Category: Publications

Migeon A, Blaudez D, Wilkins O, Montanini B, Campbell MM, Richaud P, Thomine S, Chalot M

Cell. Mol. Life Sci. 2010 Nov;67(22):3763-84

PubMed PMID: 20623158

Abstract

The specific transport of metal ions, mediated by membrane-localized metal transporters, is of fundamental importance in all eukaryotes. Genome-wide analysis of metal transporters was undertaken, making use of whole genome sequences of the green alga Chlamydomonas reinhardtii, the moss Physcomitrella patens, the lycophyte Selaginella moellendorffii, the monocots rice and sorghum, and the dicots Arabidopsis thaliana, poplar, grapevine, as well as of the yeast Saccharomyces cerevisiae. A repertoire of 430 metal transporters was found in total across eight photosynthetic plants, as well as in S. cerevisiae. Seventy-two full-length metal transporter genes were identified in the Populus genome alone, which is the largest number of metal transporters genes identified in any single species to date. Diversification of some transporter family gene clusters appears to have occurred in a lineage-specific manner. Expression analysis of Populus metal transporters indicates that some family members show tissue-specific transcript abundance. Taken together, the data provide a picture into the diversification of these important gene families.

Wilkins O, Bräutigam K, Campbell MM

Plant J. 2010 Sep;63(5):715-27

PubMed PMID: 20553421

Abstract

Under natural conditions, it is common for plants to experience water deprivation (drought) for periods of days or longer. Plants respond to drought stress by reconfiguring their transcriptome activity. Transcriptome changes in response to drought are dynamic, and are shaped by mitigating factors like time during the diurnal cycle. To date, analyses of drought-induced transcriptome remodelling have concentrated on dynamic changes induced by rapid desiccation, or changes at a single time point following gradual water stress. To gain insights into the dynamics of transcriptome reconfiguration in response to gradual drying of the soil, the drought-induced transcriptomes of Arabidopsis thaliana were examined at four time points over a single diel period – midday, late day, midnight, and pre-dawn. Transcriptome reconfigurations were induced by drought in advance of changes to relative water content, leaf water loss, and chlorophyll content. Comparative analyses support the hypothesis that the drought-responsive transcriptomes were shaped by invocation of distinct hormonal and stress response pathways at different times of the day. While a core set of genes were drought responsive at multiple time points throughout the day, the magnitude of the response varied in a manner dependent on the time of day. Moreover, analysis of a single time point would fail to identify suites of drought-responsive genes that can only be detected through assessment of the dynamics of diurnal changes, emphasising the value of characterising multiple time-of-day-specific drought transcriptomes.

Hamanishi ET, Raj S, Wilkins O, Thomas BR, Mansfield SD, Plant AL, Campbell MM

Plant Cell Environ. 2010 Oct;33(10):1742-55

PubMed PMID: 20525001

Abstract

Drought is a major limitation to the growth and productivity of trees in the ecologically and economically important genus Populus. The ability of Populus trees to contend with drought is a function of genome responsiveness to this environmental insult, involving reconfiguration of the transcriptome to appropriately remodel growth, development and metabolism. Here we test hypotheses aimed at examining the extent of intraspecific variation in the drought transcriptome using six different Populus balsamifera L. genotypes and Affymetrix GeneChip technology. Within a given genotype there was a positive correlation between the magnitude of water-deficit induced changes in transcript abundance across the transcriptome, and the capacity of that genotype to maintain growth following water deficit. Genotypes that had more similar drought-responsive transcriptomes also had fewer genotypic differences, as determined by microarray-derived single feature polymorphism (SFP) analysis, suggesting that responses may be conserved across individuals that share a greater degree of genotypic similarity. This work highlights the fact that a core species-level response can be defined; however, the underpinning genotype-derived complexities of the drought response in Populus must be taken into consideration when defining both species- and genus-level responses.

