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1.
J Exp Bot ; 69(20): 4757-4771, 2018 09 14.
Article in English | MEDLINE | ID: mdl-29945158

ABSTRACT

The shoot apical meristem (SAM) undergoes developmental transitions that include a shift from vegetative to reproductive growth. This transition is triggered by flowering time genes, which up-regulate floral meristem (FM) identity genes that, in turn, control flower development by activating floral organ identity genes. This cascade of transcriptional activation is refined by repression mechanisms that temporally and spatially restrict gene expression to ensure proper development. Here, we demonstrate that HISTONE DEACETYLASE 19 (HDA19) maintains the identity of the reproductive SAM, or inflorescence meristem (IM), late in Arabidopsis thaliana development. At late stages of growth, hda19 IMs display a striking patterning defect characterized by ectopic expression of floral organ identity genes and the replacement of flowers with individual stamenoid organs. We further show that the flowering time gene FD has a specific function in this regulatory process, as fd hastens the emergence of these patterning defects in hda19 growth. Our work therefore identifies a new role for FD in reproductive patterning, as FD regulates IM function together with HDA19 in an age-dependent fashion. To effect these abnormalities, hda19 and fd may accentuate the weakening of transcriptional repression that occurs naturally with reproductive meristem proliferation.


Subject(s)
Arabidopsis Proteins/genetics , Arabidopsis/genetics , Flowers/growth & development , Gene Expression Regulation, Plant , Histone Deacetylases/genetics , Transcription Factors/genetics , Arabidopsis/growth & development , Arabidopsis/metabolism , Arabidopsis Proteins/metabolism , Flowers/genetics , Flowers/metabolism , Histone Deacetylases/metabolism , Longevity , Meristem/genetics , Meristem/physiology , Reproduction/genetics , Transcription Factors/metabolism
2.
Int J Mol Sci ; 20(1)2018 Dec 28.
Article in English | MEDLINE | ID: mdl-30597831

ABSTRACT

As multicellular organisms grow, spatial and temporal patterns of gene expression are strictly regulated to ensure that developmental programs are invoked at appropriate stages. In this work, we describe a putative transcriptional regulator in Arabidopsis, TACO LEAF (TCO), whose overexpression results in the ectopic activation of reproductive genes during vegetative growth. Isolated as an activation-tagged allele, tco-1D displays gene misexpression and phenotypic abnormalities, such as curled leaves and early flowering, characteristic of chromatin regulatory mutants. A role for TCO in this mode of transcriptional regulation is further supported by the subnuclear accumulation patterns of TCO protein and genetic interactions between tco-1D and chromatin modifier mutants. The endogenous expression pattern of TCO and gene misregulation in tco loss-of-function mutants indicate that this factor is involved in seed development. We also demonstrate that specific serine residues of TCO protein are targeted by the ubiquitous kinase CK2. Collectively, these results identify TCO as a novel regulator of gene expression whose activity is likely influenced by phosphorylation, as is the case with many chromatin regulators.


Subject(s)
Arabidopsis Proteins/metabolism , Arabidopsis/genetics , Arabidopsis/metabolism , Casein Kinase II/metabolism , Gene Expression Regulation, Plant , Transcription Factors/metabolism , Amino Acid Sequence , Arabidopsis Proteins/chemistry , Arabidopsis Proteins/genetics , Chromatin/genetics , Chromatin/metabolism , Ectopic Gene Expression , Fluorescent Antibody Technique , Mutation , Organ Specificity/genetics , Phenotype , Phosphorylation , Promoter Regions, Genetic , Protein Binding , Reproduction/genetics , Seeds/genetics , Seeds/metabolism
3.
Curr Biol ; 27(19): 2940-2950.e4, 2017 Oct 09.
Article in English | MEDLINE | ID: mdl-28943086

ABSTRACT

The flattening of leaves to form broad blades is an important adaptation that maximizes photosynthesis. However, the molecular mechanism underlying this process remains unclear. The WUSCHEL-RELATED HOMEOBOX (WOX) genes WOX1 and PRS are expressed in the leaf marginal domain to enable leaf flattening, but the nature of WOX expression establishment remains elusive. Here, we report that adaxial-expressed MONOPTEROS (MP) and abaxial-enriched auxin together act as positional cues for patterning the WOX domain. MP directly binds to the WOX1 and PRS promoters and activates their expression. Furthermore, redundant abaxial-enriched ARF repressors suppress WOX1 and PRS expression, also through direct binding. In particular, we show that ARF2 is redundantly required with ARF3 and ARF4 to maintain the abaxial identity. Taken together, these findings explain how adaxial-abaxial polarity patterns the mediolateral axis and subsequent lateral expansion of leaves.


