Artículos (Instituto de Bioquímica Vegetal y Fotosíntesis IBVF – CIC Cartuja)

URI permanente para esta colecciónhttps://hdl.handle.net/11441/10949

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Protein persulfidation: a ubiquitous modification regulating a broad spectrum of biological processes
(Oxford University Press, 2025-10-18) Romero, Luis C.; Carrillo, Reyes; Montesinos-Pereira, David; Luque Algaba, Carmen; Aroca Aguilar, Ángeles; Bioquímica Vegetal y Biología Molecular; Ministerio de Ciencia e Innovación (MICIN). España; European Union (UE); Junta de Andalucía
Hydrogen sulfide signaling occurs mainly through protein persulfidation, an important post-translational modification and a highly dynamic process in plants. Beyond enzyme activity, persulfidation affects protein localization, structure, and interactions. This review provides an overview of the mechanisms governing this modification and the pivotal role of the thioredoxin system mediating depersulfidation, which is essential for maintaining redox homeostasis. Proteomic studies in Arabidopsis thaliana and crop species reveal that protein persulfidation is a widespread posttranslational modification in plants, with over 11 700 polypeptides identified as susceptible. Functional analysis shows a high proportion of these proteins are involved in primary metabolic pathways and secondary metabolism. Gene Ontology analysis highlights additional processes regulated by persulfidation, underscoring its broad regulatory role in plant biology. These processes are mainly related to proteolysis, defense response, embryo development, protein transport, and response to cold. In addition, other processes regulated by hydrogen sulfide, which have been widely studied, also stand out, such as response to abscisic acid, response to oxidative stress, and response to water deprivation. A comprehensive description of the persulfidated proteins implicated in the processes highlighted by the Gene Ontology analysis is provided. This integrated role of H2S with other molecules offers a panoramic overview of its importance in plant biology which also helps to raise new questions and working directions for the future research.
Phenolic acid-induced phase separation and translation inhibition mediate plant interspecific competition
(Nature Portfolio, 2023-08-23) Xie, Zhouli; Zhao, Shuai; Li, Ying; Deng, Yuhua; Shi, Yabo; Chen, Xiaoyuan; Li, Yue; Li, Haiwei; Gutiérrez Beltrán, Emilio; Wang, Wei; Bioquímica Vegetal y Biología Molecular; Natural Science Foundation of China; Beijing Nova Program of Science and Technology; European Commission (EC); Natural Science Foundation of Fujian Province
Phenolic acids (PAs) secreted by donor plants suppress the growth of their susceptible plant neighbours. However, how structurally diverse ensembles of PAs are perceived by plants to mediate interspecific competition remains a mystery. Here we show that a plant stress granule (SG) marker, RNA-BINDING PROTEIN 47B (RBP47B), is a sensor of PAs in Arabidopsis. PAs, including salicylic acid, 4-hydroxybenzoic acid, protocatechuic acid and so on, directly bind RBP47B, promote its phase separation and trigger SG formation accompanied by global translation inhibition. Salicylic acid-induced global translation inhibition depends on RBP47 family members. RBP47s regulate the proteome rather than the absolute quantity of SG. The rbp47 quadruple mutant shows a reduced sensitivity to the inhibitory effect of the PA mixture as well as to that of PA-rich rice when tested in a co-culturing ecosystem. In this Article, we identified the long sought-after PA sensor as RBP47B and illustrated that PA-induced SG-mediated translational inhibition was one of the PA perception mechanisms.
