Gilberto Sachetto-Martins

Signals from Chloroplasts Converge to Regulate Nuclear Gene Expression

One proposed strategy for controlling the transmission of insect-borne pathogens uses a drive mechanism to ensure the rapid spread of transgenes conferring disease refractoriness throughout wild populations. Here, we report the creation of maternal-effect selfish genetic elements in Drosophila that drive population replacement and are resistant to recombination-mediated dissociation of drive and disease refractoriness functions. These selfish elements use microRNA-mediated silencing of a maternally expressed gene essential for embryogenesis, which is coupled with early zygotic expression of a rescuing transgene.The phosphoinositide phosphatase PTEN is mutated in many human cancers. Although the role of PTEN has been studied extensively, the relative contributions of its numerous potential downstream effectors to deregulated growth and tumorigenesis remain uncertain. We provide genetic evidence in Drosophila melanogaster for the paramount importance of the protein kinase Akt [also called protein kinase B (PKB)] in mediating the effects of increased phosphatidylinositol 3,4,5-trisphosphate (PIP3) concentrations that are caused by the loss of PTEN function. A mutation in the pleckstrin homology (PH) domain of Akt that reduces its affinity for PIP3 sufficed to rescue the lethality of flies devoid of PTEN activity. Thus, Akt appears to be the only critical target activated by increased PIP3 concentrations in Drosophila.Using genomic and mass spectrometry-based proteomic methods, we evaluated gene expression, identified key activities, and examined partitioning of metabolic functions in a natural acid mine drainage (AMD) microbial biofilm community. We detected 2033 proteins from the five most abundant species in the biofilm, including 48% of the predicted proteins from the dominant biofilm organism, Leptospirillum group II. Proteins involved in protein refolding and response to oxidative stress appeared to be highly expressed, which suggests that damage to biomolecules is a key challenge for survival. We validated and estimated the relative abundance and cellular localization of 357 unique and 215 conserved novel proteins and determined that one abundant novel protein is a cytochrome central to iron oxidation and AMD formation.

Plant glycine-rich proteins: a family or just proteins with a common motif?

Twelve years ago a set of glycine-rich proteins (GRP) of plants were characterized and since then a wealth of new GRPs have been identified. The highly specific but diverse expression pattern of grp genes, taken together with the distinct sub-cellular localisation of some GRP groups, clearly indicate that these proteins are implicated in several independent physiological processes. Notwithstanding the absence of a clear definition of the role of GRPs in plant cells, studies conducted with these proteins have provided new and interesting insights on the molecular and cell biology of plants. Complex regulated promoters and distinct mechanisms of gene expression regulation have been demonstrated. New protein targeting pathways, as well as the exportation of GRPs from different cell types have been discovered. These data show that GRPs can be useful as markers and/or models to understand distinct aspects of plant biology. In this review, the structural and functional features of this family of plant proteins will be summarised. Special emphasis will be given to the gene expression regulation of GRPs isolated from different plant species, as it can help to unravel their possible biological functions.

Complete genome sequence of the sugarcane nitrogen-fixing endophyte Gluconacetobacter diazotrophicus Pal5

BackgroundGluconacetobacter diazotrophicus Pal5 is an endophytic diazotrophic bacterium that lives in association with sugarcane plants. It has important biotechnological features such as nitrogen fixation, plant growth promotion, sugar metabolism pathways, secretion of organic acids, synthesis of auxin and the occurrence of bacteriocins.ResultsGluconacetobacter diazotrophicus Pal5 is the third diazotrophic endophytic bacterium to be completely sequenced. Its genome is composed of a 3.9 Mb chromosome and 2 plasmids of 16.6 and 38.8 kb, respectively. We annotated 3,938 coding sequences which reveal several characteristics related to the endophytic lifestyle such as nitrogen fixation, plant growth promotion, sugar metabolism, transport systems, synthesis of auxin and the occurrence of bacteriocins. Genomic analysis identified a core component of 894 genes shared with phylogenetically related bacteria. Gene clusters for gum-like polysaccharide biosynthesis, tad pilus, quorum sensing, for modulation of plant growth by indole acetic acid and mechanisms involved in tolerance to acidic conditions were identified and may be related to the sugarcane endophytic and plant-growth promoting traits of G. diazotrophicus. An accessory component of at least 851 genes distributed in genome islands was identified, and was most likely acquired by horizontal gene transfer. This portion of the genome has likely contributed to adaptation to the plant habitat.ConclusionThe genome data offer an important resource of information that can be used to manipulate plant/bacterium interactions with the aim of improving sugarcane crop production and other biotechnological applications.

Functional diversity of the plant glycine-rich proteins superfamily.

