Allelic Polymorphism of GIGANTEA is Responsible for Naturally Occurring Variation in Circadian Period in Brassica Rapa

GIGANTEA (GI) was originally identified by a late-flowering mutant in Arabidopsis, but subsequently has been shown to act in circadian period determination, light inhibition of hypocotyl elongation, and responses to multiple abiotic stresses, including tolerance to high salt and cold (freezing) temperature. Genetic mapping and analysis of families of heterogeneous inbred lines showed that natural variation in GI is responsible for a major quantitative trait locus in circadian period in Brassica rapa. We confirmed this conclusion by transgenic rescue of an Arabidopsis gi-201 loss of function mutant. The two B. rapa GI alleles each fully rescued the delayed flowering of Arabidopsis gi-201 but showed differential rescue of perturbations in red light inhibition of hypocotyl elongation and altered cold and salt tolerance. The B. rapa R500 GI allele, which failed to rescue the hypocotyl and abiotic stress phenotypes, disrupted circadian period determination in Arabidopsis. Analysis of chimeric B. rapa GI alleles identified the causal nucleotide polymorphism, which results in an amino acid substitution (S264A) between the two GI proteins. This polymorphism underlies variation in circadian period, cold and salt tolerance, and red light inhibition of hypocotyl elongation. Loss-of-function mutations of B. rapa GI confer delayed flowering, perturbed circadian rhythms in leaf movement, and increased freezing and increased salt tolerance, consistent with effects of similar mutations in Arabidopsis. Collectively, these data suggest that allelic variation of GI—and possibly of clock genes in general—offers an attractive target for molecular breeding for enhanced stress tolerance and potentially for improved crop yield

Arabidopsis bHLH100 and bHLH101 Control Iron Homeostasis via a FIT-Independent Pathway

Iron deficiency induces a complex set of responses in plants, including developmental and physiological changes, to increase iron uptake from soil. In Arabidopsis, many transporters involved in the absorption and distribution of iron have been identified over the past decade. However, little is known about the signaling pathways and networks driving the various responses to low iron. Only the basic helix–loop–helix (bHLH) transcription factor FIT has been shown to control the expression of the root iron uptake machinery genes FRO2 and IRT1. Here, we characterize the biological role of two other iron-regulated transcription factors, bHLH100 and bHLH101, in iron homeostasis. First direct transcriptional targets of FIT were determined in vivo. We show that bHLH100 and bHLH101 do not regulate FIT target genes, suggesting that they play a non-redundant role with the two closely related bHLH factors bHLH038 and bHLH039 that have been suggested to act in concert with FIT. bHLH100 and bHLH101 play a crucial role in iron-deficiency responses, as attested by their severe growth defects and iron homeostasis related phenotypes on low-iron media. To gain further insight into the biological role of bHLH100 and bHLH101, we performed microarray analysis using the corresponding double mutant and showed that bHLH100 and bHLH101 likely regulate genes involved in the distribution of iron within the plant. Altogether, this work establishes bHLH100 and bHLH101 as key regulators of iron-deficiency responses independent of the master regulator FIT and sheds light on new regulatory networks important for proper growth and development under low iron conditions

The Importance of Ambient Temperature to Growth and the Induction of Flowering

Plant development is exquisitely sensitive to the environment. Light quantity, quality, and duration (photoperiod) have profound effects on vegetative morphology and flowering time. Recent studies have demonstrated that ambient temperature is a similarly potent stimulus influencing morphology and flowering. In Arabidopsis, ambient temperatures that are high, but not so high as to induce a heat stress response, confer morphological changes that resemble the shade avoidance syndrome. Similarly, these high but not stressful temperatures can accelerate flowering under short day conditions as effectively as exposure to long days. Photoperiodic flowering entails a series of external coincidences, in which environmental cycles of light and dark must coincide with an internal cycle in gene expression established by the endogenous circadian clock. It is evident that a similar model of external coincidence applies to the effects of elevated ambient temperature on both vegetative morphology and the vegetative to reproductive transition. Further study is imperative, because global warming is predicted to have major effects on the performance and distribution of wild species and strong adverse effects on crop yields. It is critical to understand temperature perception and response at a mechanistic level and to integrate this knowledge with our understanding of other environmental responses, including biotic and abiotic stresses, in order to improve crop production sufficiently to sustainably feed an expanding world population

