Arabinogalactan proteins 6 and 11 are required for stamen and pollen function in ArabidopsisLevitin, Bella; Richter, Dganit; Markovich, Inbal; Zik, Moriyah
doi: 10.1111/j.1365-313X.2008.03607.xpmid: 18644001
Successful male reproductive function in plants is dependent on the correct development and functioning of stamens and pollen. AGP6 and AGP11 are two homologous Arabidopsis genes encoding cell wall‐associated arabinogalactan glycoproteins (AGPs). Both genes were found to be specifically expressed in stamens, pollen grains and pollen tubes, suggesting that these genes may play a role in male organ development and function. RNAi lines with reduced AGP6 and AGP11 expression were generated. These, together with lines harboring point mutations in the coding region of AGP6, were used to show that loss of function in AGP6 and AGP11 led to reduced fertility, at least partly as a result of inhibition of pollen tube growth. Our results also suggest that AGP6 and AGP11 play an additional role in the release of pollen grains from the mature anther. Thus, our study demonstrates the involvement of specific AGPs in pollen tube growth and stamen function.
Molecular basis of the functional specificities of phototropin 1 and 2Aihara, Yusuke; Tabata, Ryohei; Suzuki, Tomomi; Shimazaki, Ken‐ichiro; Nagatani, Akira
doi: 10.1111/j.1365-313X.2008.03605.xpmid: 18643969
A blue‐light photoreceptor in plants, phototropin, mediates phototropism, chloroplast relocation, stomatal opening, and leaf‐flattening responses. Phototropin is divided into two functional moieties, the N‐terminal photosensory and the C‐terminal signaling moieties. Phototropin perceives light stimuli by the light, oxygen or voltage (LOV) domain in the N‐terminus; the signal is then transduced intramolecularly to the C‐terminal kinase domain. Two phototropins, phot1 and phot2, which have overlapping and distinct functions, exist in Arabidopsis thaliana. Phot1 mediates responses with higher sensitivity than phot2. Phot2 mediates specific responses, such as the chloroplast avoidance response and chloroplast dark positioning. To elucidate the molecular basis for the functional specificities of phot1 and phot2, we exchanged the N‐ and C‐terminal moieties of phot1 and phot2, fused them to GFP and expressed them under the PHOT2 promoter in the phot1 phot2 mutant background. With respect to phototropism and other responses, the chimeric phototropin consisting of phot1 N‐terminal and phot2 C‐terminal moieties (P1n/2cG) was almost as sensitive as phot1; whereas the reverse combination (P2n/1cG) functioned with lower sensitivity. Hence, the N‐terminal moiety mainly determined the sensitivity of the phototropins. Unexpectedly, both P1n/2cG and P2n/1cG mediated the chloroplast avoidance response, which is specific to phot2. Hence, chloroplast avoidance activity appeared to be suppressed specifically in the combination of N‐ and C‐terminal moieties of phot1. Unlike the chloroplast avoidance response, chloroplast dark positioning was observed for P2G and P2n/1cG but not for P1G or P1n/2cG, suggesting that a specific structure in the N‐terminal moiety of phot2 is required for this activity.
HAHB4, a sunflower HD‐Zip protein, integrates signals from the jasmonic acid and ethylene pathways during wounding and biotic stress responsesManavella, Pablo A.; Dezar, Carlos A.; Bonaventure, Gustavo; Baldwin, Ian T.; Chan, Raquel L.
doi: 10.1111/j.1365-313X.2008.03604.xpmid: 18643970
The Helianthus annuus (sunflower) HAHB4 transcription factor belongs to the HD‐Zip family and its transcript levels are strongly induced when sunflower plants are attacked by herbivores, mechanically damaged or treated with methyl‐jasmonic acid (MeJA) or ethylene (ET). Promoter fusion analysis, in Arabidopsis and in sunflower, demonstrated that induction of HAHB4 expression by these treatments is regulated at the transcriptional level. In transiently transformed sunflower plants HAHB4 expression upregulates the transcript levels of several genes involved in JA biosynthesis and defense‐related processes such as the production of green leaf volatiles and trypsin protease inhibitors (TPI). In HAHB4 sunflower overexpressing tissue, increased activities of lipoxygenase, hydroperoxide lyase and TPI are detected whereas in HAHB4‐silenced tissue these activities are reduced. Transgenic Arabidopsis thaliana and Zea mays plants ecotopically expressing HAHB4 also exhibit higher transcript levels of defense‐related genes and when Spodoptera littoralis or Spodoptera frugiperda larvae are placed on each species, respectively, larvae consumed less and gain less mass compared with larvae feeding on control plants. Arabidopsis plants ectopically expressing HAHB4 had higher amounts of JA, JA‐isoleucine and ET compared with control plants both before and after wounding, but reduced levels of salicylic acid (SA) after wounding and bacterial infection. We conclude that HAHB4 coordinates the production of phytohormones during biotic stress responses and mechanical damage, specifically by positively regulating JA and ET production and negatively regulating ET sensitivity and SA accumulation.