The first plant glycine–rich proteins (GRPs) have been isolated more than 20 years ago based on their specific expression pattern and/or modulation by several biotic and abiotic factors. This superfamily is characterized by the presence of a glycine-rich domain arranged in (Gly)n-X repeats. The presence of additional motifs, as well as the nature of the glycine repeats, groups them in different classes. The diversity in structure as well as in expression pattern, modulation and sub-cellular localization have always indicated that these proteins, although classified as members of the same superfamily, would perform different functions in planta. Only now, two decades later, with the first functional characterizations of plant GRPs their involvement in diverse biological and biochemical processes are being uncovered. Here, we review the so far ascribed functions of plant GRPs.

AtGRP2, a cold-induced nucleo-cytoplasmic RNA-binding protein, has a role in flower and seed development

The glycine-rich protein AtGRP2 is one of the four members of the cold-shock domain (CSD) protein family in Arabidopsis. It is characterized by the presence of a nucleic acid-binding CSD domain, two glycine-rich domains and two CCHC zinc-fingers present in nucleic acid-binding proteins. In an attempt to further understand the role of CSD/GRP proteins in plants, we have proceeded to the functional characterization of the AtGRP2 gene. Here, we demonstrate that AtGRP2 is a nucleo-cytoplasmic protein involved in Arabidopsis development with a possible function in cold-response. Expression analysis revealed that the AtGRP2 gene is active in meristematic tissues, being modulated during flower development. Down-regulation of AtGRP2 gene, using gene-silencing techniques resulted in early flowering, altered stamen number and affected seed development. A possible role of AtGRP2 as an RNA chaperone is discussed.

New Insights into Aluminum Tolerance in Rice: The ASR5 Protein Binds the STAR1 Promoter and Other Aluminum-Responsive Genes

Aluminum (Al) toxicity in plants is one of the primary constraints in crop production. Al³⁺, the most toxic form of Al, is released into soil under acidic conditions and causes extensive damage to plants, especially in the roots. In rice, Al tolerance requires the ASR5 gene, but the molecular function of ASR5 has remained unknown. Here, we perform genome-wide analyses to identify ASR5-dependent Al-responsive genes in rice. Based on ASR5_RNAi silencing in plants, a global transcriptome analysis identified a total of 961 genes that were responsive to Al treatment in wild-type rice roots. Of these genes, 909 did not respond to Al in the ASR5_RNAi plants, indicating a central role for ASR5 in Al-responsive gene expression. Under normal conditions, without Al treatment, the ASR5_RNAi plants expressed 1.756 genes differentially compared to the wild-type plants, and 446 of these genes responded to Al treatment in the wild-type plants. Chromatin immunoprecipitation followed by deep sequencing identified 104 putative target genes that were directly regulated by ASR5 binding to their promoters, including the STAR1 gene, which encodes an ABC transporter required for Al tolerance. Motif analysis of the binding peak sequences revealed the binding motif for ASR5, which was confirmed via in vitro DNA-binding assays using the STAR1 promoter. These results demonstrate that ASR5 acts as a key transcription factor that is essential for Al-responsive gene expression and Al tolerance in rice.

NIK1-mediated translation suppression functions as a plant antiviral immunity mechanism

Plants and plant pathogens are subject to continuous co-evolutionary pressure for dominance, and the outcomes of these interactions can substantially impact agriculture and food security. In virus–plant interactions, one of the major mechanisms for plant antiviral immunity relies on RNA silencing, which is often suppressed by co-evolving virus suppressors, thus enhancing viral pathogenicity in susceptible hosts. In addition, plants use the nucleotide-binding and leucine-rich repeat (NB-LRR) domain-containing resistance proteins, which recognize viral effectors to activate effector-triggered immunity in a defence mechanism similar to that employed in non-viral infections. Unlike most eukaryotic organisms, plants are not known to activate mechanisms of host global translation suppression to fight viruses. Here we demonstrate in Arabidopsis that the constitutive activation of NIK1, a leucine-rich repeat receptor-like kinase (LRR-RLK) identified as a virulence target of the begomovirus nuclear shuttle protein (NSP), leads to global translation suppression and translocation of the downstream component RPL10 to the nucleus, where it interacts with a newly identified MYB-like protein, L10-INTERACTING MYB DOMAIN-CONTAINING PROTEIN (LIMYB), to downregulate translational machinery genes fully. LIMYB overexpression represses ribosomal protein genes at the transcriptional level, resulting in protein synthesis inhibition, decreased viral messenger RNA association with polysome fractions and enhanced tolerance to begomovirus. By contrast, the loss of LIMYB function releases the repression of translation-related genes and increases susceptibility to virus infection. Therefore, LIMYB links immune receptor LRR-RLK activation to global translation suppression as an antiviral immunity strategy in plants.

