Biology subjects

Abreu, I.

Publications and source records attributed to Abreu, I..

11 recordsLinked to original sources

bioRxiv2026

Enhanced production of nitrogenase components in Nicotiana benthamiana through co-expression with Bacterioferritin A

O_LIEngineering nitrogen fixing crops requires not only transferring the nitrogenase structural genes, but also the accessory genes to synthesize its iron-sulphur cofactors. Scaffold protein NifU is a critical element in this system as the starting point of nitrogenase cofactor assembly. NifU has been successfully produced in plants, however, its optimal production required high levels of iron in the medium. This is likely due to a faulty connection with the endogenous iron trafficking network C_LIO_LITo identify specific elements targeting iron to NifU, pull-down assays were performed to identify showing bacterioferritin A (BfrA) as a likely candidate. Co-immunopurification, mutant characterization, iron transfer assays, and co-expression in Nicotiana benthamiana assays were carried out. C_LIO_LIBfrA transfers iron to NifU through protein-protein interactions. When these two proteins were co-expressed in N. benthamiana leaves, there was an increase in NifU production. In turn, it led to doubling NifH synthesis, a nitrogenase structural protein that is also required for the synthesis of the more complex nitrogenase cofactors. C_LIO_LIOur results provide a new element towards engineering nitrogen-fixing crops. They also underscore the importance of transferring the metal delivery systems when expressing metalloproteins in heterologous systems. C_LI

Plant Biology↗

bioRxiv2026

Chemodiverse cell systems responses to UV in an algal sister of land plants

Plant terrestrialization necessitated that a barrage of stressors had to be overcome1. Land plants use an integrated response network in adjusting their molecular physiology to terrestrial stressors2--one of the foremost being UV irradiance. The zygnematophytes are the closest streptophyte algal relatives of land plants3-5, are renowned for their resilience to UV stress6-8, and thus allow to glean key information for inferring the UV response toolkit of the earliest land plants9,10. Throughout streptophyte evolution, specialised metabolism radiated into creating diverse compounds used for responses to environmental challenges, such as sun-shielding compounds and antioxidants11-14. This includes UV-shielding compounds like flavonoids and coumarins but also the land plant specific polymer lignin, giving structural support in vascular plants15; homologs of the underpinning core pathway occur in streptophyte algae16. Here, we exposed the zygnematophyte Mesotaenium to UV-B irradiation and profiled its physiology, morphology, transcriptomics as well as metabolomic features. After UV-B exposure, cells showed rapid photophysiological responses and progressively growing terminal vacuoles. Our transcriptome data capture dynamic changes in gene expression of (i) core downstream responses such as genes homologous to phenol metabolic enzymes, photophysiological homeostats, and DNA repair factors; but also (ii) upstream components featuring key homologs of kinase-mediated signalling cascades, as well as light quality and abscisic acid-mediated signalling components. To scrutinize the acclimatory chassis, we created a metabolite feature database specifically for the Mesotaenium metabolome. Upon UV-B exposure, the metabolome displayed pronounced temporal shifts, with several phenolic features that accumulate along the stress-acclimation kinetics. Overall, we capture a chemodiverse response including various phenolics such as purpurogallin-like, methoxypsoralen-like derivatives and coumarins. Our data establish an integrated model for UV responses in the closest algal relatives of land plants, shedding light on the toolkit that allowed the progenitors of land plants to move out of a protective water column.

evolutionary biology↗

bioRxiv2025

KDM7-mediated oxygen sensing reprograms chromatin to enhance hypoxia tolerance in the root

Roots frequently encounter low oxygen (hypoxia) from soil compaction or water saturation and must adapt to this stress. We investigated how root tip cells sense hypoxia and adjust the meristem epigenome to activate genes that promote tolerance and growth under oxygen limitation. In Arabidopsis root tips, hypoxia tolerance was linked to increased trimethylation of histone H3 at lysine 4 (H3K4me3). We also found that group 7 demethylases (KDM7s) are directly inhibited by hypoxia, and that genetic inactivation of KDM7s, like hypoxia, induces expression of genes essential for meristem survival under oxygen deprivation. We propose that KDM7s function as root-specific oxygen sensors that prime and support hypoxia tolerance.