Subject(s)
Arabidopsis Proteins/genetics , Arabidopsis/genetics , DNA-Binding Proteins/genetics , Homeodomain Proteins/genetics , Indoleacetic Acids/metabolism , Plant Growth Regulators/metabolism , Signal Transduction , Transcription Factors/genetics , Arabidopsis/growth & development , Arabidopsis/metabolism , Arabidopsis Proteins/metabolism , DNA-Binding Proteins/metabolism , Homeodomain Proteins/metabolism , Plant Leaves/genetics , Plant Leaves/growth & development , Plant Leaves/metabolism , Transcription Factors/metabolism
4.
Plant Signal Behav ; 12(4): e1307492, 2017 04 03.
Article in English | MEDLINE | ID: mdl-28340328

ABSTRACT

Plants react to environmental cues by altering their growth and development, which can include organ tropic responses. These differential growth responses are triggered by the hormone auxin, and AUXIN RESPONSE FACTORs (ARFs) have been implicated in numerous organ tropisms in Arabidopsis thaliana. Surprisingly, despite being critical for light capture and overall plant morphology, inflorescence stem tropic responses remain relatively understudied, with presumed direct links to ARF function yet to be established. Here, we show that the expression patterns of ARF5/MONOPTEROS and ARF7/NONPHOTOTROPIC HYPOCOTYL4 are consistent with roles in inflorescence stem tropisms. Mutation of these factors does not alter inflorescence stem responses to gravity or unilateral auxin application, meaning their participation in these processes is presumably masked by functional redundancies. Future resolution of these redundancies will likely require higher order arf mutant combinations, guided by detailed expression analyses of ARFs in the inflorescence stem.


Subject(s)
Arabidopsis Proteins/metabolism , Arabidopsis/metabolism , Plant Roots/metabolism , Arabidopsis/genetics , Arabidopsis Proteins/genetics , DNA-Binding Proteins/genetics , DNA-Binding Proteins/metabolism , Gene Expression Regulation, Plant/genetics , Gene Expression Regulation, Plant/physiology , Indoleacetic Acids/metabolism , Inflorescence/genetics , Inflorescence/physiology , Plant Roots/genetics , Signal Transduction/genetics , Signal Transduction/physiology , Transcription Factors/genetics , Transcription Factors/metabolism
5.
New Phytol ; 212(1): 42-50, 2016 10.
Article in English | MEDLINE | ID: mdl-27441727

ABSTRACT

The regulatory effect auxin has on its own transport is critical in numerous self-organizing plant patterning processes. However, our understanding of the molecular mechanisms linking auxin signal transduction and auxin transport is still fragmentary, and important regulatory genes remain to be identified. To track a key link between auxin signaling and auxin transport in development, we established an Arabidopsis thaliana genetic background in which fundamental patterning processes in both shoot and root were essentially abolished and the expression of PIN FORMED (PIN) auxin efflux facilitators was dramatically reduced. In this background, we demonstrate that activating a steroid-inducible variant of the auxin response factor (ARF) MONOPTEROS (MP) is sufficient to restore patterning and PIN gene expression. Further, we show that MP binds to distinct promoter elements of multiple genetically defined PIN genes. Our work identifies a direct regulatory link between central, well-characterized genes involved in auxin signal transduction and auxin transport. The steroid-inducible MP system directly demonstrates the importance of this molecular link in multiple patterning events in embryos, shoots and roots, and provides novel options for interrogating the properties of self-regulated auxin-based patterning in planta.


