The Translationally Controlled Tumor Protein (TCTP) is a highly conserved protein at the level of sequence, considered to play an essential role in the regulation of growth and development in eukaryotes. However, this function has been inferred from studies in a few model systems, such as mice and mammalian cell lines, Drosophila and Arabidopsis. Thus, the knowledge regarding this protein is far from complete. In the present study bioinformatic analysis showed the presence of one or more TCTP genes per genome in plants with highly conserved signatures and subtle variations at the level of primary structure but with more noticeable differences at the level of predicted three-dimensional structures. These structures show differences in the "pocket" region close to the center of the protein and in its flexible loop domain. In fact, all predictive TCTP structures can be divided into two groups: (1) AtTCTP1-like and (2) CmTCTP-like, based on the predicted structures of an Arabidopsis TCTP and a Cucurbita maxima TCTP; according to this classification we propose that their probable function in plants may be inferred in principle. Thus, different TCTP genes in a single organism may have different functions; additionally, in those species harboring a single TCTP gene this could carry multiple functions. On the other hand, in silico analysis of AtTCTP1-like and CmTCTP-like promoters suggest that these share common motifs but with different abundance, which may underscore differences in their gene expression patterns. Finally, the absence of TCTP genes in most chlorophytes with the exception of Coccomyxa subellipsoidea, indicates that other proteins perform the roles played by TCTP or the pathways regulated by TCTP occur through alternative routes. These findings provide insight into the evolution of this gene family in plants.
Ectomycorrhizal symbiosis results in profound morphological and physiological modifications in both plant and fungus. This in turn is the product of differential gene expression in both co-symbionts, giving rise to specialized cell types capable of performing novel functions. During the precolonization stage, chemical signals from root exudates are sensed by the ectomycorrizal fungus, and vice versa, which are in principle responsible for the observed change in the developmental symbionts program. Little is known about the molecular mechanisms involved in the signaling and recognition between ectomycorrhizal fungi and their host plants. In the present work, we characterized a novel lactone, termed pinelactone, and identified a gene encoding for a histidine kinase in Pisolithus tictorius, which function is proposed to be the perception of the aforementioned metabolites. In this study, the use of closantel, a specific inhibitor of histidine kinase phosphorylation, affected the capacity for fungal colonization in the symbiosis between Pisolithus tinctorius and Pinus greggii, indicating that a 2-component system (TCS) may operate in the early events of plant-fungus interaction. Indeed, the metabolites induced the accumulation of Pisolithus tinctorius mRNA for a putative histidine kinase (termed Pthik1). Of note, Pthik1 was able to partially complement a S. cerevisiae histidine kinase mutant, demonstrating its role in the response to the presence of the aforementioned metabolites. Our results indicate a role of a 2-component pathway in the early stages of ectomycorrhizal symbiosis before colonization. Furthermore, a novel lactone from Pinus greggii root exudates may activate a signal transduction pathway that contributes to the establishment of the ectomycorrhizal symbiosis.
The causal agents of Citrus leprosis are viruses; however, extant diagnostic methods to identify them have failed to detect known viruses in orange, mandarin, lime and bitter orange trees with severe leprosis symptoms in Mexico, an important citrus producer. Using high throughput sequencing, a virus associated with citrus leprosis was identified, belonging to the proposed Dichorhavirus genus. The virus was termed Citrus Necrotic Spot Virus (CNSV) and contains two negative-strand RNA components; virions accumulate in the cytoplasm and are associated with plasmodesmata-channels interconnecting neighboring cells-suggesting a mode of spread within the plant. The present study provides insights into the nature of this pathogen and the corresponding plant response, which is likely similar to other pathogens that do not spread systemically in plants.
Plasmodium falciparum secretes a homologue of the translationally controlled tumor protein (TCTP) into serum of infected individuals, although its role in pathogenesis or virulence is unknown. To determine the effect of P. falciparum TCTP on B cells as compared to human TCTP, fluorescently labeled proteins were incubated on primary cultures of mouse splenic B cells and analyzed by flow cytometry and confocal microscopy. Our results indicate that both recombinant proteins are incorporated into B cells, but differ significantly in their rate and percentage of incorporation, being significantly higher for P. falciparum TCTP. Furthermore, P. falciparum TCTP showed a lower B cell proliferative effect than human TCTP, suggesting a mechanism through which the former could interfere in the host's immune response.
