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cellular and molecular Requirements for Polar PIN Targeting and Transcytosis in Plants

molecular Plant 2008年第6期1卷 1056-1066页

作者： Jurgen Kleine-Vehn tukasz tangowski Justyna Wisniewska Pankaj Dhonukshe Philip B Brewer Jiri Friml department of Plant Systems biology VIB and Department of Molecular Genetics Ghent University 9052 Gent Belgium Zentrum fur Molekularbiologie der Pflanzen (ZMBP) Universitat Tubingen 72076 Tubingen Germany present address: Institute of General and molecular biology Nicolas Copernicus University 87-100 Torun Poland present address: department of biology Utrecht University 3508 TB Utrecht The Netherlands present address: Centre for Integrative Legume Research University of Queensland Brisbane Australia

The polar, sub-cellular localization of PIN auxin efflux carriers determines the direction of intercellular auxin flow, thus defining the spatial aspect of auxin signalling. Dynamic, transcytosis-like relocalizations of PIN proteins occur in response to external and internal signals, integrating these signals into changes in auxin distribution. Here, we examine the cellular and molecular mechanisms of polar PIN delivery and transcytosis. The mechanisms of the ARF-GEF-dependent polar targeting and transcytosis are well conserved and show little variations among diverse Arabidopsis ecotypes consistent with their fundamental importance in regulating plant development. At the cellular level, we refine previous findings on the role of the actin cytoskeleton in apical and basal PIN targeting, and identify a previously unknown role for microtubules, specifically in basal targeting. PIN protein delivery to different sides of the cell is mediated by ARFdependent trafficking with a previously unknown complex level of distinct ARF-GEF vesicle trafficking regulators. Our data suggest that alternative recruitment of PIN proteins by these distinct pathways can account for cell type- and cargo-specific aspects of polar targeting, as well as for polarity changes in response to different signals. The resulting dynamic PIN positioning to different sides of cells defines a three-dimensional pattern of auxin fluxes within plant tissues.

关键词： cytoskeleton polarity protein targeting.

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Regulation of the Arabidopsis GSK3-like Kinase BRASSINOSTEROID-INSENSITIVE 2 through Proteasome-Mediated Protein Degradation

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molecular Plant 2008年第2期1卷 338-346页

作者： Peng Peng Zhenyan Yan Yongyou Zhu Jianming Li department of molecular Cellular and Development Biology University of Michigan Ann Arbor MI 48109-1048 USA present address: Howard Hughes Medical Institute Department of Molecular Cell and Developmental Biology University of California-Los AngelesLos Angeles CA g0095 USA

Glycogen synthase kinase 3 （GSK3） is a unique serine/threonine kinase that is implicated in a variety of cellular processes and is regulated by phosphorylation or protein-protein interaction in animal cells. BIN2 is an Arabidopsis GSK3- like kinase that negatively regulates brassinosteroid （BR） signaling. Genetic studies suggested that BIN2 is inhibited in response to BR perception at the cell surface to relieve its inhibitory effects on downstream targets; however, little is known about biochemical mechanisms of its inhibition. Here, we show that BIN2 is regulated by proteasome-mediated protein degradation. Exogenous application of a BR biosynthesis inhibitor and an active BR increased and decreased the amount of BIN2 proteins, respectively. Interestingly, the gain-of-function bin2-1 mutation significantly stabilizes BIN2, making it unresponsive to BR-induced BIN2 depletion. Exogenous application of different plant growth hormones revealed that BIN2 depletion is specifically induced by BR through a functional BR receptor, while treatment of a protea- some inhibitor, MG132, not only prevented the BR-induced BIN2 depletion but also nullified the inhibitory effect of BR on the BIN2 kinase activity. Taken together, our results strongly suggest that proteasome-mediated protein degradation constitutes an important regulatory mechanism for restricting the BIN2 activity.