Classification and Nomenclature of Metacaspases and Paracaspases: No More Confusion with Caspases
(Elsevier, 2020-03-05) Minina, Elena A.; Staal, Jens; Alvarez, Vanina E.; Berges, John A.; Berman-Frank, Ilana; Beyaert, Rudi; Bidle, Kay D.; Bornancin, Frédéric; Gutiérrez Beltrán, Emilio; Bozhkov, Peter V.; Bioquímica Vegetal y Biología Molecular
2-Cys peroxiredoxins and the chaperone cpHSP70 act in concert in chloroplast biogenesis in Arabidopsis seedlings
(American Society of Plant Biologists, 2026-03-27) Rodríguez Marín, Fernando; Gallardo Martínez, Antonia María; Hernández Jiménez, María Luisa; González García, María de la Cruz; Cejudo Fernández, Francisco Javier; Pérez Ruiz, Juan Manuel; Bioquímica Vegetal y Biología Molecular; Ministerio de Ciencia e Innovación (MICIN). España; Agencia Estatal de Investigación. España; European Union (UE)
The redox balance between NADPH-dependent thioredoxin reductase C (NTRC) and 2-Cys peroxiredoxins (PRXs) help chloroplast photosynthetic performance acclimate rapidly to environmental cues. The Arabidopsis (Arabidopsis thaliana) 2cpab mutant, which lacks chloroplast 2-Cys PRXs A and B, shows impaired embryogenesis and cotyledon development. This phenotype indicates that these enzymes have a relevant function at early stages of plant development, although this function is poorly understood. Here, we show that the albino cotyledons of the 2cpab mutant have unstructured chloroplasts and decreased chloroplast lipid contents, revealing a key function of 2-Cys PRXs in cotyledon chloroplast differentiation. These phenotypes are mimicked by NTRC overexpression, whereas loss of NTRC function has no effect. RNA-Seq analyses showed transcriptomic changes resembling the response to proteotoxic stress in 2cpab seedlings, which was confirmed by the high sensitivity of 2cpab seedlings to heat stress. In de-etiolating seedlings 2-Cys PRXs interact with chloroplast-localized chaperone heat shock protein 70 (cpHSP70); moreover, Arabidopsis seedlings simultaneously lacking 2-Cys PRXs and cpHSP70-1 exhibit a dramatic alteration of seedling development. Based on these results, we propose that 2-Cys PRXs contribute to cotyledon chloroplast differentiation by affecting organellar proteostasis. This function is exerted in concert with chaperone cpHSP70 and is essential for seedling establishment.
NTRC's novel role: Bridging chloroplast redox oscillations and nuclear circadian clock
(Cell Press, 2025) Cejudo Fernández, Francisco Javier; Bioquímica Vegetal y Biología Molecular; Ministerio de Ciencia e Innovación (MICIN). España; Agencia Estatal de Investigación. España
Gránulos de estrés: posibles dianas biotecnológicas contra el cambio climático
(Universidad de Málaga, 2025) Gutiérrez Beltrán, Emilio; Bioquímica Vegetal y Biología Molecular
Alimentar a 9 mil millones de personas para 2050 es uno de los principales retos que se enfrenta nuestra sociedad. Los cultivos representan la principal fuente de alimento, sin embargo, los eventos atmosféricos ligados al actual cambio climático están afectando de forma muy negativa a la producción agrícola. Si este fenómeno no cambia en los próximos años, la sociedad se enfrentará un grave problema de abastecimiento para una población mundial en crecimiento exponencial. El crecimiento de la temperatura global acompañado de sequias extremas y una alta salinidad de los suelos son las principales causas de perdida de rendimiento de los cultivos. Por lo tanto, desarrollar cultivos resilientes al estrés ambiental es una estrategia prioritaria para mantener un rendimiento óptimo, asegurando la seguridad alimentaria fututa. Para ello, es de máxima importancia comprender cómo las plantas son capaces de enfrentarse a las condiciones de estrés extremas. Uno de los primeros eventos que ocurre tras la percepción de estrés a nivel celular es la formación en el citoplasma de unos gránulos conocidos como gránulos de estrés (SGs, stress granules). Los SGs son condensados biomoleculares constituidos principalmente por ARN y proteínas y que se ensamblan por un proceso de separación de fase líquido-líquido (liquid-liquid phase separation, LLPS). Aunque el conocimiento actual sobre su composición, ensamblaje y función proviene principalmente de organismos modelos como levaduras o animales, estudios recientes en plantas sugieren que los SGs podrían jugar un papel central en la resiliencia vegetal, sugiriendo que el desarrollo de métodos dirigidos sobre los SGs para creación de plantas más resistentes es una posibilidad.
