Characterization of four bifunctional plant IAM/PAM-amidohydrolases capable of contributing to auxin biosynthesis.

Amidases [EC 3.5.1.4] capable of converting indole-3-acetamide (IAM) into the major plant growth hormone indole-3-acetic acid (IAA) are assumed to be involved in auxin de novo biosynthesis. With the emerging amount of genomics data, it was possible to identify over forty proteins with substantial homology to the already characterized amidases from Arabidopsis and tobacco. The observed high conservation of amidase-like proteins throughout the plant kingdom may suggest an important role of theses enzymes in plant development. Here, we report cloning and functional analysis of four, thus far, uncharacterized plant amidases from Oryza sativa, Sorghum bicolor, Medicago truncatula, and Populus trichocarpa. Intriguingly, we were able to demonstrate that the examined amidases are also capable of converting phenyl-2-acetamide (PAM) into phenyl-2-acetic acid (PAA), an auxin endogenous to several plant species including Arabidopsis. Furthermore, we compared the subcellular localization of the enzymes to that of Arabidopsis AMI1, providing further evidence for similar enzymatic functions. Our results point to the presence of a presumably conserved pathway of auxin biosynthesis via IAM, as amidases, both of monocot, and dicot origins, were analyzed

Mechanisms of glacial-to-future atmospheric CO2 effects on plant immunity

• Impacts of rising atmospheric CO2 concentrations on plant disease have received much attention recently. Nonetheless, evidence about the direct mechanisms by which CO2 shapes plant immunity remains fragmented and controversial. Furthermore, the impact of sub-ambient CO2 concentrations, which plants have experienced repeatedly over the past 800,000 years, has been largely overlooked. • A combination of gene expression analysis, phenotypic characterisation of mutants and mass spectrometry-based metabolic profiling, was implemented to determine development-independent effects of sub-ambient CO2 (saCO2) and elevated CO2 (eCO2) on Arabidopsis immunity. • Resistance to the necrotrophic Plectosphaerella cucumerina (Pc) was repressed at saCO2 and enhanced at eCO2. This CO2-dependent resistance was associated with priming of jasmonic acid (JA)-dependent gene expression and required intact JA biosynthesis and signalling. Resistance to the biotrophic oomycete Hyaloperonospora arabidopsidis (Hpa) increased at both eCO2 and saCO2. Although eCO2 primed salicylic acid (SA)-dependent gene expression, mutations affecting SA signalling only partially suppressed Hpa resistance at eCO2, suggesting additional mechanisms are involved. Induced production of intracellular reactive oxygen species (ROS) at saCO2 corresponded to a loss of resistance in glycolate oxidase (GOX) mutants and increased transcription of the peroxisomal catalase gene CAT2, unveiling a mechanism by which photorespiration-derived ROS determined Hpa resistance at saCO2. • By separating indirect developmental impacts from direct immunological effects, we uncover distinct mechanisms by which CO2 shapes plant immunity and discuss their evolutionary significance

The role of CYP71A12 monooxygenase in pathogen-triggered tryptophan metabolism and Arabidopsis immunity

13 Pág.Effective defense of Arabidopsis against filamentous pathogens requires two mechanisms, both of which involve biosynthesis of tryptophan (Trp)-derived metabolites. Extracellular resistance involves products of PEN2-dependent metabolism of indole glucosinolates (IGs). Restriction of further fungal growth requires PAD3-dependent camalexin and other, as yet uncharacterized, indolics. This study focuses on the function of CYP71A12 monooxygenase in pathogen-triggered Trp metabolism, including the biosynthesis of indole-3-carboxylic acid (ICA). Moreover, to investigate the contribution of CYP71A12 and its products to Arabidopsis immunity, we analyzed infection phenotypes of multiple mutant lines combining pen2 with pad3, cyp71A12, cyp71A13 or cyp82C2. Metabolite profiling of cyp71A12 lines revealed a reduction in ICA accumulation. Additionally, analysis of mutant plants showed that low amounts of ICA can form during an immune response by CYP71B6/AAO1-dependent metabolism of indole acetonitrile, but not via IG hydrolysis. Infection assays with Plectosphaerella cucumerina and Colletotrichum tropicale, two pathogens with different lifestyles, revealed cyp71A12-, cyp71A13- and cyp82C2-associated defects associated with Arabidopsis immunity. Our results indicate that CYP71A12, but not CYP71A13, is the major enzyme responsible for the accumulation of ICA in Arabidopsis in response to pathogen ingression. We also show that both enzymes are key players in the resistance of Arabidopsis against selected filamentous pathogens after they invade.This work was supported by the National Science Centre SONATA BIS grant (UMO-2012/07/E/NZ2/04098) to PB and PRELUDIUM grant (UMO-2013/09/N/NZ2/02080) to KK, and Spanish Ministry of Economy and Competitiveness (MINECO) grant BIO2015-64077-R to AM, and Grants-in-Aid for Scientific Research (18H04780, 18K19212) (KAKENHI) and the Asahi Glass Foundation to YT. EG has been supported by a Heisenberg Fellowship of the Deutsche Forschungsgemeinschaft (GL346/5) and the TUM Junior Fellow Fund.Peer reviewe

