard error. LS, liquid smoke; dph, days post-harvesting; GA₃ solution, 2.89 mM GA₃ + 30 mM KNO₃ in distilled water.</p

Liquid smoke promotes <i>S. viridis</i> seed germination.

<p>(<b>A</b>) Dose-dependent effect of liquid smoke on stimulating <i>S. viridis</i> seed germination (n = 25 seeds per replicate, seed age 30 dph). Error bars represent the standard error.</p

Effect of GA₃ and liquid smoke on breaking seed dormancy in different <i>S. viridis</i> accessions.

<p>Effect of GA₃ and liquid smoke on promoting seed germination in 13 different <i>S. viridis</i> accessions (n = 20 to 30 seeds). Graph is plotted using the difference in percent germination observed between the control seeds (incubated in water at 29°C for 24 hours) and GA₃ + KNO₃ or liquid smoke treated seeds (seed age 30 to 65 dph).</p

A microbially derived tyrosine-sulfated peptide mimics a plant peptide hormone

The biotrophic pathogen Xanthomonas oryzae pv. oryzae (Xoo) produces a sulfated peptide named RaxX, which shares similarity to peptides in the PSY (plant peptide containing sulfated tyrosine) family. We hypothesize that RaxX mimics the growth-stimulating activity of PSY peptides. Root length was measured in Arabidopsis and rice treated with synthetic RaxX peptides. We also used comparative genomic analyses and reactive oxygen species burst assays to evaluate the activity of RaxX and PSY peptides. Here we found that a synthetic sulfated RaxX derivative comprising 13 residues (RaxX13-sY), highly conserved between RaxX and PSY, induces root growth in Arabidopsis and rice in a manner similar to that triggered by PSY. We identified residues that are required for activation of immunity mediated by the rice XA21 receptor but that are not essential for root growth induced by PSY. Finally, we showed that a Xanthomonas strain lacking raxX is impaired in virulence. These findings suggest that RaxX serves as a molecular mimic of PSY peptides to facilitate Xoo infection and that XA21 has evolved the ability to recognize and respond specifically to the microbial form of the peptide.This work was supported by NIH GM59962 and NSF IOS1237975. The work conducted by the Joint BioEnergy Institute was supported by the Office of Science, Office of Biological and Environmental Research, of the US Department of Energy under contract no. DE-AC02-05CH1123

Cytokinin functions as an asymmetric and anti-gravitropic signal in lateral roots

Directional organ growth allows the plant root system to strategically cover its surroundings. Intercellular auxin transport is aligned with the gravity vector in the primary root tips, facilitating downward organ bending at the lower rootflank. Here we show that cytokinin signaling functions as a lateral root specific anti-gravitropic component, promoting the radial distribution of the root system. We performed a genome-wide association study and reveal that signal peptide processing of Cytokinin Oxidase 2 (CKX2) affects its enzymatic activity and, thereby, determines the degradation of cytokinins in naturalArabidopsis thaliana accessions. Cytokinin signaling interferes with growth at the upper lateral rootflank and thereby prevents downward bending. Our interdisciplinary approach proposes that two phytohormonal cues at opposite organflanks counterbalance each other’s negative impact on growth, suppressing organ growth towards gravity and allow for radial expansion of the root system

Low sugar is not always good: Impact of specific o-glycan defects on tip growth in arabidopsis

Hydroxyproline (Hyp)-rich O-glycoproteins (HRGPs) comprises several groups of O-glycoproteins including extensins (EXTs), ultimately secreted into plant cell walls. The latter are shaped by several posttranslational modifications (PTMs), mainly hydroxylation of proline residues into hydroxyproline (Hyp) and further O-glycosylation on Hyp and Serine (Ser) (Fig. S1A). EXTs contain several Ser-(Hyp)4 repeats usually O-glycosylated with chains of up to 4-5 linear arabinosyl units on each Hyp (Velasquez et al., 2011; Ogawa-Ohnishi et al., 2013) and mono-galactosylated on Ser residues (Saito et al., 2014). In this context, three groups of arabinosyltransferases (AraTs), HPAT1-HPAT3 (classified as GT8 in the Carbohydrate Active enZymes database [CAZy]), RRA1-RRA3 and XEG113 (GT77 family) have recently been implicated in the sequential addition of the innermost three L-Ara residues (Egelund et al., 2007; Ogawa-Ohnishi et al., 2013). In addition, one novel peptidyl-Ser galactosyltransferase named SERGT1 has been reported to add a single -Galp residue to each Ser residue in Ser-(Hyp)4 motifs of EXTs, thus belonging to a new family within CAZy (Table S1). Finally, glycosylated EXTs are possibly crosslinked by putative type-III peroxidases (PERs) at the Tyr residues by forming EXT linkages (Cannon et al., 2008) able to build a three-dimensional network likely to interact with other cell wall components like pectins (Cannon et al., 2008). Here, by using appropriate mutants of several known enzymes of the O-glycosylation pathway of HRGPs, we addressed to what extent each single defect on the O-glycosylation machinery impacts on root hair tip growth.Fil: Velásquez, Silvia Melina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Fisiología, Biología Molecular y Neurociencias. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Fisiología, Biología Molecular y Neurociencias; ArgentinaFil: Marzol, Eliana. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Fisiología, Biología Molecular y Neurociencias. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Fisiología, Biología Molecular y Neurociencias; ArgentinaFil: Borassi, Cecilia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Fisiología, Biología Molecular y Neurociencias. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Fisiología, Biología Molecular y Neurociencias; ArgentinaFil: Pol-Fachin, Laercio. Universidade Federal de Pernambuco; Brasil. Universidade Federal do Rio Grande do Sul; BrasilFil: Ricardi, Martiniano María. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Fisiología, Biología Molecular y Neurociencias. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Fisiología, Biología Molecular y Neurociencias; ArgentinaFil: Mangano, Silvina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Fisiología, Biología Molecular y Neurociencias. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Fisiología, Biología Molecular y Neurociencias; ArgentinaFil: Denita Juárez, Silvina Paola. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Fisiología, Biología Molecular y Neurociencias. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Fisiología, Biología Molecular y Neurociencias; ArgentinaFil: Salgado Salter, Juan David. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Fisiología, Biología Molecular y Neurociencias. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Fisiología, Biología Molecular y Neurociencias; ArgentinaFil: Gloazzo Dorosz, Javier Anselmo. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Fisiología, Biología Molecular y Neurociencias. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Fisiología, Biología Molecular y Neurociencias; ArgentinaFil: Marcus, Susan E.. University of Leeds; Reino UnidoFil: Knox, J. Paul. University of Leeds; Reino UnidoFil: Dinneny, Jose R.. Carnegie Institution for Science. Department Of Plant Biology; Estados UnidosFil: Iusem, Norberto Daniel. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Fisiología, Biología Molecular y Neurociencias. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Fisiología, Biología Molecular y Neurociencias; ArgentinaFil: Verli, Hugo. Universidade Federal do Rio Grande do Sul; BrasilFil: Estevez, Jose Manuel. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Fisiología, Biología Molecular y Neurociencias. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Fisiología, Biología Molecular y Neurociencias; Argentin