Arabidopsis protein phosphatase DBP1 nucleates a protein network with a role in regulating plant defense

Arabidopsis thaliana DBP1 belongs to the plant-specific family of DNA-binding protein phosphatases. Although recently identified as a novel host factor mediating susceptibility to potyvirus, little is known about DBP1 targets and partners and the molecular mechanisms underlying its function. Analyzing changes in the phosphoproteome of a loss-of-function dbp1 mutant enabled the identification of 14-3-3l isoform (GRF6), a previously reported DBP1 interactor, and MAP kinase (MAPK) MPK11 as components of a small protein network nucleated by DBP1, in which GRF6 stability is modulated by MPK11 through phosphorylation, while DBP1 in turn negatively regulates MPK11 activity. Interestingly, grf6 and mpk11 loss-offunction mutants showed altered response to infection by the potyvirus Plum pox virus (PPV), and the described molecular mechanism controlling GRF6 stability was recapitulated upon PPV infection. These results not only contribute to a better knowledge of the biology of DBP factors, but also of MAPK signalling in plants, with the identification of GRF6 as a likely MPK11 substrate and of DBP1 as a protein phosphatase regulating MPK11 activity, and unveils the implication of this protein module in the response to PPV infection in Arabidopsis.This work was supported by the Spanish MICINN (Grants BFU2009-09771, EUI2009-04009 to PV), Generalitat Valenciana (Prometeo2010/020 to PV) and the German DFG (SCHE 235/15-1 to DS). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.Carrasco Jiménez, JL.; Castelló Llopis, MJ.; Naumann, K.; Lassowskat, I.; Navarrete Gomez, ML.; Scheel, D.; Vera Vera, P. (2014). Arabidopsis protein phosphatase DBP1 nucleates a protein network with a role in regulating plant defense. PLoS ONE. 9:1-10. https://doi.org/10.1371/journal.pone.0090734S1109Carrasco, J. L. (2003). A novel transcription factor involved in plant defense endowed with protein phosphatase activity. The EMBO Journal, 22(13), 3376-3384. doi:10.1093/emboj/cdg323Carrasco, J. L., Ancillo, G., Castelló, M. J., & Vera, P. (2005). A Novel DNA-Binding Motif, Hallmark of a New Family of Plant Transcription Factors. Plant Physiology, 137(2), 602-606. doi:10.1104/pp.104.056002Castelló, M. J., Carrasco, J. L., & Vera, P. (2010). DNA-Binding Protein Phosphatase AtDBP1 Mediates Susceptibility to Two Potyviruses in Arabidopsis. Plant Physiology, 153(4), 1521-1525. doi:10.1104/pp.110.158923Castelló, M. J., Carrasco, J. L., Navarrete-Gómez, M., Daniel, J., Granot, D., & Vera, P. (2011). A Plant Small Polypeptide Is a Novel Component of DNA-Binding Protein Phosphatase 1-Mediated Resistance to Plum pox virus in Arabidopsis. Plant Physiology, 157(4), 2206-2215. doi:10.1104/pp.111.188953Denison, F. C., Paul, A.-L., Zupanska, A. K., & Ferl, R. J. (2011). 14-3-3 proteins in plant physiology. Seminars in Cell & Developmental Biology, 22(7), 720-727. doi:10.1016/j.semcdb.2011.08.006Carrasco, J. L., Castelló, M. J., & Vera, P. (2006). 14-3-3 Mediates Transcriptional Regulation by Modulating Nucleocytoplasmic Shuttling of Tobacco DNA-binding Protein Phosphatase-1. Journal of Biological Chemistry, 281(32), 22875-22881. doi:10.1074/jbc.m512611200Colcombet, J., & Hirt, H. (2008). ArabidopsisMAPKs: a complex signalling network involved in multiple biological processes. Biochemical Journal, 413(2), 217-226. doi:10.1042/bj20080625Kiegerl, S., Cardinale, F., Siligan, C., Gross, A., Baudouin, E., Liwosz, A., … Meskiene, I. (2000). SIMKK, a Mitogen-Activated Protein Kinase (MAPK) Kinase, Is a Specific Activator of the Salt Stress–Induced MAPK, SIMK. The Plant Cell, 12(11), 2247-2258. doi:10.1105/tpc.12.11.2247CAMPS, M., NICHOLS, A., & ARKINSTALL, S. (2000). Dual specificity phosphatases: a gene family for control of MAP kinase function. The FASEB Journal, 14(1), 