plant biology↗

bioRxiv2022

Functional characterization of the Co2+ transporter AitP in Sinorhizobium meliloti: a new player in Fe2+ homeostasis

Co2+ induces the increase of the labile-Fe pool (LIP) by Fe-S cluster damage, heme synthesis inhibition and "free" iron import, which affects cell viability. The N2-fixing bacteria, Sinorhizobium meliloti, is a suitable model to determine the roles of Co2+-transporting Cation diffusion facilitator exporters (Co-eCDF) in Fe2+ homeostasis because it has a putative member of this sub-family, AitP, and two specific Fe2+-export systems. An insertional mutant of AitP showed Co2+ sensitivity and accumulation, Fe accumulation and hydrogen peroxide sensitivity, but not Fe2+ sensitivity, despite AitP being a bona fide low affinity Fe2+ exporter as demonstrated by the kinetic analyses of Fe2+ uptake into everted membrane vesicles. Suggesting concomitant Fe2+-dependent induced stress, Co2+ sensitivity was increased in strains carrying mutations in AitP and Fe2+ exporters which did not correlate with the Co2+ accumulation. Growth in the presence of sub-lethal Fe2+ and Co2+ concentrations suggested that free Fe-import might contribute to Co2+ toxicity. Supporting this, Co2+ induced transcription of Fe-import system and genes associated with Fe homeostasis. Analyses of total protoporphyrin content indicates Fe-S cluster attack as the major source for LIP. AitP-mediated Fe2+-export is likely counterbalanced via a non-futile Fe2+-import pathway. Two lines of evidence support this: i) an increased hemin uptake in presence of Co2+ was observed in WT vs. AitP mutant, and ii) hemin reversed the Co2+ sensitivity in the AitP mutant. Thus, the simultaneous detoxification mediated by AitP aids cells to orchestrate an Fe-S cluster salvage response, avoiding the increase in the LIP caused by the disassembly of Fe-S clusters or free iron uptake. ImportanceCross-talk between iron and cobalt has been long recognized in biological systems. This is due to the capacity of cobalt to interfere with proper iron utilization. Cells can detoxify cobalt by exporting mechanisms involving membrane proteins known as exporters. Highlighting the cross-talk, the capacity of several cobalt exporters to also export iron is emerging. Although biologically less important than Fe2+, Co2+ induces toxicity by promoting intracellular Fe release, which ultimately causes additional toxic effects. In this work, we describe how the N2-fixating rhizobial cells solve this perturbation by clearing Fe through a Co2+-exporter, in order to reestablish intracellular Fe-levels by importing non-free Fe, heme. This piggyback-ride type of transport may aid bacterial cells to survive in free-living conditions where high anthropogenic Co2+ content may be encountered.

microbiology↗

bioRxiv2020

Arabidopsis thaliana Zn2+-efflux ATPases HMA2 and HMA4 are required for resistance to the necrotrophic fungus Plectosphaerella cucumerina BMM

O_LIZinc is an essential nutrient at low concentrations, but toxic at slightly higher ones. This could be used by plants to fight pathogens colonization. C_LIO_LIElemental distribution in Arabidopsis thaliana leaves inoculated with the necrotrophic fungus Plectosphaerella cucumerina BMM (PcBMM) was determined and compared to mock-inoculated ones. Infection assays were carried out in wild type and long-distance zinc trafficking double mutant hma2hma4, defective in root-to-shoot zinc partitioning. Expression levels of genes involved in zinc homeostasis or in defence phytohormone-mediated pathways were determined. C_LIO_LIZinc and manganese levels increased at the infection site. Zinc accumulation was absent in hma2hma4. HMA2 and HMA4 transcription levels were upregulated upon PcBMM inoculation. Consistent with a role of these genes in plant immunity, hma2hma4 mutants were more susceptible to PcBMM infection, phenotype rescued upon zinc supplementation. Transcript levels of genes involved in the salicylic acid, ethylene and jasmonate pathways were constitutively upregulated in hma2hma4 plants. C_LIO_LIThese data are consistent with a role of zinc in plant immunity not only of hyperaccumulator plants, but also of plants containing ordinary levels of zinc. This new layer of immunity seems to run in parallel to the already characterized defence pathways, and its removal has a direct effect on pathogen resistance. C_LI

plant biology↗

bioRxiv2019

Medicago truncatula Yellow Stripe1-Like3 gene is involved in symbiotic nitrogen fixation