关键词： Arabidopsis GSK3-like Kinase BRASSINOSTEROID-INSENSITIVE Proteasome-Mediated Protein Degradation

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Disruption of Arabidopsis CHY1 Reveals an Important Role of Metabolic Status in Plant Cold Stress Signaling

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molecular Plant 2009年第1期2卷 59-72页

作者： Chun-Hai Dong Bethany K. Zolman Bonnie Bartel Byeong-ha Lee Becky Stevenson Manu Agarwal Jian-Kang Zhu department of Botany and Plant Science 2150 Batchelor Hall University of California at Riverside Riverside CA 92521 USA department of biology University of Missouri-St Louis St Louis MO 63121 USA department of Biochemistry and cell biology Rice University Houston TX 77005 USA present address: department of cell biology and molecular Genetics University of Maryland College Park MD 20742 USA present address: department of Life Science Sogang University Seoul 121-742 South Korea

To study cold signaling, we screened for Arabidopsis mutants with altered cold-induced transcription of a firefly luciferase reporter gene driven by the CBF3 promoter （CBF3-LUC）. One mutant, chy1-10, displayed reduced cold-induction of CBF3-LUC luminescence. RNA gel blot analysis revealed that expression of endogenous CBFs also was reduced in the chyl mutant, chy1-10 mutant plants are more sensitive to freezing treatment than wild-type after cold acclimation. Both the wild-type and chyl mutant plants are sensitive to darkness-induced starvation at warm temperatures, although chyl plants are slightly more sensitive. This dark-sensitivity is suppressed by cold temperature in the wildtype but not in chyl. Constitutive CBF3 expression partially rescues the sensitivity of chyl-10 plants to dark treatment in the cold. The chyl mutant accumulates higher levels of reactive oxygen species, and application of hydrogen peroxide can reduce cold-induction of CBF3-LUC in wild-type. Map-based cloning of the gene defective in the mutant revealed a nonsense mutation in CHY1, which encodes a peroxisomal β-hydroxyisobutyryl （HIBYL）-CoA hydrolase needed for valine catabolism and fatty acid β-oxidation. Our results suggest a role for peroxisomal metabolism in cold stress signaling, and plant tolerance to cold stress and darkness-induced starvation.

关键词： Cold stress signal transduction gene regulation CHY1 β-hydroxyisobutyryI-CoA hydrolase.

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Calcineurin-B-Like Protein CBL9 Interacts with Target Kinase CIPK3 in the Regulation of ABA Response in Seed Germination

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molecular Plant 2008年第2期1卷 238-248页

作者： Girdhar K. Pandey John J. Grant Yong Hwa Cheong Beom-Gi Kim Le Gong Li Sheng Luan department of Plant and Microbial biology University of California Berkeley CA 94720 USA department of Plant molecular biology University of Delhi South Campus Benito Juarez Road New Delhi-110021 India present address: department of Bio-Environmental Science Sunchon National University Suncheon Jeonnam 540-742 Korea present address: department of molecular Physiology and Biochemistry National institute of Agricultural Biotechnology Suwon 441-707 Korea

Calcium plays a vital role as a second messenger in many signaling pathways in plants. The calcineurin B-like proteins （CBLs） represent a family of plant calcium-binding proteins that function in calcium signaling by interacting with their interacting protein kinases （CIPKs）. In our previous study, we have reported a role for one of the CBLs （CBL9） and one of the CIPKs （CIPK3） in ABA signaling. Here, we have shown that CBL9 and CIPK3 physically and functionally interact with each other in regulating the ABA responses. The CBL9 and CIPK3 proteins interacted with each other in the yeast two- hybrid system and when expressed in plant cells. The double mutant cbl9cipk3 showed the similar hypersensitive response to ABA as observed in single mutants （cbl9 or cipk3）. The constitutively active form of CIPK3 genetically complemented the cbl9 mutant, indicating that CIPK3 function downstream of CBL9. Based on these findings, we conclude that CBL9 and CIPK3 act together in the same pathway for regulating ABA responses.

关键词： ABA calcium CBL CIPK osmotic stress signal transduction.