Subject(s)
Arabidopsis Proteins/genetics , Arabidopsis/genetics , DNA-Binding Proteins/metabolism , Genes, Plant , Meristem/genetics , Organogenesis/genetics , Plant Roots/metabolism , Plant Shoots/metabolism , Transcription Factors/metabolism , Arabidopsis/metabolism , Arabidopsis Proteins/metabolism , Biological Transport , DNA-Binding Proteins/genetics , Gene Expression Regulation, Plant , Indoleacetic Acids/metabolism , Meristem/growth & development , Meristem/metabolism , Plant Roots/growth & development , Plant Shoots/growth & development , Protein Binding/genetics , Signal Transduction , Transcription Factors/genetics
6.
Plant Signal Behav ; 10(4): e992748, 2015.
Article in English | MEDLINE | ID: mdl-25830553

ABSTRACT

The current model of auxin-inducible transcription describes numerous regulatory interactions between AUXIN RESPONSE FACTORs (ARFs) and Aux/IAAs. However, specific relationships between individual members of these families in planta remain largely uncharacterized. Using a systems biology approach, the entire suite of Aux/IAA genes directly regulated by the developmentally pivotal ARF MONOPTEROS (MP) was recently determined for multiple Arabidopsis tissue types. This study showed that MP directly targets distinct subclades of Aux/IAAs, revealing potential regulatory modules of redundantly acting Aux/IAAs involved in MP-dependent processes. Further, functional analyses indicated that the protein products of these targeted Aux/IAAs negatively feedback on MP. Thus, comprehensive identification of Aux/IAAs targeted by individual ARFs will generate biologically meaningful networks of ARF-Aux/IAA regulatory modules controlling distinct plant pathways.


Subject(s)
Arabidopsis Proteins/metabolism , Arabidopsis/growth & development , Indoleacetic Acids/metabolism , Plant Development , Arabidopsis/genetics , Arabidopsis Proteins/genetics , Gene Expression Regulation, Plant , Models, Biological , Plant Development/genetics
7.
New Phytol ; 204(3): 474-483, 2014 Nov.
Article in English | MEDLINE | ID: mdl-25145395

ABSTRACT

The regulatory interactions between AUXIN RESPONSE FACTORS (ARFs) and Aux/IAA repressors play a central role in auxin signal transduction. Yet, the systems properties of this regulatory network are not well established. We generated a steroid-inducible ARF5/MONOPTEROS (MP) transgenic background to survey the involvement of this factor in the transcriptional regulation of the entire Aux/IAA family in Arabidopsis thaliana. Target genes of ARF5/MP identified by this approach were confirmed by chromatin immunoprecipitation, in vitro gel retardation assays and gene expression analyses. Our study shows that ARF5/MP is indispensable for the correct regulation of nearly one-half of all Aux/IAA genes, and that these targets coincide with distinct subclades. Further, genetic analyses demonstrate that the protein products of multiple Aux/IAA targets negatively feed back onto ARF5/MP activity. This work indicates that ARF5/MP broadly influences the expression of the Aux/IAA gene family, and suggests that such regulation involves the activation of specific subsets of redundantly functioning factors. These groups of factors may then act together to control various processes within the plant through negative feedback on ARF5. Similar detailed analyses of other Aux/IAA-ARF regulatory modules will be required to fully understand how auxin signal transduction influences virtually every aspect of plant growth and development.


Subject(s)
Arabidopsis Proteins/metabolism , Arabidopsis/metabolism , DNA-Binding Proteins/metabolism , Gene Expression Regulation, Plant/physiology , Indoleacetic Acids/metabolism , Transcription Factors/metabolism , Arabidopsis/drug effects , Arabidopsis/genetics , Arabidopsis Proteins/genetics , Chromatin Immunoprecipitation , DNA-Binding Proteins/genetics , Dexamethasone/pharmacology , Gene Expression Regulation, Plant/drug effects , Multigene Family , Plants, Genetically Modified , Seedlings/growth & development , Transcription Factors/genetics
8.
Curr Biol ; 24(13): 1518-24, 2014 Jul 07.
Article in English | MEDLINE | ID: mdl-24954045