COP1 destabilizes DELLA proteins in Arabidopsis

DELLA transcriptional regulators are central components in the control of plant growth responses to the environment. This control is considered to be mediated by changes in the metabolism of the hormones gibberellins (GAs), which promote the degradation of DELLAs. However, here we show that warm temperature or shade reduced the stability of a GA-insensitive DELLA allele in Arabidopsis thaliana. Furthermore, the degradation of DELLA induced by the warmth preceded changes in GA levels and depended on the E3 ubiquitin ligase CONSTITUTIVELY PHOTOMORPHOGENIC1 (COP1). COP1 enhanced the degradation of normal and GAinsensitive DELLA alleles when coexpressed in Nicotiana benthamiana. DELLA proteins physically interacted with COP1 in yeast, mammalian, and plant cells. This interaction was enhanced by the COP1 complex partner SUPRESSOR OF phyA-105 1 (SPA1). The level of ubiquitination of DELLA was enhanced by COP1 and COP1 ubiquitinated DELLA proteins in vitro. We propose that DELLAs are destabilized not only by the canonical GA-dependent pathway but also by COP1 and that this control is relevant for growth responses to shade and warm temperature.Fil: Blanco Touriñán, Noel. Universidad Politécnica de Valencia; EspañaFil: Legris, Martina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquímicas de Buenos Aires. Fundación Instituto Leloir. Instituto de Investigaciones Bioquímicas de Buenos Aires; ArgentinaFil: Minguet, Eugenio G.. Universidad Politécnica de Valencia; EspañaFil: Costigliolo Rojas, María Cecilia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquímicas de Buenos Aires. Fundación Instituto Leloir. Instituto de Investigaciones Bioquímicas de Buenos Aires; ArgentinaFil: Nohales, María A.. University of Southern California; Estados UnidosFil: Iniesto, Elisa. Consejo Superior de Investigaciones Científicas; EspañaFil: García León, Marta. Consejo Superior de Investigaciones Científicas; EspañaFil: Pacín, Manuel. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Fisiológicas y Ecológicas Vinculadas a la Agricultura. Universidad de Buenos Aires. Facultad de Agronomía. Instituto de Investigaciones Fisiológicas y Ecológicas Vinculadas a la Agricultura; ArgentinaFil: Heucken, Nicole. Universitat Dusseldorf; AlemaniaFil: Blomeier, Tim. Universitat Dusseldorf; AlemaniaFil: Locascio, Antonella. Universidad Politécnica de Valencia; EspañaFil: Cerný, Martin. Mendel University in Brno; República ChecaFil: Esteve Bruna, David. Universidad Politécnica de Valencia; EspañaFil: Díez Díaz, Mónica. Univerdiad Catolica de Valencia; EspañaFil: Brzobohatý, Bretislav. Mendel University in Brno; República ChecaFil: Frerigmann, Henning. Max Planck Institute for Plant Breeding Research; AlemaniaFil: Zurbriggen, Matías D.. Universitat Dusseldorf; AlemaniaFil: Kay, Steve A.. University of Southern California; Estados UnidosFil: Rubio, Vicente. Consejo Superior de Investigaciones Científicas; EspañaFil: Blázquez, Miguel A.. Universidad Politécnica de Valencia; EspañaFil: Casal, Jorge José. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquímicas de Buenos Aires. Fundación Instituto Leloir. Instituto de Investigaciones Bioquímicas de Buenos Aires; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Fisiológicas y Ecológicas Vinculadas a la Agricultura. Universidad de Buenos Aires. Facultad de Agronomía. Instituto de Investigaciones Fisiológicas y Ecológicas Vinculadas a la Agricultura; ArgentinaFil: Alabadí, David. Universidad Politécnica de Valencia; Españ

