Attallah C.V.,CONICET |
Welchen E.,CONICET |
Martin A.P.,Institute Biologia Molecular Y Celular Of Rosario |
Spinelli S.V.,Institute Biologia Molecular Y Celular Of Rosario |
And 3 more authors.
Journal of Experimental Botany | Year: 2011
Two Arabidopsis thaliana genes (HCC1 and HCC2), resulting from a duplication that took place before the emergence of flowering plants, encode proteins with homology to the SCO proteins involved in copper insertion during cytochrome c oxidase (COX) assembly in other organisms. Heterozygote HCC1 mutant plants produce 25% abnormal seeds with defective embryos arrested at the heart or torpedo stage. These embryos lack COX activity, suggesting that the requirement of HCC1 during the early stages of plant development is related with its COX assembly function. Homozygote HCC2 mutant plants develop normally and do not show changes in COX2 levels. These plants display increased sensitivity of root growth to increased copper and a higher expression of miR398 and other genes that respond to copper limitation, in spite of the fact that they have a higher copper content than the wild type. HCC2 mutant plants also show increased expression of stress-responsive genes. The results suggest that HCC1 is the protein involved in COX biogenesis and that HCC2, that lacks the cysteines and histidine putatively involved in copper binding, functions in copper sensing and redox homeostasis. In addition, plants that overexpress HCC1 have an altered response of root elongation to changes in copper in the growth medium and increased expression of two low-copper-responsive genes, suggesting that HCC1 may also have a role in copper homeostasis. © 2011 The Author.
Mehrnia M.,Max Planck Institute of Molecular Plant Physiology |
Balazadeh S.,Max Planck Institute of Molecular Plant Physiology |
Balazadeh S.,University of Potsdam |
Zanor M.-I.,Max Planck Institute of Molecular Plant Physiology |
And 3 more authors.
Plant Physiology | Year: 2013
We report about ERF BUD ENHANCER (EBE; At5g61890), a transcription factor that affects cell proliferation as well as axillary bud outgrowth and shoot branching in Arabidopsis (Arabidopsis thaliana). EBE encodes a member of the APETALA2/ETHYLENE RESPONSE FACTOR (AP2/ERF) transcription factor superfamily; the gene is strongly expressed in proliferating cells and is rapidly and transiently up-regulated in axillary meristems upon main stem decapitation. Overexpression of EBE promotes cell proliferation in growing calli, while the opposite is observed in EBE-RNAi lines. EBE overexpression also stimulates axillary bud formation and outgrowth, while repressing it results in inhibition of bud growth. Global transcriptome analysis of estradiol-inducible EBE overexpression lines revealed 48 EBE early-responsive genes, of which 14 were up-regulated and 34 were down-regulated. EBE activates several genes involved in cell cycle regulation and dormancy breaking, including D-type cyclin CYCD3;3, transcription regulator DPa, and BRCA1-ASSOCIATED RING DOMAIN1. Among the down-regulated genes were DORMANCY-ASSOCIATED PROTEIN1 (AtDRM1), AtDRM1 homolog, MEDIATOR OF ABA-REGULATED DORMANCY1, and ZINC FINGER HOMEODOMAIN5. Our data indicate that the effect of EBE on shoot branching likely results from an activation of genes involved in cell cycle regulation and dormancy breaking. © 2013 American Society of Plant Biologists. All Rights Reserved.
Mascali F.C.,Institute Biologia Molecular Y Celular Of Rosario |
Ching H.Y.V.,CNRS Institute of Integrative Biology |
Rasia R.M.,Institute Biologia Molecular Y Celular Of Rosario |
Un S.,CNRS Institute of Integrative Biology |
Tabares L.C.,CNRS Institute of Integrative Biology
Angewandte Chemie - International Edition | Year: 2016
Double electron–electron resonance (DEER) can be used to study the structure of a protein in its native cellular environment. Until now, this has required isolation, in vitro labeling, and reintroduction of the protein back into the cells. We describe a completely biosynthetic approach that avoids these steps. It exploits genetically encodable lanthanide-binding tags (LBT) to form self-assembling GdIII metal-based spin labels and enables direct in-cell measurements. This approach is demonstrated using a pair of LBTs encoded one at each end of a 3-helix bundle expressed in E. coli grown on GdIII-supplemented medium. DEER measurements directly on these cells produced readily detectable time traces from which the distance between the GdIII labels could be determined. This work is the first to use biosynthetically produced self-assembling metal-containing spin labels for non-disruptive in-cell structural measurements. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
Rodriguez R.E.,Institute Biologia Molecular Y Celular Of Rosario |
Mecchia M.A.,Institute Biologia Molecular Y Celular Of Rosario |
Debernardi J.M.,Institute Biologia Molecular Y Celular Of Rosario |
Schommer C.,Institute Biologia Molecular Y Celular Of Rosario |
And 2 more authors.