PharaohFUN: phylogenomic analysis for plant protein history and function elucidation
(Oxford University Press, 2026-01-22) Ramos González, Marcos; Ramos González, Víctor; Serrano Pérez, Emma; Arvanitidou, Christina; Hernández García, Jorge; García González, Mercedes; Romero Campero, Francisco José; Ciencias de la Computación e Inteligencia Artificial; Bioquímica Vegetal y Biología Molecular; Ministerio de Ciencia, Innovación y Universidades (MICIU). España; Agencia Estatal de Investigación. España
Since DNA sequencing has become commonplace, the development of efficient methods and tools to explore gene sequences has become indispensable. In particular, despite photosynthetic eukaryotes constituting the largest percentage of terrestrial biomass, computational functional characterization of gene sequences in these organisms still predominantly relies on comparisons with Arabidopsis thaliana and other angiosperms. This paper introduces PharaohFUN, a web application designed for the evolutionary and functional analysis of protein sequences in photosynthetic eukaryotes, leveraging orthology relationships between them. PharaohFUN incorporates a homogeneous representative sampling of key species in this group, bridging clades that have traditionally been studied separately, thus establishing a comprehensive evolutionary framework to draw conclusions about sequence evolution and function. For this purpose, it incorporates modules for exploring gene tree evolutionary history, expansion and contraction events, ancestral states, domain identification, multiple sequence alignments, and diverse functional annotation. It also incorporates different search modes to facilitate its use and increase its reach within the community. Tests were performed on the whole transcription factor toolbox of A. thaliana and on CCA1 protein to assess its utility for both large-scale and fine-grained phylogenetic studies. These exemplify how PharaohFUN accurately traces the corresponding evolutionary histories of these proteins by unifying results for land plants, streptophyte and chlorophyte microalgae. Thus, PharaohFUN democratices access to these kinds of analyses in photosynthetic organisms for every user, independently of their prior training in bioinformatics.
Enhanced sucrose production by controlling carbon flux through CfrA expression in Synechocystis sp. PCC 6803
(Springer Nature, 2025-12-31) Domínguez-Lobo, María Teresa; Ortega Martínez, Pablo; Florencio Bellido, Francisco Javier; Muro-Pastor, M. Isabel; Bioquímica Vegetal y Biología Molecular; Ministerio de Ciencia, Innovación y Universidades (MICIU). España; Junta de Andalucía
Background Cyanobacteria, as phototrophic organisms with low nutritional requirements and great metabolic versatility, are attractive for the sustainable production of value-added chemicals from CO2 and sunlight. One limitation of these strategies is that carbon is partitioned towards biomass synthesis rather than product synthesis. An alternative to conventional metabolic engineering approaches involves controlling regulatory circuits to enhance the flow of carbon towards the synthesis of desired compounds. The carbon-flow-regulator A (CfrA) is pivotal in redirecting carbon flux during nitrogen deficiency in cyanobacteria, promoting glycogen accumulation by inhibiting 2,3-phosphoglycerate mutase enzyme. The moderately halotolerant cyanobacterium Synechocystis sp. PCC 6803 accumulates sucrose and glucosylglycerol (GG) as compatible solutes under salt stress. Sucrose is a valuable carbon source for heterotrophic organisms, whether they are cultivated independently or in co-cultures. In this context, we explored the potential biotechnological relevance of CfrA in redirecting carbon flow towards sucrose production. Results A strain that overexpresses cfrA, independently of nitrogen growth conditions, and carries a plasmid that expresses sucrose-phosphate synthase (SPS) from Synechocystis sp. PCC 6803 and the heterologous sucrose permease CscB inducibly (Pars-cfrA/suc strain) was constructed and analysed. In this strain, cfrA expression increased sucrose production by 40% compared to non-induced levels. The fixed carbon was partially redirected towards sucrose production at the expense of glycogen accumulation and biomass generation. Furthermore, an improvement in the photosynthetic activity of this strain was observed due to the presence of this carbon sink. The effect of eliminating GG synthesis (ΔggpS/Pars-cfrA/suc strain) on sucrose production was also analyzed. Under high salinity conditions (400 mM NaCl), this strain exhibited a maximum sucrose accumulation of 2.72 g/L. Encapsulation of the Pars-cfrA/suc strain has also been studied. Conclusions Our results indicate that modulating carbon flow through CfrA overexpression can substantially boost sucrose production. Glycogen accumulation, mediated by CfrA, enhances sucrose production, which is partly derived from the use of stored glycogen. Furthermore, immobilising Synechocystis cells in alginate improves sucrose production and facilitates its utilisation. Given the widespread occurrence of the cfrA gene in cyanobacteria, its potential as a target in various biotechnological strategies that require the redirection of carbon flow should be considered.