ABSTRACT

Circadian clocks allow organisms to anticipate daily changes in the environment to enhance overall fitness. Transcription factors (TFs) play a prominent role in the molecular mechanism but are incompletely described possibly due to functional redundancy, gene family proliferation, and/or lack of context-specific assays. To overcome these, we performed a high-throughput yeast one-hybrid screen using the LUX ARRYHTHMO (LUX) gene promoter as bait against an Arabidopsis TF library. LUX is a unique gene because its mutation causes severe clock defects and transcript maintains high-amplitude cycling in the cold. We report the well-characterized cold-inducible C-repeat (CRT)/drought-responsive element (DRE) binding factor CBF1/DREB1b is a transcriptional regulator of LUX. We show that CBF1 binds the CRT in the LUX promoter, and both genes overlap in temporal and spatial expression. CBF1 overexpression causes upregulation of LUX and also alters other clock gene transcripts. LUX promoter regions including the CRT and Evening Element (EE) are sufficient for high-amplitude transcriptional cycling in the cold, and cold-acclimated lux seedlings are sensitive to freezing stress. Our data show cold signaling is integrated into the clock by CBF-mediated regulation of LUX expression, thereby defining a new transcriptional mechanism for temperature input to the circadian clock.


Subject(s)
Arabidopsis Proteins/metabolism , Arabidopsis/metabolism , Circadian Clocks/physiology , Cold Temperature , Gene Expression Regulation, Plant/physiology , Trans-Activators/metabolism , Transcription Factors/metabolism , Arabidopsis/genetics , Gene Expression Regulation, Plant/genetics , Gene Library , High-Throughput Screening Assays , Two-Hybrid System Techniques
9.
Development ; 139(22): 4180-90, 2012 Nov.
Article in English | MEDLINE | ID: mdl-23034631

ABSTRACT

The development and coordination of complex tissues in eukaryotes requires precise spatial control of fate-specifying genes. Although investigations of such control have traditionally focused on mechanisms of transcriptional activation, transcriptional repression has emerged as being equally important in the establishment of gene expression territories. In the angiosperm flower, specification of lateral organ fate relies on the spatial regulation of the ABC floral organ identity genes. Our understanding of how the boundaries of these expression domains are controlled is not complete. Here, we report that the A-class organ identity gene APETALA2 (AP2), which is known to repress the C-class gene AGAMOUS, also regulates the expression borders of the B-class genes APETALA3 and PISTILLATA, and the E-class gene SEPALLATA3. We show that AP2 represses its target genes by physically recruiting the co-repressor TOPLESS and the histone deacetylase HDA19. These results demonstrate that AP2 plays a broad role in flower development by controlling the expression domains of numerous floral organ identity genes.


Subject(s)
Arabidopsis Proteins/metabolism , Arabidopsis/genetics , Arabidopsis/metabolism , Histone Deacetylases/metabolism , Homeodomain Proteins/metabolism , Nuclear Proteins/metabolism , AGAMOUS Protein, Arabidopsis/genetics , AGAMOUS Protein, Arabidopsis/metabolism , Arabidopsis/growth & development , Arabidopsis Proteins/genetics , Flowers/genetics , Flowers/growth & development , Gene Expression Regulation, Developmental , Gene Expression Regulation, Plant , Homeodomain Proteins/genetics , MADS Domain Proteins/genetics , MADS Domain Proteins/metabolism , Plants, Genetically Modified , Transcription Factors/genetics
10.
Plant Signal Behav ; 7(8): 1027-30, 2012 Aug.
Article in English | MEDLINE | ID: mdl-22827953

ABSTRACT

The molecularly well-characterized auxin signal transduction pathway involves two evolutionarily conserved families interacting through their C-terminal domains III and IV: the Auxin Response Factors (ARFs) and their repressors the Aux/IAAs, to control auxin-responsive genes, among them genes involved in auxin transport. ( 1) (,) ( 2) We have developed a new genetic tool to study ARF function. Using MONOPTEROS (MP)/ARF5, we have generated a truncated version of MP (MPΔ), ( 3) which has lost the target domains for repression by Aux/IAA proteins. Besides exploring genetic interactions between MP and Aux/IAAs, we used this construct to trace MP's role in vascular patterning, a previously characterized auxin dependent process. ( 4) (,) ( 5) Here we summarize examples of naturally occurring truncated ARFs and summarize potential applications of truncated ARFs as analytical tools.