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A Light-Independent Allele of Phytochrome B Faithfully Recapitulates Photomorphogenic Transcriptional Networks

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molecular Plant 2009年第1期2卷 166-182页

作者： Wei Hu Yi-Shin Su J. Clark Lagarias department of molecular and cellular biology College of Biological Sciences University of California Davis CA 95616 USA present address: Carnegie Institution of Washington -Department of Plant Biology Stanford CA 94305 USA

Dominant gain-of-function alleles of Arabidopsis phytochrome B were recently shown to confer lightindependent, constitutive photomorphogenic （cop） phenotypes to transgenic plants （Su and Lagarias, 2007）. In the present study, comparative transcription profiling experiments were performed to assess whether the pattern of gene expression regulated by these alleles accurately reflects the process of photomorphogenesis in wild-type Arabidopsis. Whole- genome transcription profiles of dark-grown phyAphyB seedlings expressing the Y^276H mutant of phyB （YHB） revealed that YHB reprograms about 13% of the Arabidopsis transcriptome in a light-independent manner. The YHB-regulated transcriptome proved qualitatively similar to but quantitatively greater than those of wild-type seedlings grown under 15 or 50 μmol m^-2 m^-1 continuous red light （Rc）. Among the 2977 genes statistically significant two-fold （SSTF） regulated by YHB in the absence of light include those encoding components of the photosynthetic apparatus, tetrapyrrole/pigment biosynthetic pathways, and early light-responsive signaling factors. Approximately 80% of genes SSTF regulated by Rc were also YHB-regulated. Expression of a notable subset of 346 YHB-regulated genes proved to be strongly attenuated by Rc, indicating compensating regulation by phyC-E and/or other Rc-dependent processes. Since the majority of these 346 genes are regulated by the circadian clock, these results suggest that phyA- and phyB-independent light signaling pathway（s） strongly influence clock output. Together with the unique plastid morphology of dark-grown YHB seedlings, these analyses indicate that the YHB mutant induces constitutive photomorphogenesis via faithful reconstruction of phyB signaling pathways in a light-independent fashion.

关键词： light signaling signal transduction transcriptome analysis photomorphogenesis Arabidopsis phytochrome.

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Immunophilin-like FKBP42/TWISTED DWARF1 Interacts with the Receptor Kinase BRI1 to Regulate Brassinosteroid Signaling in Arabidopsis

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molecular Plant 2016年第4期9卷 593-600页

作者： Juthamas Chaiwanon Veder J. Garcia Heather Cartwright Ying Sun Zhi-Yong Wang department of Plant biology Carnegie Institution for Science Stanford CA 94305 USA department of Botany Center of Excellence in Environment and Plant Physiology Faculty of Science Chulalongkorn University Bangkok 10330 Thailand present address: Institute of molecular cell biology Hebei Key Laboratory of Molecular and Cellular Biology Hebei Normal University Shijiazhuang 050024 China

Mutation of the immunophilin-like FK506-binding protein TWlSTED DWARF1 （FKBP42/TWD1） causes dwarf and twisted-organ phenotypes in Arabidopsis. However, the function of FKBP42 is not fully understood at the molecular level. Using genetic, physiological, and immunological experiments, we show here that FKBP42/TWD1 is necessary for brassinosteroid （BR） signal transduction. The twdl mutant showed reduced BR sensitivity in growth responses and activation of the BZR1 transcription factor. However, twdl showed normal responses to an inhibitor of BIN2/GSK3, suggesting that twdl has a defect upstream of BIN2 in the BR signaling pathway. In vitro and in vivo assays showed that TWD1 interacts physically with the kinase domains of the BR receptor kinases BRI1 and BAK1. TWD1 is not required for normal localization of BRI1-GFP to the plasma membrane or for activation of the flagellin receptor kinase FLS2. Our results suggest that FKBP42/TWD1 plays a specific role in the activation of BRI1 receptor kinase.

