The Arabidopsis SUPPRESSOR OF AUXIN RESISTANCE Proteins Are Nucleoporins with an Important Role in Hormone Signaling and Development (original) (raw)
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The Nuclear Pore Protein AtTPR Is Required for RNA Homeostasis, Flowering Time, and Auxin Signaling
PLANT PHYSIOLOGY, 2007
Nuclear pore complexes (NPCs) mediate the transport of RNA and other cargo between the nucleus and the cytoplasm. In vertebrates, the NPC protein TRANSLOCATED PROMOTER REGION (TPR) is associated with the inner filaments of the nuclear basket and is thought to serve as a scaffold for the assembly of transport machinery. In a screen for mutants that suppress the expression of the floral inhibitor FLOWERING LOCUS C, we identified lesions in the Arabidopsis (Arabidopsis thaliana) homolog of TPR (AtTPR). attpr mutants exhibit early-flowering and other pleiotropic phenotypes. A possible explanation for these developmental defects is that attpr mutants exhibit an approximately 8-fold increase in nuclear polyA RNA. Thus AtTPR is required for the efficient export of RNA from the nucleus. Microarray analysis shows that, in wild type, transcript abundance in the nuclear and total RNA pools are highly correlated; whereas, in attpr mutants, a significantly larger fraction of transcripts is enric...
Reduced expression of AtNUP62 nucleoporin gene affects auxin response in Arabidopsis
BMC Plant Biology, 2016
Background: The plant nuclear pore complex has strongly attracted the attention of the scientific community during the past few years, in particular because of its involvement in hormonal and pathogen/symbiotic signalling. In Arabidopsis thaliana, more than 30 nucleoporins have been identified, but only a few of them have been characterized. Among these, AtNUP160, AtNUP96, AtNUP58, and AtTPR have been reported to modulate auxin signalling, since corresponding mutants are suppressors of the auxin resistance conferred by the axr1 (auxin-resistant) mutation. The present work is focused on AtNUP62, which is essential for embryo and plant development. This protein is one of the three nucleoporins (with AtNUP54 and AtNUP58) of the central channel of the nuclear pore complex. Results: AtNUP62 promoter activity was detected in many organs, and particularly in the embryo sac, young germinating seedlings and at the adult stage in stipules of cauline leaves. The atnup62-1 mutant, harbouring a T-DNA insertion in intron 5, was identified as a knock-down mutant. It displayed developmental phenotypes that suggested defects in auxin transport or responsiveness. Atnup62 mutant plantlets were found to be hypersensitive to auxin, at the cotyledon and root levels. The phenotype of the AtNUP62-GFP overexpressing line further supported the existence of a link between AtNUP62 and auxin signalling. Furthermore, the atnup62 mutation led to an increase in the activity of the DR5 auxin-responsive promoter, and suppressed the auxin-resistant root growth and leaf serration phenotypes of the axr1 mutant. Conclusion: AtNUP62 appears to be a major negative regulator of auxin signalling. Auxin hypersensitivity of the atnup62 mutant, reminding that of atnup58 (and not observed with other nucleoporin mutants), is in agreement with the reported interaction between AtNUP62 and AtNUP58 proteins, and suggests closely related functions. The effect of AtNUP62 on auxin signalling likely occurs in relation to scaffold proteins of the nuclear pore complex (AtNUP160, AtNUP96 and AtTPR).
Arabidopsis TRANSCURVATA1 Encodes NUP58, a Component of the Nucleopore Central Channel
PLoS ONE, 2013
The selective trafficking of proteins and RNAs through the nuclear envelope regulates nuclear-cytoplasmic segregation of macromolecules and is mediated by nucleopore complexes (NPCs), which consist of about 400 nucleoporins (Nups) of about 30 types. Extensive studies of nucleoporin function in yeast and vertebrates showed that Nups function in nucleocytoplasmic trafficking and other processes. However, limited studies of plant Nups have identified only a few mutations, which cause pleiotropic phenotypes including reduced growth and early flowering. Here, we describe loss-offunction alleles of Arabidopsis TRANSCURVATA1 (TCU1); these mutations cause increased hypocotyl and petiole length, reticulate and asymmetrically epinastic leaf laminae of reduced size, and early flowering. TCU1 is transcribed in all of the organs and tissues examined, and encodes the putative ortholog of yeast and vertebrate Nup58, a nucleoporin of the Nup62 subcomplex. Nup58 forms the central channel of the NPC and acts directly in translocation of proteins through the nuclear envelope in yeast and vertebrates. Yeast two-hybrid (Y2H) assays identified physical interactions between TCU1/ NUP58 and 34 proteins, including nucleoporins, SCF (Skp1/Cul1/F-box) ubiquitin ligase complex components and other nucleoplasm proteins. Genetic interactions were also found between TCU1 and genes encoding nucleoporins, soluble nuclear transport receptors and components of the ubiquitin-proteasome and auxin signaling pathways. These genetic and physical interactions indicate that TCU1/NUP58 is a member of the Nup62 subcomplex of the Arabidopsis NPC. Our findings also suggest regulatory roles for TCU1/NUP58 beyond its function in nucleocytoplasmic trafficking, a hypothesis that is supported by the Y2H and genetic interactions that we observed. Citation: Ferrández-Ayela A, Alonso-Peral MM, Sánchez-García AB, Micol-Ponce R, Pérez-Pérez JM, et al. (2013) Arabidopsis TRANSCURVATA1 Encodes NUP58, a Component of the Nucleopore Central Channel. PLoS ONE 8(6): e67661.