Subject(s)
Gene Expression Regulation, Plant , Indoleacetic Acids/metabolism , Plant Proteins/genetics , Plant Proteins/metabolism , Amino Acid Sequence , Evolution, Molecular , Molecular Sequence Data , Plant Proteins/chemistry , Plant Vascular Bundle/growth & development , Plant Vascular Bundle/metabolism , Sequence Deletion
11.
Plant Signal Behav ; 7(8): 940-3, 2012 Aug.
Article in English | MEDLINE | ID: mdl-22751359

ABSTRACT

The establishment of adaxial-abaxial polarity in plant lateral organs involves elaborate interactions between members of several transcription factor families, including the Auxin Response Factors (ARFs). We previously described a dominant allele of ARF5/MONOPTEROS (MP), termed MPΔ, which causes severe vascular hypertrophy in shoot lateral organs. Here we report that these organs are also disrupted in adaxial-abaxial polarity. Other MPΔ lateral organs with decreased vasculature show similar disruptions, suggesting that MP impinges on organ polarity through pathways separate from its role in promoting vascularization. Furthermore, we demonstrate that MPΔ exhibits an adaxial-abaxial asymmetry in its ability to influence organ development. Since ARFs previously implicated in polarity establishment function as transcriptional repressors, the transcriptional activator MP represents a novel link between auxin signal transduction and adaxial-abaxial polarity.


Subject(s)
Arabidopsis Proteins/metabolism , Arabidopsis/growth & development , Arabidopsis/genetics , Body Patterning/genetics , DNA-Binding Proteins/metabolism , Genes, Dominant/genetics , Mutation/genetics , Plant Shoots/growth & development , Transcription Factors/metabolism , Flowers/genetics , Flowers/growth & development , Organ Specificity , Plant Shoots/genetics , Plant Vascular Bundle/growth & development
12.
New Phytol ; 194(2): 391-401, 2012 Apr.
Article in English | MEDLINE | ID: mdl-22320407

ABSTRACT

Combinatorial interactions of AUXIN RESPONSE FACTORs (ARFs) and auxin/indole acetic acid (Aux/IAA) proteins through their common domains III and IV regulate auxin responses, but insight into the functions of individual proteins is still limited. As a new tool to explore this regulatory network, we generated a gain-of-function ARF genotype by eliminating domains III and IV from the functionally well-characterized ARF MONOPTEROS(MP)/ARF5. This truncated version of MP, termed MPΔ, conferred complementing MP activity, but also displayed a number of semi-dominant traits affecting auxin signaling and organ patterning. In MPΔ, the expression levels of many auxin-inducible genes, as well as rooting properties and vascular tissue abundance, were enhanced. Lateral organs were narrow, pointed and filled with parallel veins. This effect was epistatic over the vascular hypotrophy imposed by certain Aux/IAA mutations. Further, in MPΔ leaves, failure to turn off the procambium-selecting gene PIN1 led to the early establishment of oversized central procambial domains and very limited subsequent lateral growth of the leaf lamina. We conclude that MPΔ can selectively uncouple a single ARF from regulation by Aux/IAA proteins and can be used as a genetic tool to probe auxin pathways and explore leaf development.


Subject(s)
Arabidopsis Proteins/chemistry , Arabidopsis Proteins/metabolism , Arabidopsis/embryology , Body Patterning , DNA-Binding Proteins/chemistry , DNA-Binding Proteins/metabolism , Indoleacetic Acids/metabolism , Plant Leaves/embryology , Plant Vascular Bundle/embryology , Sequence Deletion/genetics , Transcription Factors/chemistry , Transcription Factors/metabolism , Alleles , Arabidopsis Proteins/genetics , DNA-Binding Proteins/genetics , Genetic Complementation Test , Green Fluorescent Proteins/metabolism , Models, Biological , Phenotype , Plant Leaves/anatomy & histology , Protein Structure, Tertiary , Signal Transduction , Structure-Activity Relationship , Transcription Factors/genetics , Transgenes/genetics
13.
Proc Natl Acad Sci U S A ; 107(52): 22734-9, 2010 Dec 28.
Article in English | MEDLINE | ID: mdl-21149702