Thioredoxins m are major players in the multifaceted light-adaptive response in Arabidopsis thaliana
(Wiley, 2021-07-20) Serrato, Antonio J.; Rojas González, José A.; Torres Romero, Diego; Vargas, Paola; Mérida, Ángel; Sahrawy, Mariam; Bioquímica Vegetal y Biología Molecular; Ministerio de Economía y Competitividad (MINECO). España; Ministerio de Ciencia e Innovación (MICIN). España; European Union (UE)
Thioredoxins (TRXs) are well-known redox signalling players, which carry out post-translational modiﬁca-tions in target proteins. Chloroplast TRXs are divided into different types and have central roles in lightenergy uptake and the regulation of primary metabolism. The isoforms TRX m1, m2, and m4 from Arabidop-sis thaliana are considered functionally related. Knowing their key position in the hub of plant metabolism,we hypothesized that the impairment of the TRX m signalling would not only have harmful consequenceson chloroplast metabolism but also at different levels of plant development. To uncover the physiologicaland developmental processes that depend on TRX m signalling, we carried out a comprehensive study ofArabidopsis single, double, and triple mutants defective in the TRX m1, m2, and m4 proteins. As light andredox signalling are closely linked, we investigated the response to high light (HL) of the plants that aregradually compromised in TRX m signalling. We provide experimental evidence relating the lack of TRX mand the appearance of novel phenotypic features concerning mesophyll structure, stomata biogenesis, andstomatal conductance. We also report new data indicating that the isoforms of TRX m ﬁne-tune theresponse to HL, including the accumulation of the protective pigment anthocyanin. These results revealnovel signalling functions for the TRX m and underline their importance for plant growth and fulﬁlment ofthe acclimation/response to HL conditions.
The Heterocyst-Specific Small RNA NsiR1 Regulates the Commitment to Differentiation in Nostoc
(American Society for Microbiology, 2022-03-01) Brenes-Álvarez, Manuel; Vioque Peña, Agustín; Muro Pastor, Alicia M.; Bioquímica Vegetal y Biología Molecular; Ministerio de Ciencia e Innovación (MICIN). España; Agencia Estatal de Investigación. España; European Union (UE)
Heterocysts are specialized cells that ﬁlamentous cyanobacteria differentiatefor the ﬁxation of atmospheric nitrogen when other nitrogen sources are not available.Heterocyst differentiation at semiregular intervals along the ﬁlaments requires complexstructural and metabolic changes that are under the control of the master transcriptionalregulator HetR. NsiR1 (nitrogen stress-induced RNA 1) is a HetR-dependent noncoding RNAthat is expressed from multiple chromosomal copies, some identical, some slightly divergentin sequence, speciﬁcally in heterocysts from very early stages of differentiation. We havepreviously shown that NsiR1 inhibits translation of the overlapping hetF mRNA by anantisense mechanism. Here, we identify alr3234, a hetP-like gene involved in the regula-tion of commitment (point of no return) to heterocyst differentiation, as a target of NsiR1.A strain overexpressing one of the identical copies of NsiR1 commits to heterocyst devel-opment earlier than the wild type. The posttranscriptional regulation exerted by NsiR1 onthe expression of two genes involved in heterocyst differentiation and commitment, hetFand alr3234, adds a new level of complexity to the network of transcriptional regulationand protein-protein interactions that participate in heterocyst differentiation. IMPORTANCE Heterocysts are nitrogen-ﬁxing specialized cells that appear at semiregularintervals along cyanobacterial ﬁlaments upon nitrogen starvation. The differentiation andpatterning of heterocysts is a model for the study of cell differentiation in multicellularprokaryotes. The regulation of differentiation, which is only partially understood, includestranscriptional changes, factor diffusion between cells, and protein-protein interactions. Thiswork describes the identiﬁcation of a novel target for NsiR1, a small RNA (sRNA) encodedin multiple slightly divergent copies, and shows how different copies of “sibling” sRNAs reg-ulate the expression of different targets involved in one of the few examples of a differen-tiation process in prokaryotes.