Development | Year: 2010
Cell proliferation is an important determinant of plant form, but little is known about how developmental programs control cell division. Here, we describe the role of microRNA miR396 in the coordination of cell proliferation in Arabidopsis leaves. In leaf primordia, miR396 is expressed at low levels that steadily increase during organ development. We found that miR396 antagonizes the expression pattern of its targets, the GROWTH-REGULATING FACTOR (GRF) transcription factors. miR396 accumulates preferentially in the distal part of young developing leaves, restricting the expression of GRF2 to the proximal part of the organ. This, in turn, coincides with the activity of the cell proliferation marker CYCLINB1;1. We show that miR396 attenuates cell proliferation in developing leaves, through the repression of GRF activity and a decrease in the expression of cell cycle genes. We observed that the balance between miR396 and the GRFs controls the final number of cells in leaves. Furthermore, overexpression of miR396 in a mutant lacking GRF-INTERACTING FACTOR 1 severely compromises the shoot meristem. We found that miR396 is expressed at low levels throughout the meristem, overlapping with the expression of its target, GRF2. In addition, we show that miR396 can regulate cell proliferation and the size of the meristem. Arabidopsis plants with an increased activity of the transcription factor TCP4, which reduces cell proliferation in leaves, have higher miR396 and lower GRF levels. These results implicate miR396 as a significant module in the regulation of cell proliferation in plants.
Abriata L.A.,Institute Biologia Molecular Y Celular Of Rosario |
Abriata L.A.,Ecole Polytechnique Federale de Lausanne |
M. Salverda M.L.,Wageningen University |
Tomatis P.E.,Institute Biologia Molecular Y Celular Of Rosario |
Tomatis P.E.,University of Zürich
FEBS Letters | Year: 2012
A dataset of TEM lactamase variants with different substrate and inhibition profiles was compiled and analyzed. Trends show that loops are the main evolvable regions in these enzymes, gradually accumulating mutations to generate increasingly complex functions. Notably, many mutations present in evolved enzymes are also found in simpler variants, probably originating functional promiscuity. Following a function-stability tradeoff, the increase in functional complexity driven by accumulation of mutations fosters the incorporation of other stability-restoring substitutions, although our analysis suggests they might not be as "global" as generally accepted and seem instead specific to different networks of protein sites. Finally, we show how this dataset can be used to model functional changes in TEMs based on the physicochemical properties of the amino acids. © 2012 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.
Asencion Diez M.D.,CONICET |
Peiru S.,Institute Biologia Molecular Y Celular Of Rosario |
Demonte A.M.,CONICET |
Gramajo H.,Institute Biologia Molecular Y Celular Of Rosario |
Journal of Bacteriology | Year: 2012
Streptomyces coelicolor exhibits a major secondary metabolism, deriving important amounts of glucose to synthesize pigmented antibiotics. Understanding the pathways occurring in the bacterium with respect to synthesis of oligo- and polysaccharides is of relevance to determine a plausible scenario for the partitioning of glucose-1-phosphate into different metabolic fates. We report the molecular cloning of the genes coding for UDP- and ADP-glucose pyrophosphorylases as well as for glycogen synthase from genomic DNA of S. coelicolor A3(2). Each gene was heterologously expressed in Escherichia coli cells to produce and purify to electrophoretic homogeneity the respective enzymes. UDP-glucose pyrophosphorylase (UDP-Glc PPase) was characterized as a dimer exhibiting a relatively high V max in catalyzing UDP-glucose synthesis (270 units/mg) and with respect to dTDP-glucose (94 units/mg). ADP-glucose pyrophosphorylase (ADP-Glc PPase) was found to be tetrameric in structure and specific in utilizing ATP as a substrate, reaching similar activities in the directions of ADP-glucose synthesis or pyrophosphorolysis (V max of 0.15 and 0.27 units/mg, respectively). Glycogen synthase was arranged as a dimer and exhibited specificity in the use of ADP-glucose to elongate α-1,4-glucan chains in the polysaccharide. ADP-Glc PPase was the only of the three enzymes exhibiting sensitivity to allosteric regulation by different metabolites. Mannose-6-phosphate, phosphoenolpyruvate, fructose-6-phosphate, and glucose-6-phosphate behaved as major activators, whereas NADPH was a main inhibitor of ADP-Glc PPase. The results support a metabolic picture where glycogen synthesis occurs via ADP-glucose in S. coelicolor, with the pathway being strictly regulated in connection with other routes involved with oligo- and polysaccharides, as well as with antibiotic synthesis in the bacterium. © 2012, American Society for Microbiology.