ABSTRACT

A central question in developmental biology is how multicellular organisms coordinate cell division and differentiation to determine organ size. In Arabidopsis roots, this balance is controlled by cytokinin-induced expression of SHORT HYPOCOTYL 2 (SHY2) in the so-called transition zone of the meristem, where SHY2 negatively regulates auxin response factors (ARFs) by protein-protein interaction. The resulting down-regulation of PIN-FORMED (PIN) auxin efflux carriers is considered the key event in promoting differentiation of meristematic cells. Here we show that this regulation involves additional, intermediary factors and is spatio-temporally constrained. We found that the described cytokinin-auxin crosstalk antagonizes BREVIS RADIX (BRX) activity in the developing protophloem. BRX is an auxin-responsive target of the prototypical ARF MONOPTEROS (MP), a key promoter of vascular development, and transiently enhances PIN3 expression to promote meristem growth in young roots. At later stages, cytokinin induction of SHY2 in the vascular transition zone restricts BRX expression to down-regulate PIN3 and thus limit meristem growth. Interestingly, proper SHY2 expression requires BRX, which could reflect feedback on the auxin responsiveness of SHY2 because BRX protein can directly interact with MP, likely acting as a cofactor. Thus, cross-regulatory antagonism between BRX and SHY2 could determine ARF activity in the protophloem. Our data suggest a model in which the regulatory interactions favor BRX expression in the early proximal meristem and SHY2 prevails because of supplementary cytokinin induction in the later distal meristem. The complex equilibrium of this regulatory module might represent a universal switch in the transition toward differentiation in various developmental contexts.


Subject(s)
Arabidopsis Proteins/metabolism , Arabidopsis/metabolism , Meristem/metabolism , Plant Roots/metabolism , Arabidopsis/genetics , Arabidopsis/growth & development , Arabidopsis Proteins/genetics , Blotting, Western , Cytokinins/pharmacology , DNA-Binding Proteins/genetics , DNA-Binding Proteins/metabolism , Gene Expression Regulation, Developmental/drug effects , Gene Expression Regulation, Plant/drug effects , Glucuronidase/genetics , Glucuronidase/metabolism , Green Fluorescent Proteins/genetics , Green Fluorescent Proteins/metabolism , Indoleacetic Acids/pharmacology , Meristem/genetics , Meristem/growth & development , Microscopy, Confocal , Mutation , Nuclear Proteins/genetics , Nuclear Proteins/metabolism , Plant Growth Regulators/pharmacology , Plant Roots/genetics , Plant Roots/growth & development , Plants, Genetically Modified , Reverse Transcriptase Polymerase Chain Reaction , Time Factors , Transcription Factors/genetics , Transcription Factors/metabolism
14.
Curr Opin Plant Biol ; 12(5): 628-36, 2009 Oct.
Article in English | MEDLINE | ID: mdl-19700365

ABSTRACT

To ensure correct patterns of gene expression, eukaryotes use a variety of strategies to repress transcription. The transcriptional regulators mediating this repression can be broadly categorized as either passive or active repressors. While passive repressors rely on mechanisms such as steric hindrance of transcriptional activators to repress gene expression, active repressors display inherent repressive abilities commonly conferred by discrete repression domains. Recent studies have indicated that both categories of regulators function in a variety of plant processes, including hormone signal transduction, developmental pathways, and response to biotic and abiotic stresses.


Subject(s)
Gene Expression Regulation, Plant , Plant Development , Repressor Proteins/metabolism , Transcription, Genetic , Gene Expression Regulation, Developmental , Histone Deacetylases/metabolism , Plant Growth Regulators/metabolism , Signal Transduction , Stress, Physiological
15.
Development ; 134(14): 2561-7, 2007 Jul.
Article in English | MEDLINE | ID: mdl-17553903

ABSTRACT

AUXIN RESPONSE FACTOR (ARF)-mediated signaling conveys positional information during embryonic and postembryonic organogenesis and mutations in MONOPTEROS (MP/ARF5) result in severe patterning defects during embryonic and postembryonic development. Here we show that MP patterning activity is largely dispensable when the presumptive carboxypeptidase ALTERED MERISTEM PROGRAM 1 (AMP1) is not functional, indicating that MP is primarily necessary to counteract AMP1 activity. Closer inspection of the single and double mutant phenotypes reveals antagonistic influences of both genes on meristematic activities throughout the Arabidopsis life cycle. In the absence of MP activity, cells in apical meristems and along the paths of procambium formation acquire differentiated identities and this is largely dependent on differentiation-promoting AMP1 activity. Positions of antagonistic interaction between MP and AMP1 coincide with MP expression domains within the larger AMP1 expression domain. These observations suggest a model in which auxin-derived positional information through MP carves out meristematic niches by locally overcoming a general differentiation-promoting activity involving AMP1.