Trans‐specific gene silencing between host and parasitic plantsTomilov, Alexey A.; Tomilova, Natalia B.; Wroblewski, Tadeusz; Michelmore, Richard; Yoder, John I.
doi: 10.1111/j.1365-313X.2008.03613.xpmid: 18643992
Species of Orobanchaceae parasitize the roots of nearby host plants to rob them of water and other nutrients. Parasitism can be debilitating to the host plant, and some of the world’s most pernicious agricultural pests are parasitic weeds. We demonstrate here that interfering hairpin constructs transformed into host plants can silence expression of the targeted genes in the parasite. Transgenic roots of the hemi‐parasitic plant Triphysaria versicolor expressing the GUS reporter gene were allowed to parasitize transgenic lettuce roots expressing a hairpin RNA containing a fragment of the GUS gene (hpGUS). When stained for GUS activity, Triphysaria roots attached to non‐transgenic lettuce showed full GUS activity, but those parasitizing transgenic hpGUS lettuce lacked activity in root tissues distal to the haustorium. Transcript quantification indicated a reduction in the steady‐state level of GUS mRNA in Triphysaria when they were attached to hpGUS lettuce. These results demonstrate that the GUS silencing signal generated by the host roots was translocated across the haustorium interface and was functional in the parasite. Movement across the haustorium was bi‐directional, as demonstrated in double‐junction experiments in which non‐transgenic Triphysaria concomitantly parasitized two hosts, one transgenic for hpGUS and the other transgenic for a functional GUS gene. Observation of GUS silencing in the second host demonstrated that the silencing trigger could be moved from one host to another using the parasite as a physiological bridge. Silencing of parasite genes by generating siRNAs in the host provides a novel strategy for controlling parasitic weeds.
Exploring the nuclear proteome of Medicago truncatula at the switch towards seed fillingRepetto, Ombretta; Rogniaux, Hélène; Firnhaber, Christian; Zuber, Hélène; Küster, Helge; Larré, Colette; Thompson, Richard; Gallardo, Karine
doi: 10.1111/j.1365-313X.2008.03610.xpmid: 18643982
Despite its importance in determining seed composition, and hence quality, regulation of the development of legume seeds is incompletely understood. Because of the cardinal role played by the nucleus in gene expression and regulation, we have characterized the nuclear proteome of Medicago truncatula at the 12 days after pollination (dap) stage that marks the switch towards seed filling. Nano‐liquid chromatography–tandem mass spectrometry analysis of nuclear protein bands excised from one‐dimensional SDS‐PAGE identified 179 polypeptides (143 different proteins), providing an insight into the complexity and distinctive feature of the seed nuclear proteome and highlighting new plant nuclear proteins with possible roles in the biogenesis of ribosomal subunits (PESCADILLO‐like) or nucleocytoplasmic trafficking (dynamin‐like GTPase). The results revealed that nuclei of 12‐dap seeds store a pool of ribosomal proteins in preparation for intense protein synthesis activity, occurring subsequently during seed filling. Diverse proteins of the molecular machinery leading to the synthesis of ribosomal subunits were identified along with proteins involved in transcriptional regulation, RNA processing or transport. Some had already been shown to play a role during the early stages of seed formation whereas for others the findings are novel (e.g. the DIP2 and ES43 transcriptional regulators or the RNA silencing‐related ARGONAUTE proteins). This study also revealed the presence of chromatin‐modifying enzymes and RNA interference proteins that have roles in RNA‐directed DNA methylation and may be involved in modifying genome architecture and accessibility during seed filling and maturation.