In higher plants, the phloem plays a central role in the delivery of nutrients and signals from source to sink tissues. These signals likely coordinate different aspects of plant development, as well as its response to environmental cues. Although some phloem-transported proteins and RNAs may function as signaling molecules in plants, their mode of action remains poorly understood. Previous analysis of transcripts from CMV-infected pumpkin (Cucurbita maxima cv Big Max) identified a Translationally-Controlled Tumor Protein (TCTP) mRNA homolog, designated CmTCTP. In the present work this transcript was analyzed in terms of its expression pattern. This RNA accumulates, both in healthy and CMV-infected plants, in developing and mature phloem in petiole and roots, as well as in apices at high levels. The protein was present at lower levels in most cell types, and almost no signal was detected in apices, suggesting translational regulation of this RNA. Additionally, CmTCTP harbored by Agrobacterium rhizogenes is capable of inducing whole plant regeneration. These data suggest a role for CmTCTP in growth regulation, possibly through long-distance signaling.
The phloem is the conduit through which photoassimilates are distributed from autotrophic to heterotrophic tissues and is involved in the distribution of signaling molecules that coordinate plant growth and responses to the environment. Phloem function depends on the coordinate expression of a large array of genes. We have previously identified conserved motifs in upstream regions of the Arabidopsis genes, encoding the homologs of pumpkin phloem sap mRNAs, displaying expression in vascular tissues. This tissue-specific expression in Arabidopsis is predicted by the overrepresentation of GA/CT-rich motifs in gene promoters. In this work we have searched for common motifs in upstream regions of the homologous genes from plants considered to possess a "primitive" vascular tissue (a lycophyte), as well as from others that lack a true vascular tissue (a bryophyte), and finally from chlorophytes. Both lycophyte and bryophyte display motifs similar to those found in Arabidopsis with a significantly low E-value, while the chlorophytes showed either a different conserved motif or no conserved motif at all. These results suggest that these same genes are expressed coordinately in non-vascular plants; this coordinate expression may have been one of the prerequisites for the development of conducting tissues in plants. We have also analyzed the phylogeny of conserved proteins that may be involved in phloem function and development. The presence of CmPP16, APL, FT, and YDA in chlorophytes suggests the recruitment of ancient regulatory networks for the development of the vascular tissue during evolution while OPS is a novel protein specific to vascular plants.
We utilized the two-compartment system to study the effect of arsenic (As) on the expression of the Glomus intraradices high-affinity phosphate transporter GiPT, and the GiArsA gene, a novel protein with a possible putative role as part of an arsenite efflux pump and similar to ArsA ATPase. Our results show that induction of GiPT expression correlates with As(V) uptake in the extra-radical mycelium of G. intraradices. We showed a time-concerted induction of transcript levels first of GiPT, followed by GiArsA, as well as the location of gene expression using laser microdissection of these two genes not only in the extra-radical mycelium but also in arbuscules. This work represents the first report showing the dissection of the molecular players involved in arbuscular mycorrhizal fungus (AMF)-mediated As tolerance in plants, and suggests that tolerance mediated by AMF may be caused by an As exclusion mechanism, where fungal structures such as the extra-radical mycelium and arbuscules may be playing an important role. Our results extend knowledge of the mechanisms underlying As efflux in arbuscular mycorrhizal fungi and mechanisms related to As tolerance.
Phloem-transported signals play an important role in regulating plant development and in orchestrating responses to environmental stimuli. Among such signals, phloem-mobile RNAs have been shown to play an important role as long-distance signaling agents. At maturity, angiosperm sieve elements are enucleate, and thus transcripts in the phloem translocation stream probably originate from the nucleate companion cells. In the present study, a pumpkin (Cucurbita maxima) phloem transcriptome was used to test for the presence of common motifs within the promoters of this unique set of genes, which may function to coordinate expression in cells of the vascular system. A bioinformatics analysis of the upstream sequences from 150 Arabidopsis genes homologous to members of the pumpkin phloem transcriptome identified degenerate sequences containing CT/GA- and GT/CA-rich motifs that were common to many of these promoters. Parallel studies performed on genes shown previously to be expressed in phloem tissues identified similar motifs. An expanded analysis, based on homologs of the pumpkin phloem transcriptome from cucumber (Cucumis sativus), identified similar sets of common motifs within the promoters of these genes. Promoter analysis offered support for the hypothesis that these motifs regulate expression within the vascular system. Our findings are discussed in terms of a role for these motifs in coordinating gene expression within the companion cell/sieve element system. These motifs could provide a useful bioinformatics tool for genome-wide screens on plants for which phloem tissues cannot readily be obtained.