The Plant Journal, 2012
In eukaryotic cells, transduction of external stimuli into the nucleus to induce transcription and export of mRNAs for translation in the cytoplasm is mediated by nuclear pore complexes (NPCs) composed of nucleoporin proteins (Nups). We previously reported that Arabidopsis MOS3, encoding the homolog of vertebrate Nup96, is required for plant immunity and constitutive resistance mediated by the de-regulated Toll interleukin 1 receptor/nucleotide-binding/leucine-rich repeat (TNL)-type R gene snc1. In vertebrates, Nup96 is a component of the conserved Nup107-160 nuclear pore sub-complex, and implicated in immunity-related mRNA export. Here, we used a reverse genetics approach to examine the requirement for additional subunits of the predicted Arabidopsis Nup107-160 complex in plant immunity. We show that, among eight putative complex members, beside MOS3, only plants with defects in Nup160 or Seh1 are impaired in basal resistance. Constitutive resistance in the snc1 mutant and immunity mediated by TNL-type R genes also depend on functional Nup160 and have a partial requirement for Seh1. Conversely, resistance conferred by coiled coil-type immune receptors operates largely independently of both genes, demonstrating specific contributions to plant defense signaling. Our functional analysis further revealed that defects in nup160 and seh1 result in nuclear accumulation of poly(A) mRNA, and, in the case of nup160, considerable depletion of EDS1, a key positive regulator of basal and TNL-triggered resistance. These findings suggest that Nup160 is required for nuclear mRNA export and full expression of EDS1-conditioned resistance pathways in Arabidopsis.
In plants, the activation of defense responses to pathogens is mediate by the recognition of elicitors produced by the pathogen (Pathogen-associated molecular patterns, PAMPs) or by the alteration of components of the plant itself (Damage-associated molecular patterns, DAMPs). This recognition induces a variety of defense responses and in the same time determines an inhibition of the developmental processes. A close relationship exists between the hormonal regulation of growth , developmental and defense processes. Auxin plays a central role in plant growth and development, regulating cellular division and elongation and triggering specific developmental events. The oligogalacturonides (OGs) deriving from the degradation of the pectic fraction of the plant cell wall are DAMPs able to activate the plant immune system. OGs also regulates growth and development of plant cells due to an antagonistic activity towards auxin. Plant expressing GUS reporter gene under the control of the synthetic auxin-responsive promoter DR5 shows an induction of the activity of the reporter gene in response to auxin treatments; this induction is decreased in response to auxin-OG co-treatment. In my PhD work I have studied IAA5 (INDOLE-3-ACETIC ACID INDUCIBLE 5), an auxin-inducible gene subjected to antagonism by the OGs. First I wanted to determine if the antagonism is played at the level of the promoter: Arabidopsis plants were transformed with a construct containing the GUS reporter gene under the control of the promoter of IAA5(PIAA5). Analysis of GUS expression through quantitative real-time PCR showed that GUS transcript was induced by auxin and that the level of induction was decreased in response to auxin-OG co-treatment showing that the antagonism between OGs and auxin takes place on the promoter of IAA5 . To identify the factors involved in the antagonism on PIAA5 and DR5 an in vitro approach called DNA affinity capture was used. Through this approach was possible to identify TGA7 (TGACG SEQUENCE-SPECIFIC BINDING PROTEIN 7), ARF5 (AUXIN RESPONSE FACTOR 5) e GT2 (AT-GT2) transcription factors as specific interactors of DR5 and PIAA5 , while TGA7 was identified as candidate for a role in the antagonism on the promoter of IAA5. Moreover a nuclear proteome analysis was performed to identify differentially expressed proteins in response to auxin or OG treatments and to auxin and OG co-treatments through a quantitative proteomic approach. It was possible to identify groups of proteins which the expression is positively regulated by auxin and also negatively regulated by the auxin and OG co-treatment. These proteins belongs to different functional classes, particularly, most of them has a role inoxidative stress response and splicing. Among the proteins that are up-regulated by OGs and down- regulated by auxin, we found different subunits of the COP9 signalosome and the related proteins CAND1 and DCAF1 suggesting a role for the COP9 signalosome in the OG- auxin antagonism.