关键词： brassinosteroid signaling BRI1 FKBP42

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Temporal and Spatial Requirement of EMF1 Activity for Arabidopsis Vegetative and Reproductive Development

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molecular Plant 2009年第4期2卷 643-653页

作者： Rosario Sanchez Minjung Y. Kim Myriam Calonje Yong-Hwan Moon Z. Renee Sung department of Plant and Microbial biology University of California Berkeley CA 94720 USA present address: Heidelberg Institute for Plant Science Biodiversity and Plant Systematics Heidelberg University 69120 Heidelberg Germany present address: department of molecular biology Pusan National University 30 Jangjeon-dong Geumjeong-gu Busan 609-735 Korea

EMBRYONIC FLOWER （EMF） genes are required to maintain vegetative development via repression of flower homeotic genes in Arabidopsis. Removal of EMF gene function caused plants to flower upon germination, producing abnormal and sterile flowers. The pleiotropic effect of ernfl mutation suggests its requirement for gene programs involved in diverse developmental processes. Transgenic plants harboring EMF1 promoter：：glucuronidase （GUS） reporter gene were generated to investigate the temporal and spatial expression pattern of EMF1. These plants displayed differential GUS activity in vegetative and flower tissues, consistent with the role of EMF1 in regulating multiple gene programs. EMFI：：GUS expression pattern in emf mutants suggests organ-specific auto-regulation. Sense- and antisense （as） EMF1 cDNA were expressed under the control of stage- and tissue-specific promoters in transgenic plants. Characterization of these transgenic plants showed that EMF1 activity is required in meristematic as well as differentiating tissues to rescue emf mutant phenotype. Temporal removal or reduction of EMF1 activity in the embryo or shoot apex of wild-type seedlings was sufficient to cause early flowering and terminal flower formation in adult plants. Such reproductive cell memory is reflected in the flower MADS-box gene activity expressed prior to flowering in these early flowering plants. However, temporal removal of EMF1 activity in flower meristem did not affect flower development. Our results are consistent with EMF1＇s primary role in repressing flowering in order to allow for vegetative growth.

关键词： EMF1 stage-specific promoter early flowering reproductive cell memory vegetative/reproductive development repression of flower MADS-box genes.

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Blocking the Metabolism of Starch Breakdown Products in Arabidopsis Leaves Triggers Chloroplast Degradation

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molecular Plant 2009年第6期2卷 1233-1246页

作者： Michaela Stettler Simona Eicke Tabea Mettler Gaelle Messerli Stefan Hortensteiner Samuel C. Zeeman department of biology ETH Zurich CH-8092 Zurich Switzerland Institute of Plant biology University of Zurich CH-8008 Zurich Switzerland present address: Max Planck Institute for molecular Plant Physiology Potsdam-Golm 14476 Germany present address: Johnson & Johnson Medical CH-8957 Spreitenbach Switzerland

In most plants, a large fraction of photo-assimilated carbon is stored in the chloroplasts during the day as starch and remobilized during the subsequent night to support metabolism. Mutations blocking either starch synthesis or starch breakdown in Arabidopsis thaliana reduce plant growth. Maltose is the major product of starch breakdown exported from the chloroplast at night. The maltose excess 1 mutant （mex1）, which lacks the chloroplast envelope maltose transporter, accumulates high levels of maltose and starch in chloroplasts and develops a distinctive but previously unexplained chlorotic phenotype as leaves mature. The introduction of additional mutations that prevent starch synthesis, or that block maltose production from starch, also prevent chlorosis of mex1. In contrast, introduction of mutations in disproportionating enzyme （DPE1） results in the accumulation of maltotriose in addition to maltose, and greatly increases chlorosis. These data suggest a link between maltose accumulation and chloroplast homeostasis. Microscopic analyses show that the mesophyll cells in chlorotic mex1 leaves have fewer than half the number of chloroplasts than wild-type cells. Transmission electron microscopy reveals autophagy-like chloroplast degradation in both mex1 and the dpe1/mex1 double mutant. Microarray analyses reveal substantial reprogramming of metabolic and cellular processes, suggesting that organellar protein turnover is increased in mex1, though leaf senescence and senescence-related chlorophyll catabolism are not induced. We propose that the accumulation of maltose and malto-oligosaccharides causes chloroplast dysfunction, which may by signaled via a form of retrograde signaling and trigger chloroplast degradation.

关键词： Carbohydrate metabolism photosynthesis senescence chloroplast biology Arabidopsis autophagy.