The root system of transgenic tobacco plants expressing the enhanced green fluorescent protein (EGFP) under the control of the 35S cauliflower mosaic virus (CaMV) promoter, were colonized with the endomycorrhizal fungus Glomus intraradices. Translocation of EGFP protein from the root to the fungus was registered by light and confocal microscopy. Immunolocalization also showed the presence of EGFP in the mycelium of Glomus intraradices. Carboxyfluorescein feeding experiments on wild type mycorrhized plants evidenced the transport of fluorescein, a symplasmic tracer, from the plant to the fungus. Our results suggest that endomycorrhiza possess the capacity to exchange functional biological macromolecules as evidenced by the transport EGFP from the plant to the fungal symbiont.
PHYVV and PepGMV are plant viruses reported in Mexico and Southern US as causal agents of an important pepper disease known as "rizado amarillo". Mixed infections with PHYVV and PepGMV have been reported in several hosts over a wide geographic area. Previous work suggested that these viruses might interact at the replication and/or movement level in a complex manner. The aim of present report was to study some aspects of a synergistic interaction between PHYVV and PepGMV in pepper plants. These include analyses of symptom severity, viral DNA concentration and tissue localization of both viruses in single and mixed infections.
Influenza viruses pose a threat to human health because of their potential to cause global disease. Between mid March and mid April a pandemic influenza A virus emerged in Mexico. This report details 202 cases of infection of humans with the 2009 influenza A virus (H1N1)v which occurred in Mexico City as well as the spread of the virus throughout the entire country.
The Cfl xyn11A gene, encoding the endo-1,4-beta-xylanase Cfl Xyn11A from Cellulomonas flavigena, was isolated from a genomic DNA library. The open reading frame of the Cfl xyn11A gene was 999 base pairs long and encoded a polypeptide (Cfl Xyn11A) of 332 amino acids with a calculated molecular mass of 35,110Da. The Cfl xyn11A gene was expressed in Escherichia coli and the recombinant enzyme, with an estimated molecular weight of 31kDa was purified and xylanase activity was measured. Cfl Xyn11A showed optimal activity at pH 6.5 and 55 degrees C. The enzyme demonstrated moderate thermal stability as Cfl Xyn11A maintained 50% of its activity when incubated at 55 degrees C for 1h or at 45 degrees C for 6h. This is the first report describing the cloning, expression and functional characterization of an endo-1,4-beta-xylanase-encoding gene from C. flavigena. Cfl Xyn11A may be suitable for industrial applications in the food and feed industries, or in the pre-treatment of lignocellulosic biomass required to improve the yields of fermentable sugars for bioethanol production.
The aim of this study was to test the effect of acetosyringone (AS) on the accumulation of the histidine kinase coding mRNA, using in vitro propagated Pisolithus tinctorius. In the precontact phase of the ectomycorrhizal symbiosis, it is hypothesized that the plants releases chemicals, which are sensed by the fungal mycelium and in turn trigger the start of the symbiosis. The nature of such molecules is largely unknown; however, plant metabolite and their structural analogues have been widely used to induce infection in different microbe-plant interactions. A histidine kinase in Agrobacterium tumefaciens was activated by AS in the first step during the infection of dicotyledonous plants. A conserved gene fragment from the ectomycorrhizal fungus Pisolithus tinctorius was cloned using degenerate primers of conserved regions characteristic of the histidine kinase gene family. Then, the accumulation of this transcript in fungi incubated with AS was analyzed. RT-PCR and in situ hybridization suggest that this mRNA is synthesized in the presence of this plant analog in AS-treated mycelia. The findings presented here suggest a role of a histidine kinase involved in the early stages of ectomycorrhizal differentiation.
Tomato production in West Africa has been severely affected by begomovirus diseases, including yellow leaf curl and a severe symptom phenotype, characterized by extremely stunted and distorted growth and small deformed leaves. Here, a novel recombinant begomovirus from Mali, Tomato yellow leaf curl Mali virus (TYLCMLV), is described that, alone, causes tomato yellow leaf curl disease or, in combination with a betasatellite, causes the severe symptom phenotype. TYLCMLV is an Old World monopartite begomovirus with a hybrid genome composed of sequences from Tomato yellow leaf curl virus-Mild (TYLCV-Mld) and Hollyhock leaf crumple virus (HoLCrV). A TYLCMLV infectious clone induced leaf curl and yellowing in tomato, leaf curl, crumpling and yellowing in Nicotiana benthamiana and common bean, mild symptoms in N. glutinosa, and a symptomless infection in Datura stramonium. In a field-collected sample from a tomato plant showing the severe symptom phenotype in Mali, TYLCMLV was detected together with a betasatellite, identified as Cotton leaf curl Gezira betasatellite (CLCuGB). Tomato plants co-agroinoculated with TYLCMLV and CLCuGB developed severely stunted and distorted growth and small crumpled leaves. These symptoms were more severe than those induced by TYLCMLV alone, and were similar to the severe symptom phenotype observed in the field in Mali and in other West African countries. TYLCMLV and CLCuGB also induced more severe symptoms than TYLCMLV in the other solanaceous hosts, but not in common bean. The increased symptom severity was associated with hyperplasia of phloem-associated cells, but relatively little increase in TYLCMLV DNA levels. In surveys of tomato virus diseases in West Africa, TYLCMLV was commonly detected in plants with leaf curl and yellow leaf curl symptoms, whereas CLCuGB was infrequently detected and always in association with the severe symptom phenotype. Together, these results indicate that TYLCMLV causes tomato yellow leaf curl disease throughout West Africa, whereas TYLCMLV and CLCuGB represent a reassortant that causes the severe symptom phenotype in tomato.