Cutter AD, Choi JY

Genome Res. 2010 Aug;20(8):1103-11

PubMed PMID: 20508143

Abstract

The combined actions of natural selection, mutation, and recombination forge the landscape of genetic variation across genomes. One frequently observed manifestation of these processes is a positive association between neutral genetic variation and local recombination rates. Two selective mechanisms and/or recombination-associated mutation (RAM) could generate this pattern, and the relative importance of these alternative possibilities remains unresolved generally. Here we quantify nucleotide differences within populations, between populations, and between species to test for genome-wide effects of selection and RAM in the partially selfing nematode Caenorhabditis briggsae. We find that nearly half of genome-wide variation in nucleotide polymorphism is explained by differences in local recombination rates. By quantifying divergence between several reproductively isolated lineages, we demonstrate that ancestral polymorphism generates a spurious signal of RAM for closely related lineages, with implications for analyses of humans and primates; RAM is, at most, a minor factor in C. briggsae. We conclude that the positive relation between nucleotide polymorphism and the rate of crossover represents the footprint of natural selection across the C. briggsae genome and demonstrate that background selection against deleterious mutations is sufficient to explain this pattern. Hill-Robertson interference also leaves a signature of more effective purifying selection in high-recombination regions of the genome. Finally, we identify an emerging contrast between widespread adaptive hitchhiking effects in species with large outcrossing populations (e.g., Drosophila) versus pervasive background selection effects on the genomes of organisms with self-fertilizing lifestyles and/or small population sizes (e.g., Caenorhabditis elegans, C. briggsae, Arabidopsis thaliana, Lycopersicon, human). These results illustrate how recombination, mutation, selection, and population history interact in important ways to shape molecular heterogeneity within and between genomes.

Wilton M, Desveaux D

Plant Signal Behav 2010 Jun;5(6):746-8

PubMed PMID: 20505348

Abstract

The Gram negative bacterial phytopathogen Pseudomonas syringae employs a molecular syringe termed the Type III secretion system (TTSS) to deliver an array of Type III secreted effector (TTSE) proteins into plant cells. The major function ascribed to type III effectors of P. syringae is their ability to suppress plant immunity. Because individual pathovars of P. syringae can possess over 30 TTSEs, functional redundancy can provide a hurdle to ascribing functions by TTSE-deletion or -overexpression in such TTSE-rich backgrounds. Approaches to overcome functional redundancy have included the deletion of multiple TTSEs from individual pathovars as well as engineering the plant commensal P. fluorescens strain to express the P. syringae TTSS and deliver P. syringae TTSEs. As we describe here, transgenic Arabidopsis plants expressing individual TTSEs have also be used to overcome problems of functional redundancy and provide invaluable insights into TTSE virulence functions.

Dang LT, Tropepe V

Neural Dev 2010;5:13

PubMed PMID: 20459606

Abstract

BACKGROUND: Mouse definitive neural stem cells (NSCs) are derived from a population of LIF-responsive primitive neural stem cells (pNSCs) within the neurectoderm, yet details on the early signaling and transcriptional mechanisms that control this lineage transition are lacking. Here we tested whether FGF and Wnt signaling pathways can regulate Zfhx1b expression to control early neural stem cell development.

RESULTS: By microinjecting FGF8b into the pro-amniotic cavity ex vivo at 7.0 days post-coitum (dpc) and culturing whole embryos, we demonstrate that neurectoderm-specific gene expression (for example, Sox2, Nestin, Zfhx1b) is increased, whereas Wnt3a represses neurectoderm gene expression. To determine whether FGF signaling also mediates the lineage transition from a pNSC to a NSC, 7.0-dpc embryos were microinjected with either FGF8b or inhibitors of the FGF receptor-MAP kinase signaling pathway ex vivo, cultured as whole embryos to approximately 8.5 dpc and assayed for clonal NSC colony formation. We show that pre-activation of FGF signaling in the anterior neurectoderm causes an increase in the number of colony forming NSCs derived later from the anterior neural plate, whereas inhibition of FGF signaling significantly reduces the number of NSC colonies. Interestingly, inhibition of FGF signaling causes the persistence of LIF-responsive pNSCs within the anterior neural plate and over-expression of Zfhx1b in these cells is sufficient to rescue the transition from a LIF-responsive pNSC to an FGF-responsive NSC.

CONCLUSION: Our data suggest that definitive NSC fate specification in the mouse neurectoderm is facilitated by FGF activation of Zfhx1b.

Cutter AD, Agrawal AF

Genetics 2010 Jun;185(2):685-93

PubMed PMID: 20382830

Abstract

Genes in nematode and ascidian genomes frequently occur in operons–multiple genes sharing a common promoter to generate a polycistronic primary transcript–and such genes comprise 15-20% of the coding genome for Caenorhabditis elegans and Ciona intestinalis. Recent work in nematodes has demonstrated that the identity of genes within operons is highly conserved among species and that the unifying feature of genes within operons is that they are expressed in germline tissue. However, it is generally unknown what processes are responsible for generating the distribution of operon sizes across the genome, which are composed of up to eight genes per operon. Here we investigate several models for operon evolution to better understand their abundance, distribution of sizes, and evolutionary dynamics over time. We find that birth-death models of operon evolution reasonably describe the relative abundance of operons of different sizes in the C. elegans and Ciona genomes and generate predictions about the number of monocistronic, nonoperon genes that likely participate in the birth-death process. This theory, and applications to C. elegans and Ciona, motivates several new and testable hypotheses about eukaryote operon evolution.