Photosynthetic assimilation of CO2 regulates TOR activity
(National Academy of Sciences, 2022-01-07) Mallén Ponce, Manuel J.; Pérez Pérez, María Esther; Crespo, José L.; Bioquímica Vegetal y Biología Molecular; Ministerio de Ciencia y Tecnología (MCYT). España
The target of rapamycin (TOR) kinase is a master regulator that integrates nutrient signals to promote cell growth in all eukaryotes. It is well established that amino acids and glucose are major regulators of TOR signaling in yeast and metazoan, but whether and how TOR responds to carbon availability in photosynthetic organisms is less understood. In this study, we showed that photosynthetic assimilation of CO2 by the Calvin–Benson–Bassham (CBB) cycle regulates TOR activity in the model single-celled microalga Chlamydomonas reinhardtii. Stimulation of CO2 fixation boosted TOR activity, whereas inhibition of the CBB cycle and photosynthesis down-regulated TOR. We uncovered a tight link between TOR activity and the endogenous level of a set of amino acids including Ala, Glu, Gln, Leu, and Val through the modulation of CO2 fixation and the use of amino acid synthesis inhibitors. Moreover, the finding that the Chlamydomonas starch-deficient mutant sta6 displayed disproportionate TOR activity and high levels of most amino acids, particularly Gln, further connected carbon assimilation and amino acids to TOR signaling. Thus, our results showed that CO2 fixation regulates TOR signaling, likely through the synthesis of key amino acids.
Soil Bacteriome Shifts along a Cultivation Gradient in Southwestern Spanish Wetlands
(Springer, 2025-11-29) González Pimentel, José Luis; Cuecas Morano, María de Piedras Alba; Álvarez Núñez, Consolación; Bioquímica Vegetal y Biología Molecular
Understanding how long-term agricultural practices affect soil bacteriome is essential for sustainable land management. In the Guadalquivir Marshes of southwestern Spain, which encompass both Doñana National Park and one of Europe’s most productive rice cultivation areas, decades of rice farming have transformed natural wetlands into artificial agroecosystems. Although bacterial degradation in cultivated soils has been previously suggested, comparative analyses between rice paddies and adjacent natural wetlands remain scarce. Here, we characterized the soil bacteriome across a cultivation gradient by comparing undisturbed natural marshes, within Doñana National Park, with rice fields cultivated for 25 years (Cantarita) and 80 years (Mínima 2). Using full 16S rRNA gene via long-read metabarcoding and standardized soil physicochemical assays, we analysed taxonomic composition, environmental associations, and predicted functional profiles. Our results reveal a progressive restructuring of bacterial communities with increased cultivation time, notably a significant enrichment of Chloroflexota (especially Anaerolineae) and a decline in Actinomycetota and Planctomycetota in paddy soils. Functional predictions indicated a higher potential for denitrification in cultivated soils—likely involving Chloroflexota taxa—compared to more diverse nitrogen pathways in natural sites. These shifts were strongly associated with changes in pH, electrical conductivity, calcium carbonate, and nitrate levels. Remarkably, most bacterial differences were already evident within the first 25 years of cultivation, underscoring the rapid ecological impact of intensive rice cultivation. Notably, we identified specific bacterial groups (Anaerolineae and Nocardioides in paddy soils; Euzebya, Rubrobacter, and Planctomycetota in natural wetlands), whose enrichment was associated with soil type. This approach highlights the value of integrating bacterial-based assessments into sustainable wetland management strategies.
Hydrogen sulfide improves performance under suppressed photorespiration in Arabidopsis thaliana and orchestrates molecular reprogramming to alleviate stress
(Elsevier, 2026-01-08) Luque, C.; García Calderón, Margarita; Gotor, C.; Márquez Cabeza, Antonio José; Aroca Aguilar, Ángeles; Bioquímica Vegetal y Biología Molecular; European Union (UE); Ministerio de Ciencia e Innovación (MICIN). España; Junta de Andalucía
High levels of atmospheric carbon dioxide result in suppression of plant photorespiration. The non-photorespiratory conditions (NPC) result in unbalancing the C/N metabolism, overproducing reactive oxygen species (ROS), and reducing stomatal activity. In plant stress responses, hydrogen sulfide (H2S) has been identified as an important signaling molecule through persulfidation of specific proteins. Previous works demonstrated that H₂S protects Arabidopsis thaliana against NPC-induced stress, and this work investigates the molecular basis of such protection. H₂S modulates a metabolic reprogramming influencing elemental homeostasis of C/N ratio, amino acids profile, central carbon metabolites and accumulation of polyunsaturated fatty acids (PUFAs). Persulfidation level under NPC was also restored after H₂S treatment. At the transcriptomic level, several well-known hypoxia marker genes, such as plant CYSTEINE OXIDASE 1 and 2, ETHYLENE-RESPONSIVE TRANSCRIPTION FACTOR ERF71 AND ETHYLENE RECEPTOR 2, are induced under NPC, and sulfide treatment decreases their expression levels to the ones in active photorespiration conditions (APC). H₂S also negatively regulates ABA signaling by targeting genes controlling ion transport and stomatal development which are involved in stomatal function. These integrated responses across metabolism, redox regulation and developmental programming emphasize the key contribution of H₂S to orchestrating plant adaptation to high CO₂ environments, positioning it as a master regulator that ensures plant resilience in the face of climate change.