Plant establishment, presence of arbuscular mycorrhizal fungi (AMF) and other rhizospheric fungi were studied in mine wastes from Zimapan, Hidalgo state, Mexico, using a holistic approach. Two long-term afforested and three non-afforested mine tailings were included in this research. Fifty-six plant species belonging to 29 families were successfully established on the afforested sites, while unmanaged tailings had only a few native plant species colonizing the surrounding soils. Almost all plant roots collected were associated to AMF in these sites. The genus Glomus was the most abundant AMF species found in their rhizosphere; however, the Acaulospora genus was also observed. Other rhizospheric fungi were identified by 18S rDNA sequencing analysis. Their role in these substrates, i.e. biocontrol, pollutant- and organic matter-degradation, and aides that increase plant metal tolerance is discussed. Our results advance the understanding of fungal diversity in sites polluted with metals and present alternative plants for remediation use.
RNA binding proteins (RBPs) are integral components of ribonucleoprotein (RNP) complexes and play a central role in RNA processing. In plants, some RBPs function in a non-cell-autonomous manner. The angiosperm phloem translocation stream contains a unique population of RBPs, but little is known regarding the nature of the proteins and mRNA species that constitute phloem-mobile RNP complexes. Here, we identified and characterized a 50-kD pumpkin (Cucurbita maxima cv Big Max) phloem RNA binding protein (RBP50) that is evolutionarily related to animal polypyrimidine tract binding proteins. In situ hybridization studies indicated a high level of RBP50 transcripts in companion cells, while immunolocalization experiments detected RBP50 in both companion cells and sieve elements. A comparison of the levels of RBP50 present in vascular bundles and phloem sap indicated that this protein is highly enriched in the phloem sap. Heterografting experiments confirmed that RBP50 is translocated from source to sink tissues. Collectively, these findings established that RBP50 functions as a non-cell-autonomous RBP. Protein overlay, coimmunoprecipitation, and cross-linking experiments identified the phloem proteins and mRNA species that constitute RBP50-based RNP complexes. Gel mobility-shift assays demonstrated that specificity, with respect to the bound mRNA, is established by the polypyrimidine tract binding motifs within such transcripts. We present a model for RBP50-based RNP complexes within the pumpkin phloem translocation stream.
Identified SSR markers ( Xcfd49 and Xbarc183 ) linked with stem rust resistance for efficient use in marker-assisted selection and stacking of resistance genes in wheat breeding programs. More than 80 % of the worldwide wheat (Triticum aestivum L.) area is currently sown with varieties susceptible to the Ug99 race group of stem rust fungus. However, wheat lines Niini, Tinkio, Coni, Pfunye, Blouk, and Ripper have demonstrated Ug99 resistance at the seedling and adult plant stages. We mapped stem rust resistance in populations derived from crosses of a susceptible parent with each of the resistant lines. The segregation of resistance in each population indicated the presence of a single gene. The resistance gene in Niini mapped to short arm of chromosome 6D and was flanked by SSR markers Xcfd49 at distances of 3.9 cM proximal and Xbarc183 8.4 cM distal, respectively. The chromosome location of this resistance was validated in three other populations: PBW343/Coni, PBW343/Tinkio, and Cacuke/Pfunye. Resistance initially postulated to be conferred by the SrTmp gene in Blouk and Ripper was also linked to Xcfd49 and Xbarc183 on 6DS, but it was mapped proximal to Xbarc183 at a similar position to previously mapped genes Sr42 and SrCad. Based on the variation in diagnostic marker alleles, it is possible that Niini and Pfunye may carry different resistance genes/alleles. Further studies are needed to determine the allelic relationships between various genes located on chromosome arm 6DS. Our results provide valuable molecular marker and genetic information for developing Ug99 resistant wheat varieties in diverse germplasm and using these markers to tag the resistance genes in wheat breeding.
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