COP1 destabilizes DELLA proteins in Arabidopsis

DELLA transcriptional regulators are central components in the control of plant growth responses to the environment. This control is considered to be mediated by changes in the metabolism of the hormones gibberellins (GAs), which promote the degradation of DELLAs. However, here we show that warm temperature or shade reduced the stability of a GA-insensitive DELLA allele in Arabidopsis thaliana. Furthermore, the degradation of DELLA induced by the warmth preceded changes in GA levels and depended on the E3 ubiquitin ligase CONSTITUTIVELY PHOTOMORPHOGENIC1 (COP1). COP1 enhanced the degradation of normal and GA-insensitive DELLA alleles when coexpressed in Nicotiana benthamiana. DELLA proteins physically interacted with COP1 in yeast, mammalian, and plant cells. This interaction was enhanced by the COP1 complex partner SUPRESSOR OF phyA-105 1 (SPA1). The level of ubiquitination of DELLA was enhanced by COP1 and COP1 ubiquitinated DELLA proteins in vitro. We propose that DELLAs are destabilized not only by the canonical GA-dependent pathway but also by COP1 and that this control is relevant for growth responses to shade and warm temperature.This work was supported by the Spanish Ministry of Economy, Industry and Competitiveness and Agencia Española de Investigación/Fondo Europeo para el Desarrollo Regional/Unión Europea (grants BIO2016-79133-P to D.A. and BIO2013-46539-R and BIO2016-80551-R to V.R.); the European Union SIGNAT-Research and Innovation Staff Exchange (Grant H2020-MSCA-RISE-2014-644435 to M.A.B., D.A., and J.J.C.); the Argentinian Agencia Nacional de Promoción Científica y Tecnológica (Grant Proyectos de Investigación Científica y Tecnológica-2016-1459 to J.J.C.); Universidad de Buenos Aires (grant 20020170100505BA to J.J.C.); the National Institute of General Medical Sciences of the National Institutes of Health (awards R01GM067837 and R01GM056006 to S.A.K.); the German Research Foundation (DFG) under Germany’s Excellence Strategy/Initiative (Cluster of Excellence on Plant Sciences – Excellence Cluster EXC-2048/1, Project ID 390686111 to M.D.Z.); the International Max Planck Research School of the Max Planck Society; the Universities of Düsseldorf and of Cologne to T.B.; Nordrhein Westfalen Bioeconomy Science Center-FocusLabs CombiCom to N.H. and M.D.Z.; and Ministry of Education, Youth and Sports of the Czech Republic (Project LQ1601 Central European Institute of Technology 2020 to B.B. and M.C.). N.B.-T., E.I., and M.G.-L. were supported by Ministerio de Economía y Competitividad-Formación de Personal Investigador Program fellowships

Genetic architecture of glucosinolate variation in Brassica napus

The diverse biological activities of glucosinolate (GSL) hydrolysis products play significant biological and economical roles in the defense system and nutritional qualities of Brassica napus (oilseed rape). Yet, genomic-based study of the B. napus GSL regulatory mechanisms are scarce due to the complexity of working with polyploid species. To address these challenges, we used transcriptome-based GWAS approach, Associative Transcriptomics (AT), across a diversity panel of 288 B. napus genotypes to uncover the underlying genetic basis controlling quantitative variation of GSLs in B. napus vegetative tissues. Single nucleotide polymorphism (SNP) markers and gene expression markers (GEMs) associations identify orthologues of MYB28/HAG1 (AT5G61420), specifically the copies on chromosome A9 and C2, to be the key regulators of aliphatic GSL variation in leaves. We show that the positive correlation observed between aliphatic GSLs in seed and leaf is due to the amount synthesized, as controlled by Bna.HAG1.A9 and Bna.HAG1.C2, rather than by variation in the transport processes. In addition, AT and differential expression analysis in root tissues implicate an orthologue of MYB29/HAG3 (AT5G07690), Bna.HAG3.A3, as controlling root aromatic GSL variation. Based on the root expression data we also propose Bna.MAM3.A3 to have a role in controlling phenylalanine chain elongation for aromatic GSL biosynthesis. This work uncovers a regulator of homophenylalanine-derived aromatic GSLs and implicates the shared biosynthetic pathways between aliphatic and aromatic GSLs