Survey of glycine-rich proteins (GRPs) in the Eucalyptus expressed sequence tag database (ForEST)

The occurrence of quasi-repetitive glycine-rich peptides has been reported in different organisms. Glycine-rich regions are proposed to be involved in protein-protein interactions in some mammalian protein families. In plants, a set of glycine-rich proteins (GRPs) was characterized several years ago, and since then a wealth of new GRPs have been identified. GRPs may have very diverse sub-cellular localization and functions. The only common feature among all different GRPs is the presence of glycine-rich repeat domains. The expression of genes encoding GRPs is developmentally regulated, and also induced, in several plant genera, by physical, chemical and biological factors. In addition to the highly modulated expression, several GRPs also show tissue-specific localization. GRPs specifically expressed in xylem, phloem, epidermis, anther tapetum and roots have been described. In this paper, the structural and functional features of these proteins in Eucalyptus are summarized. Since this is the first description of GRPs in this species, particular emphasis has been given to the expression pattern of these genes by analyzing their abundance and prevalence in the different cDNA-libraries of the Eucalyptus Genome Sequencing Project Consortium (ForEST). The comparison of GRPs from Eucalyptus and other species is also discussed.

AtGRP5, a vacuole-located glycine-rich protein involved in cell elongation

Although several glycine-rich protein (GRP) genes were isolated and characterized, very little is known about their function. The primary structure of AtGRP5 from Arabidopsis thaliana has a signal peptide followed by a region with high glycine content. In this work, green fluorescent protein fusions were obtained in order to characterize the sub-cellular localization of the AtGRP5 protein. The results indicated that this protein is the first described vacuolar GRP. Sense, antisense and RNAi transgenic A. thaliana plants were generated and analyzed phenotypically. Plants overexpressing AtGRP5 showed longer roots and an enhanced elongation of the inflorescence axis, while antisense and RNAi plants demonstrated the opposite phenotype. The analysis of a knockout T-DNA line corroborates the phenotypes obtained with the antisense and RNAi plants. Altogether, these results suggest that this vacuolar GRP could be involved in organ growth by promoting cell elongation processes.

The Arabidopsis translocator protein (AtTSPO) is regulated at multiple levels in response to salt stress and perturbations in tetrapyrrole metabolism

BackgroundThe translocator protein 18 kDa (TSPO), previously known as the peripheral-type benzodiazepine receptor (PBR), is important for many cellular functions in mammals and bacteria, such as steroid biosynthesis, cellular respiration, cell proliferation, apoptosis, immunomodulation, transport of porphyrins and anions. Arabidopsis thaliana contains a single TSPO/PBR-related gene with a 40 amino acid N-terminal extension compared to its homologs in bacteria or mammals suggesting it might be chloroplast or mitochondrial localized.ResultsTo test if the TSPO N-terminal extension targets it to organelles, we fused three potential translational start sites in the TSPO cDNA to the N-terminus of GFP (AtTSPO:eGFP). The location of the AtTSPO:eGFP fusion protein was found to depend on the translational start position and the conditions under which plants were grown. Full-length AtTSPO:eGFP fusion protein was found in the endoplasmic reticulum and in vesicles of unknown identity when plants were grown in standard conditions. However, full length AtTSPO:eGFP localized to chloroplasts when grown in the presence of 150 mM NaCl, conditions of salt stress. In contrast, when AtTSPO:eGFP was truncated to the second or third start codon at amino acid position 21 or 42, the fusion protein co-localized with a mitochondrial marker in standard conditions. Using promoter GUS fusions, qRT-PCR, fluorescent protein tagging, and chloroplast fractionation approaches, we demonstrate that AtTSPO levels are regulated at the transcriptional, post-transcriptional and post-translational levels in response to abiotic stress conditions. Salt-responsive genes are increased in a tspo-1 knock-down mutant compared to wild type under conditions of salt stress, while they are decreased when AtTSPO is overexpressed. Mutations in tetrapyrrole biosynthesis genes and the application of chlorophyll or carotenoid biosynthesis inhibitors also affect AtTSPO expression.ConclusionOur data suggest that AtTSPO plays a role in the response of Arabidopsis to high salt stress. Salt stress leads to re-localization of the AtTSPO from the ER to chloroplasts through its N-terminal extension. In addition, our results show that AtTSPO is regulated at the transcriptional level in tetrapyrrole biosynthetic mutants. Thus, we propose that AtTSPO may play a role in transporting tetrapyrrole intermediates during salt stress and other conditions in which tetrapyrrole metabolism is compromised.