Symbiotic nitrogen fixation carried out in legume root nodules requires transition metals. These nutrients are delivered by the host plant to the endosymbiotic nitrogen-fixing bacteria living with the nodule cells, a process in which vascular transport is essential. As occurs in root-to-shoot transport, members of the Yellow Stripe-Like (YSL) family of metal transporters should also be required for root-to-nodule metal delivery. The genome of the model legume Medicago truncatula encodes for eight YSL proteins, four of them with a high degree of similarity to Arabidopsis thaliana YSLs involved in long-distance metal trafficking. Among them, MtYSL3 is a plasma membrane protein expressed by vascular cells in roots and nodules, and by cortical nodule cells. Reducing expression levels of this gene had no major effect on plant physiology when assimilable nitrogen was provided in the nutrient solution. However, nodule functioning was severely impaired, with a significant reduction of nitrogen fixation capabilities. Further, iron and zinc accumulation and distribution changed. Iron was retained in the apical region of the nodule, while zinc became strongly accumulated in the nodule veins in the ysl3 mutant. These data suggest a role of MtYSL3 in vascular delivery of iron and zinc to symbiotic nitrogen fixation. HighlightMedicago truncatula YSL3 transporter is required for optimal nitrogen fixation in root nodules, mediating iron and zinc distribution in these organs.

plant biology↗

bioRxiv2019

Nicotianamine synthase 2 is required for symbiotic nitrogen fixation in Medicago truncatula nodules

Symbiotic nitrogen fixation carried out by the interaction between legumes and diazotrophic bacteria known as rhizobia requires of relatively large levels of transition metals. These elements act as cofactors of many key enzymes involved in this process. Metallic micronutrients are obtained from soil by the roots and directed to sink organs by the vasculature, in a process participated by a number of metal transporters and small organic molecules that mediate metal delivery in the plant fluids. Among the later, nicotianamine is one of the most important. Synthesized by nicotianamine synthases (NAS), this non-proteinogenic amino acid forms metal complexes participating in intracellular metal homeostasis and long-distance metal trafficking. Here we characterized the NAS2 gene from model legume Medicago truncatula. MtNAS2 is located in the root vasculature and in all nodule tissues in the infection and fixation zones. Symbiotic nitrogen fixation requires of MtNAS2 function, as indicated by the loss of nitrogenase activity in the insertional mutant nas2-1, a phenotype reverted by reintroduction of a wild-type copy of MtNAS2. This would be the result of the altered iron distribution in nas2-1 nodules, as indicated by X-ray fluorescence studies. Moreover, iron speciation is also affected in these nodules. These data suggest a role of nicotianamine in iron delivery for symbiotic nitrogen fixation.\n\nSignificance StatementNicotianamine synthesis mediated by MtNAS2 is important for iron allocation for symbiotic nitrogen fixation by rhizobia in Medicago truncatula root nodules.

plant biology↗

bioRxiv2019

Medicago truncatula Ferroportin2 mediates iron import into nodule symbiosomes

Iron is an essential cofactor for symbiotic nitrogen fixation. It is required by many of the enzymes facilitating the conversion of N2 into NH4+ by endosymbiotic bacteria living within root nodule cells, including signal transduction proteins, O2 homeostasis systems, and nitrogenase itself. Consequently, host plants have developed a transport network to deliver essential iron to nitrogen-fixing nodule cells. Model legume Medicago truncatula Ferroportin2 (MtFPN2) is a nodule-specific gene that encodes an iron-efflux protein. MtFPN2 is located in intracellular membranes in the nodule vasculature, and in the symbiosome membranes that contain the nitrogen-fixing bacteria in the differentiation and early-fixation zones of the nodules. Loss-of-function of MtFPN2 leads to altered iron distribution and speciation in nodules, which causes a reduction in nitrogenase activity and in biomass production. Using promoters with different tissular activity to drive MtFPN2 expression in MtFPN2 mutants, we determined that MtFPN2-facilitated iron delivery across symbiosomes is essential for symbiotic nitrogen fixation, while its presence in the vasculature does not seem to play a major role in in the conditions tested.