Analysis of the Plant bos1 Mutant Highlights Necrosis as an Efficient Defence Mechanism during D. dadantii/Arabidospis thaliana Interaction

Dickeya dadantii is a broad host range phytopathogenic bacterium provoking soft rot disease on many plants including Arabidopsis. We showed that, after D. dadantii infection, the expression of the Arabidopsis BOS1 gene was specifically induced by the production of the bacterial PelB/C pectinases able to degrade pectin. This prompted us to analyze the interaction between the bos1 mutant and D. dadantii. The phenotype of the infected bos1 mutant is complex. Indeed, maceration symptoms occurred more rapidly in the bos1 mutant than in the wild type parent but at a later stage of infection, a necrosis developed around the inoculation site that provoked a halt in the progression of the maceration. This necrosis became systemic and spread throughout the whole plant, a phenotype reminiscent of that observed in some lesion mimic mutants. In accordance with the progression of maceration symptoms, bacterial population began to grow more rapidly in the bos1 mutant than in the wild type plant but, when necrosis appeared in the bos1 mutant, a reduction in bacterial population was observed. From the plant side, this complex interaction between D. dadantii and its host includes an early plant defence response that comprises reactive oxygen species (ROS) production accompanied by the reinforcement of the plant cell wall by protein cross-linking. At later timepoints, another plant defence is raised by the death of the plant cells surrounding the inoculation site. This plant cell death appears to constitute an efficient defence mechanism induced by D. dadantii during Arabidopsis infection