Abriata L.A.,Institute Biologia Molecular Y Celular Of Rosario
Concepts in Magnetic Resonance Part A: Bridging Education and Research | Year: 2012
This work describes two activities designed to introduce students into the use of NMR spectroscopy for the analysis of biological fluids. In the first activity, 1H spectra of common beverages are acquired and analyzed to identify their most abundant constituents based on computer-aided comparison against known spectra of metabolites. In the second activity, the utilization of glucose by yeast is followed by collecting 1H spectra over time, and the evolution of substrates and products is analyzed. The possibility of tailoring these activities to different levels and course scopes is discussed, together with potential alternative experiments and analyses. © 2012 Wiley Periodicals, Inc.
Teijeiro J.M.,National University of Rosario |
Teijeiro J.M.,CONICET |
Roldan M.L.,National University of Rosario |
Marini P.E.,National University of Rosario |
Marini P.E.,Institute Biologia Molecular Y Celular Of Rosario
Cell and Tissue Research | Year: 2016
In many mammals, upon entry into the female reproductive tract, a subpopulation of sperm is stored in the oviduct forming a functional reservoir. In the oviducts of pig and cow, Annexin A2 (AnxA2) has been linked to the binding of sperm. This protein may exist as a monomer or bound to S100A10 and both forms are associated with different biological functions. S100A10 has not yet been reported in the oviduct. The objective of this work is to analyze for the presence of S100A10 in the oviduct and to advance the study of AnxA2 and S100A10 in this organ. This work shows the presence of both proteins, AnxA2 and S100A10, in the oviduct of human, pig, cow, cat, dog and rabbit. At least in pig, AnxA2 is found devoid of S100A10 in the outer surface of the apical plasma membrane of oviductal epithelial cells, indicating that it binds to sperm as a monomer or in association with proteins different from S100A10. In the apical cytoplasm of pig oviductal epithelial cells, AnxA2 is associated with S100A10. In primary culture of porcine oviductal cells, the expression of ANXA2 is increased by progesterone, while the expression of S100A10 is increased by progesterone and estradiol. The widespread detection of both proteins in the oviduct of mammals indicates a probable conserved function in this organ. In summary, S100A10 and AnxA2 are widespread in the mammalian oviduct but AnxA2 binds sperm in vivo devoid of S100A10 and may be related to reservoir formation. © 2015, Springer-Verlag Berlin Heidelberg.
Suarez I.P.,Institute Biologia Molecular Y Celular Of Rosario |
Suarez I.P.,National University of Rosario |
Burdisso P.,Institute Biologia Molecular Y Celular Of Rosario |
Burdisso P.,National University of Rosario |
And 4 more authors.
Nucleic Acids Research | Year: 2015
DCL1 is the ribonuclease that carries out miRNA biogenesis in plants. The enzyme has two tandem double stranded RNA binding domains (dsRBDs) in its C-terminus. Here we show that the first of these domains binds precursor RNA fragments when isolated and cooperates with the second domain in the recognition of substrate RNA. Remarkably, despite showing RNA binding activity, this domain is intrinsically disordered. We found that it acquires a folded conformation when bound to its substrate, being the first report of a complete dsRBD folding upon binding. The free unfolded form shows tendency to adopt folded conformations, and goes through an unfolded bound state prior to the folding event. The significance of these results is discussed by comparison with the behavior of other dsRBDs. © 2015 The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.
Mateos J.L.,Institute Biologia Molecular Y Celular Of Rosario |
Bologna N.G.,Institute Biologia Molecular Y Celular Of Rosario |
Chorostecki U.,Institute Biologia Molecular Y Celular Of Rosario |
Palatnik J.F.,Institute Biologia Molecular Y Celular Of Rosario
Current Biology | Year: 2010
MicroRNAs (miRNAs) are widespread posttranscriptional regulators of gene expression. They are processed from longer primary transcripts that contain foldback structures (reviewed in [1, 2]). In animals, a complex formed by Drosha and DGCR8/Pasha recognizes the transition between the single-stranded RNA sequences and the stem loop to produce the first cleavage step in miRNA biogenesis . Whereas animal precursors are of uniform size and shape, their plant counterparts comprise a collection of variable stem loops, and little is known about the structural clues recognized during their processing. Here, we designed an unbiased approach based on the random mutagenesis of the MIR172a precursor to study miRNA processing in plants. Randomly mutated precursors were overexpressed in Arabidopsis, and their activity was determined in vivo. We gathered sequence data from these transgenes and used it to build a MIR172a precursor map highlighting relevant and neutral positions for its processing. A 15 nucleotide stem segment below the miRNA/miRNA* duplex was essential for MIR172a processing. In contrast, mutations in the terminal-loop region were mostly neutral, yet a loop was required for miR172 biogenesis. The results could be extended to other precursors, suggesting the existence of common features in at least part of the plant precursors. © 2010 Elsevier Ltd. All rights reserved.