Chin K, Moeder W, Abdel-Hamid H, Shahinas D, Gupta D, Yoshioka K

J. Exp. Bot. 2010 May;61(9):2383-93

PubMed PMID: 20378667

Abstract

The involvement of cyclic nucleotide gated ion channels (CNGCs) in the signal transduction of animal light and odorant perception is well documented. Although plant CNGCs have recently been revealed to mediate multiple stress responses and developmental pathways, studies that aim to elucidate their structural and regulatory properties are still very much in their infancy. The structure-function relationship of plant CNGCs was investigated here by using the chimeric Arabidopsis AtCNGC11/12 gene that induces multiple defence responses in the Arabidopsis mutant constitutive expresser of PR genes 22 (cpr22) for the identification of functionally essential residues. A genetic screen for mutants that suppress cpr22-conferred phenotypes identified over 20 novel mutant alleles in AtCNGC11/12. One of these mutants, suppressor S58 possesses a single amino acid substitution, arginine 557 to cysteine, in the alphaC-helix of the cyclic nucleotide-binding domain (CNBD). The suppressor S58 lost all cpr22 related phenotypes, such as spontaneous cell death formation under ambient temperature conditions. However, these phenotypes were recovered at 16 degrees C suggesting that the stability of channel function is affected by temperature. In silico modelling and site-directed mutagenesis analyses suggest that arginine 557 in the alphaC-helix of the CNBD is important for channel regulation, but not for basic function. Furthermore, another suppressor mutant, S136 that lacks the entire alphaC-helix due to a premature stop codon, lost channel function completely. Our data presented here indicate that the alphaC-helix is functionally important in plant CNGCs.

Lewis JD, Wu R, Guttman DS, Desveaux D

PLoS Genet. 2010 Apr;6(4):e1000894

PubMed PMID: 20368970

Abstract

Plant resistance (R) proteins provide a robust surveillance system to defend against potential pathogens. Despite their importance in plant innate immunity, relatively few of the approximately 170 R proteins in Arabidopsis have well-characterized resistance specificity. In order to identify the R protein responsible for recognition of the Pseudomonas syringae type III secreted effector (T3SE) HopZ1a, we assembled an Arabidopsis R gene T-DNA Insertion Collection (ARTIC) from publicly available Arabidopsis thaliana insertion lines and screened it for plants lacking HopZ1a-induced immunity. This reverse genetic screen revealed that the Arabidopsis R protein HOPZ-activated resistance 1 (ZAR1; At3g50950) is required for recognition of HopZ1a in Arabidopsis. ZAR1 belongs to the coiled-coil (CC) class of nucleotide binding site and leucine-rich repeat (NBS-LRR) containing R proteins; however, the ZAR1 CC domain phylogenetically clusters in a clade distinct from other related Arabidopsis R proteins. ZAR1-mediated immunity is independent of several genes required by other R protein signaling pathways, including NDR1 and RAR1, suggesting that ZAR1 possesses distinct signaling requirements. The closely-related T3SE protein, HopZ1b, is still recognized by zar1 Arabidopsis plants indicating that Arabidopsis has evolved at least two independent R proteins to recognize the HopZ T3SE family. Also, in Arabidopsis zar1 plants HopZ1a promotes P. syringae growth indicative of an ancestral virulence function for this T3SE prior to the evolution of recognition by the host resistance protein ZAR1. Our results demonstrate that the Arabidopsis resistance protein ZAR1 confers allele-specific recognition and virulence attenuation of the Pseudomonas syringae T3SE protein HopZ1a.

Ba AN, Moses AM

Mol. Biol. Evol. 2010 Sep;27(9):2027-37

PubMed PMID: 20368267

Abstract

Phosphorylation is one of the most studied and important regulatory mechanisms that modulate protein function in eukaryotic cells. Recently, several studies have investigated the evolution of phosphorylation sites identified by high-throughput methods. These studies have revealed varying degrees of evidence for constraint and plasticity, and therefore, there is currently no consensus as to the evolutionary properties of this important regulatory mechanism. Here, we present a study of high-confidence annotated sites from budding yeast and show that these sites are significantly constrained compared with their flanking region in closely related species. We show that this property does not change in structured or unstructured regions. We investigate the birth, death and compensation rates of the phosphorylation sites and test if sites are more likely to be gained or lost in proteins with greater numbers of sites. Finally, we also show that this evolutionary conservation can yield significant improvement for kinase target predictions when the kinase recognition motif is known, and can be used to infer the recognition motif when a set of targets is known. Our analysis indicates that phosphorylation sites are under selective constraint, consistent with their functional importance. We also find that a small fraction of phosphorylation sites turnover during evolution, which may be an important process underlying the evolution of regulatory networks.