Functionnal analysis of two Heavy Metal ATPase genes in Nicotania tabacum

Le cadmium est un métal lourd non-essentiel naturellement présent dans le sol. Il a été classé par le centre international de recherche sur le cancer (CIRC) comme un élément cancérigène de type I. Contrairement à la majorité des autres plantes, le tabac (Nicotiana tabacum) accumule le cadmium à des niveaux relativement élevés dans ses parties aériennes. Ce cadmium est ensuite retrouvé dans la fumée de cigarette. La concentration en cadmium dans les vaisseaux sanguins des fumeurs est deux à trois fois supérieure à celle que l'on rencontre chez les non-fumeurs. Pour diminuer la toxicité des cigarettes, il est souhaitable de diminuer la quantité de cadmium accumulé par le tabac dans ses feuilles. Pour parvenir à cet objectif, il est nécessaire de comprendre les mécanismes impliqués dans l'accumulation du cadmium chez le tabac.Les acteurs moléculaires impliqués dans le transport et l'accumulation du cadmium in-planta ont été principalement décrits chez l'espèce modèle Arabidopsis thaliana. Chez cette plante, le cadmium est chargé dans le xylème par AtHMA2 et AtHMA4, deux transporteurs de zinc qui partagent une redondance fonctionnelle partielle et qui sont responsables de la translocation du cadmium des parties racinaires vers les parties aériennes. Deux orthologues à AtHMA2 et AtHMA4 ont été identifiés chez N. tabacum et nommés NtHMAα et NtHMAβ. Ces deux transporteurs sont principalement exprimés dans les racines mais on en trouve également dans les feuilles. NtHMAα a été localisé plus précisément au niveau des cellules du péricycle dans les racines et dans les nervures tertiaires des feuilles. L'étude de lignées mutantes a confirmé le rôle de NtHMAα et NtHMAβ dans la translocation du cadmium des parties racinaires vers les parties aériennes. Les lignées qui expriment une version tronquée de NtHMAα ont une réduction de leur teneur en cadmium foliaire de 45%. Les lignées dans lesquelles l'expression des gènes NtHMAα et NtHMAβ est réduite sont sévèrement impactées dans leur développement. Un des phénotypes observé est une diminution drastique de la quantité de graines en raison de l'incapacité du pollen à germer à cause d'un déficit en zinc. Nous avons montré que chez ces lignées, la tolérance au cadmium était accrue. Dans l'ensemble, nos résultats montrent une grande redondance entre NtHMAα et NtHMAβ.Cadmium is a heavy metal naturally present in the soil. It is classified as a Group 1 carcinogen by the International Agency for Research on Cancer (IARC). Unlike most other plants, tobacco (Nicotiana tabacum) translocates most of the cadmium taken up from the soil out of the roots and into the shoots. As a result, cadmium content in cigarettes is a problem for smokers who have four to five times higher blood cadmium concentrations than nonsmokers. In order to reduce cigarette toxicity it is desired to reduce cadmium accumulated in tobacco leaves. For this purpose, it is important to understand the mechanisms controlling cadmium accumulation in shoots.Molecular actors involved in cadmium repartition in plants have been well described in the model plant Arabidopsis thaliana. In Arabidopsis, cadmium is loaded into the xylem vessels by HMA2 and HMA4, two zinc transporters with partial functional redundancy. Two orthologous proteins of AtHMA2 and AtHMA4 were identified in N. tabacum and named NtHMAα and NtHMAβ. These two transporters are mainly expressed in roots but their expression was also found in shoots. NtHMAα expression was more precisely found in root pericycle cells and in shoot tertiary nerves. The analysis of mutant lines confirmed that NtHMAα and NtHMAβ are involved in cadmium translocation from roots to shoots. Lines which expressed a truncated version of NtHMAα had a 45% reduction in shoot cadmium content. Lines where both NtHMAα and NtHMAβ were silenced were severely impacted in their development. One of the phenotypes that was identified was a drastic reduction in the number of seeds due to the lack of pollen germination. We also found an enhanced tolerance to cadmium in the silenced lines. Altogether, our results show a great redundancy between NtHMAα and NtHMAβ

Functional analysis of HMA genes in tobacco and production of low-cadmium tobacco lines

International audienc

Functional analysis of two Heavy Metal ATPases (HMA) in tobacco

International audienc

Expression and activity of selected FIT direct targets in <i>bhlh100/bhlh101</i> and wild-type backgrounds.

<p>Relative gene expression level measured by quantitative RT-PCR of <i>IRT1</i> (A), <i>FIT</i> (B), and <i>IRT2</i> (C) in wild-type (WT) and <i>bhlh100/bhlh101</i> plants grown on 1/2 MS without iron (black) or in the presence of 100 µM Fe (white). (D) Root ferric reductase activity from WT and <i>bhlh100/bhlh101</i> plants grown as in A–C. Error bars show standard error (n = 3–4). * indicates a statistically significant difference (p-value <0.05) between wild-type and <i>bhlh100/101</i> for the indicated condition.</p

<i>bhlh100/bhlh101</i> plants grow poorly on media with low iron as compared to wild-type.

<p>(A) 2-week-old wild-type (WT) and <i>bhlh100/bhlh101</i> plants were grown on 1/2 MS supplemented with a range of iron concentration or ferrozine (frz) as indicated on the left-hand side. (B) A close-up picture of wild-type (WT, left) and <i>bhlh100/bhlh101</i> (right) plants the grown as in (A) on media without added iron. (C) Complementation of low-iron growth defects by expression of a <i>bHLH100</i> genomic clone in <i>bhlh100/bhlh101.</i> Plants were grown as in (B). (D) 3-week-old wild-type and <i>bhlh100/bhlh101</i> plants were grown on soil supplemented with the indicated percent (w/w) of lime (CaO).</p