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Emerging Perspectives on the Mechanisms, Regulation, and Distribution of Light Color Acclimation in Cyanobacteria

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molecular Plant 2012年第1期5卷 1-13页

作者： Andrian Gutu David M. Kehoe department of biology 1001 East Third Street indiana University Bloomington iN 47405 USA present address： HoWard Hughes Medical Institute/department Of molecular and cellular biology FAS Center for Systems Biology Harvard UniversityCambridge MA 02138 USA

Chromatic acclimation （CA） provides many cyanobacteria with the ability to tailor the properties of their light-harvesting antennae to the spectral distribution of ambient light. CA was originally discovered as a result of its dramatic cellular phenotype in red and green light. However, discoveries over the past decade have revealed that many pairs of light colors, ranging from blue to infrared, can trigger CA responses. The capacity to undergo CA is widespread geographically, occurs in most habitats around the world, and is found within all major cyanobacterial groups. In addition, many other cellular activities have been found to be under CA control, resulting in distinct physiological and morphological states for cells under different light-color conditions. Several types of CA appear to be the result of convergent evolution, where different strategies are used to achieve the final goal of optimizing light-harvesting antenna composition to maximize photon capture. The regulation of CA has been found to occur primarily at the level of RNA abundance. The CA-regulatory pathways uncovered thus far are two-component systems that use phytochrome-class photoreceptors with sensor-kinase domains to control response regulators that function as transcription factors. However, there is also at least one CA- regulatory pathway that operates at the post-transcriptional level. It is becoming increasingly clear that large numbers of cyanobacterial species have the capacity to acclimate to a wide variety of light colors through the use of a range of different CA processes.

关键词： Chromatic adaptation phycobilisome cyanobacteria light regulation gene regulation signal transduction light harvesting phenotypic plasticity.

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Functional Analysis of Semi-conserved Transit Peptide Motifs and Mechanistic Implications in Precursor Targeting and Recognition

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molecular Plant 2016年第9期9卷 1286-1301页

作者： Kristen Holbrook Chitra Subramanian Prakitchai Chotewutmontri L. Evan Reddick Sarah Wright Huixia Zhang Lily Moncrief Barry D. Bruce department of Biochemistry Cellular and Molecular Biology Graduate Program in Plant Physiology and Genetics Graduate School of Genome Science and Technology department of Botany department of BotanyUniversity of Tennessee Knoxville TN 37996 USA present address: department of Infectious Disease St. Jude Children's Research Hospital Memphis TN 38105 USA present address: Institute of molecular biology University of Oregon Eugene OR 97403 USA present address: department of Microbiology University of Texas Southwestern Medical Center 5323 Harry Hines Boulevard Dallas TX 75390 USA present address: Albert R. Mann Library Cornell University Ithaca NY 14850 USA present address: U.S. Food and Drug Administration 10903 New Hampshire Avenue Silver Spring MD 20993 USA

Over 95% of plastid proteins are nuclear-encoded as their precursors containing an N-terminal extension known as the transit peptide （TP）. Although highly variable, TPs direct the precursors through a conserved, posttranslational mechanism involving translocons in the outer （TOC） and inner envelope （TOC）. The organelle import specificity is mediated by one or more components of the Toc complex. However, the high TP diversity creates a paradox on how the sequences can be specifically recognized. An emerging model of TP design is that they contain multiple loosely conserved motifs that are recognized at different steps in the targeting and transport process. Bioinformatics has demonstrated that many TPs contain semiconserved physicochemical motifs, termed FGLK. In order to characterize FGLK motifs in TP recognition and import, we have analyzed two well-studied TPs from the precursor of RuBisCO small subunit （SStp） and ferredoxin （Fdtp）. Both SStp and Fdtp contain two FGLK motifs. Analysis of large set mutations （-85） in these two motifs using in vitro, in organello, and in vivo approaches support a model in which the FGLK domains mediate interaction with TOC34 and possibly other TOC components. In vivo import analysis suggests that multiple FGLK motifs are functionally redundant. Furthermore, we discuss how FGLK motifs are required for efficient precursor protein import and how these elements may permit a convergent function of this highly variable class of targeting sequences.

关键词： chloroplast biology protein translocation Toc34 transit peptide cell biology

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