plant biology↗

bioRxiv2018

SSU72 phosphatase is a telomere replication terminator

Telomeres, the protective ends of eukaryotic chromosomes, are replicated through concerted actions by conventional DNA polymerases and telomerase, though the regulation of this process is not fully understood. Telomere replication requires (C)-Stn1-Ten1, a telomere ssDNA-binding complex that is homologous to RPA. Here, we show that the evolutionarily conserved phosphatase Ssu72 is responsible for terminating the cycle of telomere replication in fission yeast. Ssu72 controls the recruitment of Stn1 to telomeres by regulating Stn1 phosphorylation at S74, a residue that lies within the conserved OB fold domain. Consequently, ssu72{Delta} mutants are defective in telomere replication and exhibit long 3 overhangs, which are indicative of defective lagging strand DNA synthesis. We also show that hSSU72 regulates telomerase activation in human cells by controlling the recruitment of hSTN1 to telomeres. Thus, in this study, we demonstrate a previously unknown yet conserved role for the phosphatase SSU72, whereby this enzyme controls telomere homeostasis by activating lagging strand DNA synthesis, thus terminating the cycle of telomere replication.

cell biology↗

bioRxiv2017

Medicago truncatula copper transporter1 (MtCOPT1) delivers copper for symbiotic nitrogen fixation

* Copper is an essential nutrient for symbiotic nitrogen fixation. This element is delivered by the host plant to the nodule, where membrane copper transporter would introduce it into the cell to synthesize cupro-proteins.\n\n* COPT family members in model legume Medicago truncatula were identified and their expression determined. Yeast complementation assays, confocal microscopy, and phenotypical characterization of a Tnt1 insertional mutant line were carried out in the nodule-specific M.truncatula COPT family member.\n\n* Medicago truncatula genome encodes eight COPT transporters. MtCOPT1 (Medtr4g019870) is the only nodule-specific COPT gene. It is located in the plasma membrane of the differentiation, interzone and early fixation zones. Loss of MtCOPT1 function results in a copper-mitigated reduction of biomass production when the plant obtains its nitrogen exclusively from symbiotic nitrogen fixation. Mutation of MtCOPT1 results in diminished nitrogenase activity in nodules, likely an indirect effect from the loss of a copper-dependent function, such as cytochrome oxidase activity in copt1-1 bacteroids.\n\n* These data are consistent with a model in which MtCOPT1 transports copper from the apoplast into nodule cells to provide copper for essential metabolic processes associated with symbiotic nitrogen fixation.

plant biology↗

bioRxiv2017

Medicago truncatula Zinc-Iron Permease6 provides zinc to rhizobia-infected nodule cells

AbstractZinc is a micronutrient required for symbiotic nitrogen fixation. It has been proposed that in model legume Medicago truncatula, zinc is delivered in a similar fashion as iron, i.e. by the root vasculature into the nodule and released in the infection/differentiation zone. There, zinc transporters must introduce this element into rhizobia-infected cells to metallate the apoproteins that use zinc as a cofactor. MtZIP6 (Medtr4g083570) is a M. truncatula Zinc-Iron Permease (ZIP) that is expressed only in roots and nodules, with the highest expression levels in the infection/differentiation zone. Immunolocalization studies indicate that it is located in the plasma membrane of rhizobia-infected cells in the nodule. Down-regulating MtZIP6 expression levels with RNAi does not result in any strong phenotype when plants are being watered with mineral nitrogen. However, these silenced plants displayed severe growth defects when they depended on nitrogen fixed by their nodules, as a consequence of the loss of 80% of their nitrogenase activity. The reduction of this activity was not the result of iron not reaching the nodule, but an indirect effect of zinc being retained in the infection/differentiation zone and not reaching the cytosol of rhizobia-infected cells. These data are consistent with a model in which MtZIP6 would be responsible for zinc uptake by rhizobia-infected nodule cells in the infection/differentiation zone.

plant biology↗