6-16. doi:10.1096/fasebj.14.1.6Bethke, G., Pecher, P., Eschen-Lippold, L., Tsuda, K., Katagiri, F., Glazebrook, J., … Lee, J. (2012). Activation of the Arabidopsis thaliana Mitogen-Activated Protein Kinase MPK11 by the Flagellin-Derived Elicitor Peptide, flg22. Molecular Plant-Microbe Interactions, 25(4), 471-480. doi:10.1094/mpmi-11-11-0281Wolschin, F., Wienkoop, S., & Weckwerth, W. (2005). Enrichment of phosphorylated proteins and peptides from complex mixtures using metal oxide/hydroxide affinity chromatography (MOAC). PROTEOMICS, 5(17), 4389-4397. doi:10.1002/pmic.200402049Petersen, M., Brodersen, P., Naested, H., Andreasson, E., Lindhart, U., Johansen, B., … Mundy, J. (2000). Arabidopsis MAP Kinase 4 Negatively Regulates Systemic Acquired Resistance. Cell, 103(7), 1111-1120. doi:10.1016/s0092-8674(00)00213-0Asai, T., Tena, G., Plotnikova, J., Willmann, M. R., Chiu, W.-L., Gomez-Gomez, L., … Sheen, J. (2002). MAP kinase signalling cascade in Arabidopsis innate immunity. Nature, 415(6875), 977-983. doi:10.1038/415977aKosetsu, K., Matsunaga, S., Nakagami, H., Colcombet, J., Sasabe, M., Soyano, T., … Machida, Y. (2010). The MAP Kinase MPK4 Is Required for Cytokinesis in Arabidopsis thaliana. The Plant Cell, 22(11), 3778-3790. doi:10.1105/tpc.110.077164Koroleva, O. A., Tomlinson, M. L., Leader, D., Shaw, P., & Doonan, J. H. (2004). High-throughput protein localization in Arabidopsis using Agrobacterium-mediated transient expression of GFP-ORF fusions. The Plant Journal, 41(1), 162-174. doi:10.1111/j.1365-313x.2004.02281.xVierstra, R. D. (2009). The ubiquitin–26S proteasome system at the nexus of plant biology. Nature Reviews Molecular Cell Biology, 10(6), 385-397. doi:10.1038/nrm2688Gökirmak, T., Paul, A.-L., & Ferl, R. J. (2010). Plant phosphopeptide-binding proteins as signaling mediators. Current Opinion in Plant Biology, 13(5), 527-532. doi:10.1016/j.pbi.2010.06.001Keyse, S. M. (2000). Protein phosphatases and the regulation of mitogen-activated protein kinase signalling. Current Opinion in Cell Biology, 12(2), 186-192. doi:10.1016/s0955-0674(99)00075-7Gupta, R., & Luan, S. (2003). Redox Control of Protein Tyrosine Phosphatases and Mitogen-Activated Protein Kinases in Plants. Plant Physiology, 132(3), 1149-1152. doi:10.1104/pp.103.020792Katou, S., Karita, E., Yamakawa, H., Seo, S., Mitsuhara, I., Kuchitsu, K., & Ohashi, Y. (2005). Catalytic Activation of the Plant MAPK Phosphatase NtMKP1 by Its Physiological Substrate Salicylic Acid-induced Protein Kinase but Not by Calmodulins. Journal of Biological Chemistry, 280(47), 39569-39581. doi:10.1074/jbc.m508115200Schweighofer, A., Kazanaviciute, V., Scheikl, E., Teige, M., Doczi, R., Hirt, H., … Meskiene, I. (2007). The PP2C-Type Phosphatase AP2C1, Which Negatively Regulates MPK4 and MPK6, Modulates Innate Immunity, Jasmonic Acid, and Ethylene Levels in Arabidopsis. The Plant Cell, 19(7), 2213-2224. doi:10.1105/tpc.106.049585Ulm, R. (2001). Mitogen-activated protein kinase phosphatase is required for genotoxic stress relief in Arabidopsis. Genes & Development, 15(6), 699-709. doi:10.1101/gad.192601Yamakawa, H., Katou, S., Seo, S., Mitsuhara, I., Kamada, H., & Ohashi, Y. (2003). Plant MAPK Phosphatase Interacts with Calmodulins. Journal of Biological Chemistry, 279(2), 928-936. doi:10.1074/jbc.m310277200Popescu, S. C., Popescu, G. V., Bachan, S., Zhang, Z., Gerstein, M., Snyder, M., & Dinesh-Kumar, S. P. (2008). MAPK target networks in Arabidopsis thaliana revealed using functional protein microarrays. Genes & Development, 23(1), 80-92. doi:10.1101/gad.1740009Sato, T., Maekawa, S., Yasuda, S., Domeki, Y., Sueyoshi, K., Fujiwara, M., … Yamaguchi, J. (2011). Identification of 14-3-3 proteins as a target of ATL31 ubiquitin ligase, a regulator of the C/N response in Arabidopsis. The Plant Journal, 68(1), 137-146. doi:10.1111/j.1365-313x.2011.04673.xHunter, T. (2007). The Age of Crosstalk: Phosphorylation, Ubiquitination, and Beyond. Molecular Cell, 28(5), 730-738. doi:10.1016/j.molcel.2007.11.01