Subject(s)
Arabidopsis Proteins/metabolism , Arabidopsis/embryology , Carboxypeptidases/metabolism , DNA-Binding Proteins/metabolism , Meristem/embryology , Transcription Factors/metabolism , Arabidopsis/growth & development , Arabidopsis/metabolism , Arabidopsis Proteins/genetics , Carboxypeptidases/genetics , Cell Differentiation/physiology , Cell Lineage/physiology , Cotyledon/cytology , Cotyledon/embryology , Cotyledon/growth & development , Cotyledon/metabolism , DNA-Binding Proteins/genetics , Indoleacetic Acids/metabolism , Meristem/cytology , Meristem/growth & development , Meristem/metabolism , Mutation , Plant Roots/cytology , Plant Roots/embryology , Plant Roots/growth & development , Plant Roots/metabolism , Protein Binding , Transcription Factors/genetics
16.
J Cell Biochem ; 100(3): 642-52, 2007 Feb 15.
Article in English | MEDLINE | ID: mdl-16983699

ABSTRACT

Here, we report the use of an in vivo protein-protein interaction detection approach together with focused follow-up experiments to study the function of the DeaD protein in Escherichia coli. In this method, functions are assigned to proteins based on the interactions they make with others in the living cell. The assigned functions are further confirmed using follow-up experiments. The DeaD protein has been characterized in vitro as a putative prokaryotic factor required for the formation of translation initiation complexes on structured mRNAs. Although the RNA helicase activity of DeaD has been demonstrated in vitro, its in vivo activity remains controversial. Here, using a method called sequential peptide affinity (SPA) tagging, we show that DeaD interacts with certain ribosomal proteins as well as a series of other nucleic acid binding proteins. Focused follow-up experiments provide evidence for the mRNA helicase activity of the DeaD protein complex during translation initiation. DeaD overexpression compensates for the reduction of the translation activity caused by a structure placed at the initiation region of a chloramphenicol acetyltransferase gene (cat) used as a reporter. Deletion of the deaD gene, encoding DeaD, abolishes the translation activity of the mRNA with an inhibitory structure at its initiation region. Increasing the growth temperature disrupts RNA secondary structures and bypasses the DeaD requirement. These observations suggest that DeaD is involved in destabilizing mRNA structures during translation initiation. This study also provides further confirmation that large-scale protein-protein interaction data can be suitable to study protein functions in E. coli.


Subject(s)
Chloramphenicol O-Acetyltransferase/genetics , DEAD-box RNA Helicases/physiology , Escherichia coli Proteins/physiology , Protein Biosynthesis , RNA, Messenger/genetics , Base Sequence , DEAD-box RNA Helicases/genetics , DEAD-box RNA Helicases/metabolism , DNA Primers , Electrophoresis, Polyacrylamide Gel , Escherichia coli/genetics , Escherichia coli Proteins/genetics , Escherichia coli Proteins/metabolism , Nucleic Acid Conformation , Protein Binding , RNA, Messenger/chemistry
17.
Curr Opin Plant Biol ; 7(5): 553-63, 2004 Oct.
Article in English | MEDLINE | ID: mdl-15337098

ABSTRACT

The extremely wide spectrum of the plant processes that are influenced by auxin raises the question of how signals conveyed by a single molecule can trigger such a variety of responses. Although many aspects of auxin function remain elusive, others have become genetically tractable. The identification of crucial genes in auxin signal transduction and auxin transport in the past few years has led to molecularly testable concepts of how auxin signals regulate gene activities in individual cells, and how the polar transport of auxin could impact on patterning processes throughout the plant.


Subject(s)
Arabidopsis/metabolism , Gene Expression Regulation, Plant , Genes, Plant , Indoleacetic Acids/metabolism , Signal Transduction , Arabidopsis Proteins/metabolism , Biological Transport , DNA-Binding Proteins/metabolism , Models, Biological , Nuclear Proteins/metabolism
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