Dihydroxyacetone phosphate generated in the chloroplast mediates the activation of TOR by CO2 and light
(American Association for the Advancement of Science, 2025-04-18) Mallén Ponce, Manuel J.; Quintero Moreno, Andrea M.; Gámez Arcas, Samuel; Grossman, Arthur R.; Pérez Pérez, María Esther; Crespo, José L.; Bioquímica Vegetal y Biología Molecular; Ministerio de Ciencia, Innovación y Universidades (MICIU). España
Light and CO2 assimilation activate the target of rapamycin (TOR) kinase in photosynthetic cells, but how thesesignals are transmitted to TOR is unknown. Using the green alga Chlamydomonas reinhardtii as a model system,we identified dihydroxyacetone phosphate (DHAP) as the key metabolite regulating TOR in response to carbonand light cues. Metabolomic analyses of synchronized cells revealed that DHAP levels change more than any oth-er metabolite between dark- and light-grown cells and that the addition of the DHAP precursor, dihydroxyacetone(DHA), was sufficient to activate TOR in the dark. We also demonstrated that TOR was insensitive to light or inor-ganic carbon but not to exogenous DHA in a Chlamydomonas mutant defective in the export of DHAP from thechloroplast. Our results provide a metabolic basis for the mode of TOR control by light and inorganic carbon andindicate that cytoplasmic DHAP is an important metabolic regulator of TOR.
Redox partner interactions in the ATG8 lipidation system in microalgae
(Elsevier, 2023-04-06) Mallén Ponce, Manuel J.; Gámez Arcas, Samuel; Pérez Pérez, María Esther; Bioquímica Vegetal y Biología Molecular; Ministerio de Ciencia e Innovación (MICIN). España
Autophagy is a catabolic pathway that functions as a degradative and recycling process to maintain cellular homeostasis in most eukaryotic cells, including photosynthetic organisms such as microalgae. This process involves the formation of double-membrane vesicles called autophagosomes, which engulf the material to be degraded and recycled in lytic compartments. Autophagy is mediated by a set of highly conserved autophagyrelated (ATG) proteins that play a fundamental role in the formation of the autophagosome. The ATG8 ubiquitin-like system catalyzes the conjugation of ATG8 to the lipid phosphatidylethanolamine, an essential reaction in the autophagy process. Several studies identified the ATG8 system and other core ATG proteins in photosynthetic eukaryotes. However, how ATG8 lipidation is driven and regulated in these organisms is not fully understood yet. A detailed analysis of representative genomes from the entire microalgal lineage revealed a high conservation of ATG proteins in these organisms with the remarkable exception of red algae, which likely lost ATG genes before diversification. Here, we examine in silico the mechanisms and dynamic interactions between different components of the ATG8 lipidation system in plants and algae. Moreover, we also discuss the role of redox post-translational modifications in the regulation of ATG proteins and the activation of autophagy in these organisms by reactive oxygen species.