ITN1, a novel gene encoding an ankyrin‐repeat protein that affects the ABA‐mediated production of reactive oxygen species and is involved in salt‐stress tolerance in Arabidopsis thalianaSakamoto, Hikaru; Matsuda, Osamu; Iba, Koh
doi: 10.1111/j.1365-313X.2008.03614.xpmid: 18643991
Salt stress and abscisic acid (ABA) induce accumulation of reactive oxygen species (ROS) in plant cells. ROS not only act as second messengers for the activation of salt‐stress responses, but also have deleterious effects on plant growth due to their cytotoxicity. Therefore, the timing and degree of activation of ROS‐producing or ROS‐scavenging enzymes must be tightly regulated under salt‐stress conditions. We identified a novel locus of Arabidopsis, designated itn1 (increased tolerance to NaCl1), whose disruption leads to increased salt‐stress tolerance in vegetative tissues. ITN1 encodes a transmembrane protein with an ankyrin‐repeat motif that has been implicated in diverse cellular processes such as signal transduction. Comparative microarray analysis between wild‐type and the itn1 mutant revealed that induction of genes encoding the ROS‐producing NADPH oxidases (RBOHC and RBOHD) under salt‐stress conditions was suppressed in the mutant. This suppression was accompanied by a corresponding reduction in ROS accumulation. The ABA‐induced expression of RBOHC and RBOHD was also suppressed in the mutant, as was the case for RD29A, an ABA‐inducible marker gene. However, the ABA‐induced expression of another marker gene, RD22, was not impaired in the mutant. These results suggest that the itn1 mutation partially impairs ABA signaling pathways, possibly leading to the reduction in ROS accumulation under salt‐stress conditions. We discuss the possible mechanisms underlying the salt‐tolerant phenotype of the itn1 mutant.
Involvement of RTE1 in conformational changes promoting ETR1 ethylene receptor signaling in ArabidopsisResnick, Josephine S.; Rivarola, Maximo; Chang, Caren
doi: 10.1111/j.1365-313X.2008.03615.xpmid: 18643990
Ethylene is an important regulator of plant growth, development and responses to environmental stresses. Arabidopsis perceives ethylene through five homologous receptors that negatively regulate ethylene responses. RTE1, a novel gene conserved in plants, animals and some protists, was recently identified as a positive regulator of the ETR1 ethylene receptor. Here, we genetically analyze the dependence of ETR1 on RTE1 in order to obtain further insight into RTE1 function. The function of RTE1 was found to be independent and distinct from that of RAN1, which encodes a copper transporter required for ethylene receptor function. We tested the ability of an rte1 loss‐of‐function mutation to suppress 11 etr1 ethylene‐binding domain mis‐sense mutations, all of which result in dominant ethylene insensitivity due to constitutive signaling. This suppression test uncovered two classes of etr1 mutations –RTE1‐dependent and RTE1‐independent. The nature of these mutations suggests that the ethylene‐binding domain is a possible target of RTE1 action. Based on these findings, we propose that RTE1 promotes ETR1 signaling through a conformational effect on the ethylene‐binding domain.
GLK transcription factors regulate chloroplast development in a cell‐autonomous mannerWaters, Mark T.; Moylan, Elizabeth C.; Langdale, Jane A.
doi: 10.1111/j.1365-313X.2008.03616.xpmid: 18643989
In a number of land plants, Golden2‐like (GLK) genes encode a pair of partially redundant nuclear transcription factors that are required for the expression of nuclear photosynthetic genes and for chloroplast development. As chloroplast biogenesis depends on close co‐operation between the nuclear and plastid compartments, GLK gene function must be dependent on tight intracellular control. However, the extent to which GLK‐mediated chloroplast development depends on intercellular communication is not known. Here we used sector analysis to show that GLK proteins operate cell‐autonomously in leaf mesophyll cells. To establish whether GLK proteins are able to influence adjacent cell layers, we used tissue‐specific promoters to restrict GLK gene expression to the epidermis and to the phloem. GLK genes driven by the Arabidopsis epidermal FIDDLEHEAD (FDH) and MERISTEM LAYER1 (AtML1) promoters failed to rescue the pale‐green Atglk1 Atglk2 mutant phenotype, confirming the suggestion that GLK proteins can only influence chloroplast development intracellularly. An exception to this rule was seen in lines in which GLK genes were expressed in the phloem. However, we believe that the partial complementation of the mutant phenotype that was observed resulted from phloem unloading, as opposed to inherent properties of GLK proteins. We conclude that GLK proteins act in a cell‐autonomous manner to coordinate and maintain the photosynthetic apparatus within individual cells. Significantly, this suggests that GLK proteins provide a means to fine‐tune photosynthesis according to the differential requirements of cells within the leaf.