Repeated long-distance dispersal and convergent evolution in hazel

Closely related species with a worldwide distribution provide an opportunity to understand evolutionary and biogeographic processes at a global scale. Hazel (Corylus) is an economically important genus of tree and shrub species found in temperate regions of Asia, North America and Europe. Here we use multiple nuclear and chloroplast loci to estimate a time-calibrated phylogenetic tree of the genus Corylus. We model the biogeographic history of this group and the evolutionary history of tree and shrub form. We estimate that multiple Corylus lineages dispersed long distances between Europe and Asia and colonised North America from Asia in multiple independent events. The geographic distribution of tree versus shrub form of species appears to be the result of 4–5 instances of convergent evolution in the past 25 million years. We find extensive discordance between our nuclear and chloroplast trees and potential evidence for chloroplast capture in species with overlapping ranges, suggestive of past introgression. The important crop species C. avellana is estimated to be closely related to C. maxima, C. heterophylla var. thunbergii and the Colurnae subsection. Our study provides a new phylogenetic hypothesis or Corylus and reveals how long-distance dispersal can shape the distribution of biodiversity in temperate plants

A survey of the ATP-binding cassette (ABC) gene superfamily in the salmon louse (Lepeophtheirus salmonis)

Salmon lice,Lepeophtheirus salmonis(Krøyer, 1837), are fish ectoparasites causing significant economic damage in the mariculture of Atlantic salmon,Salmo salarLinnaeus, 1758. The control ofL.salmonisat fish farms relies to a large extent on treatment with anti-parasitic drugs. A problem related to chemical control is the potential for development of resistance, which inL.salmonisis documented for a number of drug classes including organophosphates, pyrethroids and avermectins. The ATP-binding cassette (ABC) gene superfamily is found in all biota and includes a range of drug efflux transporters that can confer drug resistance to cancers and pathogens. Furthermore, some ABC transporters are recognised to be involved in conferral of insecticide resistance. While a number of studies have investigated ABC transporters inL.salmonis, no systematic analysis of the ABC gene family exists for this species. This study presents a genome-wide survey of ABC genes inL.salmonisfor which, ABC superfamily members were identified through homology searching of theL.salmonisgenome. In addition, ABC proteins were identified in a reference transcriptome of the parasite generated by high-throughput RNA sequencing (RNA-seq) of a multi-stage RNA library. Searches of both genome and transcriptome allowed the identification of a total of 33 genes / transcripts coding for ABC proteins, of which 3 were represented only in the genome and 4 only in the transcriptome. Eighteen sequences were assigned to ABC subfamilies known to contain drug transporters,i.e. subfamilies B (4 sequences), C (11) and G (2). The results suggest that the ABC gene family ofL.salmonispossesses fewer members than recorded for other arthropods. The present survey of theL.salmonisABC gene superfamily will provide the basis for further research into potential roles of ABC transporters in the toxicity of salmon delousing agents and as potential mechanisms of drug resistance

Global marine pollutants inhibit P-glycoprotein: Environmental levels, inhibitory effects, and cocrystal structure.

The world's oceans are a global reservoir of persistent organic pollutants to which humans and other animals are exposed. Although it is well known that these pollutants are potentially hazardous to human and environmental health, their impacts remain incompletely understood. We examined how persistent organic pollutants interact with the drug efflux transporter P-glycoprotein (P-gp), an evolutionarily conserved defense protein that is essential for protection against environmental toxicants. We identified specific congeners of organochlorine pesticides, polychlorinated biphenyls, and polybrominated diphenyl ethers that inhibit mouse and human P-gp, and determined their environmental levels in yellowfin tuna from the Gulf of Mexico. In addition, we solved the cocrystal structure of P-gp bound to one of these inhibitory pollutants, PBDE (polybrominated diphenyl ether)-100, providing the first view of pollutant binding to a drug transporter. The results demonstrate the potential for specific binding and inhibition of mammalian P-gp by ubiquitous congeners of persistent organic pollutants present in fish and other foods, and argue for further consideration of transporter inhibition in the assessment of the risk of exposure to these chemicals