The PS-IAA4/5-like Family of Early Auxin-inducible mRNAs in Arabidopsis thaliana
Journal of Molecular Biology, 1995
The plant hormone auxin transcriptionally activates early genes. We have isolated a 14-member family of DNA sequences complementary to indoleacetic acid (IAA)-inducible transcripts inArabidopsis thaliana. The corresponding genes,IAA1andIAA14, are homologs ofPS-1AA4/5andPS-IAA6from pea,AUX22andAUX28from soybean,ARG3andARG4from mungbean, andAtAux2-11andAtAux2-27fromArabidopsis. The members of the family are differentially expressed in matureArabidopsisplants. Characterization ofIAAgene expression in etiolated seedlings demonstrates specificity for auxin inducibility. The response of most family members to IAA is rapid (within 4 to 30 minutes) and insensitive to cyclohexamide. Cyclohexamide alone induces all the early genes. Auxin-induction of two late genes,IAA7andIAA8, is inhibited by cyclohexamide, indicating requirement of protein synthesis for their activation. AllIAAgenes display a biphasic dose response that is optimal at 10 μM IAA. However, individual genes respond differentially between 10 nM and 5μM IAA. Expression of all genes is defective in theArabidopsisauxin-resistant mutant linesaxr1, axr2,andaux1.The encoded polypeptides share four conserved domains, and seven invariant residues in the intervening regions. The spaces vary considerably in length, rendering the calculated molecular mass of IAA proteins to range from 19 kDa to 36 kDa. Overall sequence identity between members of the family is highly variable (36 to 87%). Their most significant structural features are functional nuclear transport signals, and a putative βαα-fold whose modeled three dimensional structure appears to be compatible with the prokaryotic β-ribbon DNA recognition motif. The data suggest that auxin induces in a differential and hierarchical fashion a large family of early genes that encode a structurally diverse class of nuclear proteins. These proteins are proposed to mediate tissue-specific and cell-type restricted responses to the hormone during plant growth and development.
Identification and localisation of a nucleoporin-like protein component of the plant nuclear matrix
Planta, 1992
Salt-detergent extraction of purified plant nuclei yields a fraction enriched in putative structural proteins known as the "nuclear matrix". Compared with mammalian nuclear matrices, which contain three major proteins, plant nuclear matrices are complex, containing at least 100 polypeptides. In order to characterise more fully the plant nuclear matrix we have used antibodies raised against both yeast (Saccharomyces cerevisiae) and mammalian (rat) nuclear pore proteins. We have shown that the nuclear matrix of carrot (Daucus carota L.) contains at least one nucleoporin-like protein of about 100 kDa which is immunologically related to both the yeast nuclear pore protein NSP1 and mammalian nucleoporins (p62). Antibody labelling of a variety of plant cells at the light-microscope and electron-microscope levels confirms that this antigen is located at the nuclear pores. This, to our knowledge, is the first identification of a nuclear pore protein in plants.
Nuclear RNA Export and Its Importance in Abiotic Stress Responses of Plants
Transduction of developmental and environmental cues into the nucleus to induce transcription and the export of RNAs to the cytoplasm through the nuclear pore complex (NPC) play pivotal roles in regulation of gene expression. The process of bulk export of mRNAs from nucleus to cytoplasm is highly conserved across eukaryotes. Assembly of export-competent mRNA ribonucleoprotein (mRNP) is coupled with both transcription and mRNA processing. The export-competent mRNP consists of mRNAs and a dozen nucleocytoplasmic shuttling nuclear proteins, including RNA export factors (Mex67-Mtr2 heterodimer, Npl3), poly(A)binding proteins, DEAD-box protein 5 (Dbp5), and nucleoporins (NUPs) in yeast. Mobile NUPs help docking of mRNP to the NPC nuclear basket. A partially
Auxin cross-talk: integration of signalling pathways to control plant development
Auxin Molecular Biology, 2002
We address here organellar genetic regulation and intercompartment genome coordination. We developed earlier a strategy relying on a tRNA-like shuttle to mediate import of nuclear transgene-encoded custom RNAs into mitochondria in plants. In the present work, we used this strategy to drive trans-cleaving hammerhead ribozymes into the organelles, to knock down specific mitochondrial RNAs and analyze the regulatory impact. In a similar approach, the tRNA mimic was used to import into mitochondria in Arabidopsis thaliana the orf77, an RNA associated with cytoplasmic male sterility in maize and possessing sequence identities with the atp9 mitochondrial RNA. In both cases, inducible expression of the transgenes allowed to characterise early regulation and signaling responses triggered by these respective manipulations of the organellar transcriptome. The results imply that the mitochondrial transcriptome is tightly controlled by a "bu↵ering" mechanism at the early and intermediate stages of plant development, a control that is released at later stages. On the other hand, high throughput analyses showed that knocking down a specific mitochondrial mRNA triggered a retrograde signaling and an anterograde nuclear transcriptome response involving a series of transcription factor genes and small RNAs. Our results strongly support transcriptome coordination mechanisms within the organelles and between the organelles and the nucleus.
A functional genetic assay for nuclear trafficking in plants
2007
Summary The receptor importin-α mediates the nuclear import of functionally diverse cargo proteins that contain arginine/lysine-rich nuclear localization signals (NLSs). Functional homologs of importin-α have been characterized in a wide range of species including yeast, human and plants. However, the differential cargo selectivity of plant importin-α homologs has not been established.