Redox-mediated activation of ATG3 promotes ATG8 lipidation and autophagy progression in Chlamydomonas reinhardtii
(Oxford University Press, 2023-09-29) Mallén Ponce, Manuel J.; Pérez Pérez, María Esther; Bioquímica Vegetal y Biología Molecular; Ministerio de Economía y Competitividad (MINECO). España; Ministerio de Ciencia e Innovación (MICIN). España
Autophagy is one of the main degradative pathways used by eukaryotic organisms to eliminate useless or damaged intracellular material to maintain cellular homeostasis under stress conditions. Mounting evidence indicates a strong interplay between the generation of reactive oxygen species and the activation of autophagy. Although a tight redox regulation of autophagy has been shown in several organisms, including microalgae, the molecular mechanisms underlying this control remain poorly understood. In this study, we have performed an in-depth in vitro and in vivo redox characterization of ATG3, an E2-activating enzyme involved in ATG8 lipidation and autophagosome formation, from 2 evolutionary distant unicellular model organisms: the green microalga Chlamydomonas (Chlamydomonas reinhardtii) and the budding yeast Saccharomyces cerevisiae. Our results indicated that ATG3 activity from both organisms is subjected to redox regulation since these proteins require reducing equivalents to transfer ATG8 to the phospholipid phosphatidylethanolamine. We established the catalytic Cys of ATG3 as a redox target in algal and yeast proteins and showed that the oxidoreductase thioredoxin efficiently reduces ATG3. Moreover, in vivo studies revealed that the redox state of ATG3 from Chlamydomonas undergoes profound changes under autophagyactivating stress conditions, such as the absence of photoprotective carotenoids, the inhibition of fatty acid synthesis, or high light irradiance. Thus, our results indicate that the redox-mediated activation of ATG3 regulates ATG8 lipidation under oxidative stress conditions in this model microalga.
Lipid turnover through lipophagy in the newly identiﬁedextremophilic green microalga Chlamydomonas urium
(Wiley, 2024-07-05) Pérez Pérez, María Esther; Mallén Ponce, Manuel J.; Odriozola Gil, Yosu; Rubio, Alejandro; Salas, Joaquín J.; Martínez Force, Enrique; Pérez Pulido, Antonio J.; Crespo, José Luis; Bioquímica Vegetal y Biología Molecular; Ministerio de Ciencia, Innovación y Universidades (MICIU). España; Junta de Andalucía; Consejo Superior de Investigaciones Científicas (CSIC)
Autophagy is a central degradative pathway highly conserved among eukaryotes, includingmicroalgae, which remains unexplored in extremophilic organisms. In this study, we describedand characterized autophagy in the newly identiﬁed extremophilic green microalga Chlamy-domonas urium, which was isolated from an acidic environment. The nuclear genome of C. urium was sequenced, assembled and annotated in order toidentify autophagy-related genes. Transmission electron microscopy, immunoblotting, meta-bolomic and photosynthetic analyses were performed to investigate autophagy in this extre-mophilic microalga. The analysis of the C. urium genome revealed the conservation of core autophagy-relatedgenes. We investigated the role of autophagy in C. urium by blocking autophagic ﬂux withthe vacuolar ATPase inhibitor concanamycin A. Our results indicated that inhibition of autop-hagic ﬂux in this microalga resulted in a pronounced accumulation of triacylglycerols and lipiddroplets (LDs). Metabolomic and photosynthetic analyses indicated that C. urium cells withimpaired vacuolar function maintained an active metabolism. Such effects were not observedin the neutrophilic microalga Chlamydomonas reinhardtii. Inhibition of autophagic ﬂux in C. urium uncovered an active recycling of LDs through lipo-phagy, a selective autophagy pathway for lipid turnover. This study provided the metabolicbasis by which extremophilic algae are able to catabolize lipids in the vacuole.
Retention of a SulP-family bicarbonate transporter in a periplasmic N2-fixing cyanobacterial endosymbiont of an open ocean diatom
(Oxford University Press, 2025-09-04) Nieves Morión, Mercedes; Romero García, Rubén; Bardi, Sepehr; López Maury, Luis; Hagemann, Martin; Flores, Enrique; Foster, Rachel A.; Bioquímica Vegetal y Biología Molecular; Swedish Research Council; Junta de Andalucía; Knut and Alice Wallenberg Foundation
Symbioses between diatoms and the N2-fixing, heterocyst-forming cyanobacteria Richelia spp. are widespread and contribute to primary production. Unique to these symbioses is a variation in the symbiont location: one lives in the host cytoplasm (endobiont) vs. residing between the host frustule and plasmalemma (periplasmic endobiont). Both partners are photosynthetic, yet how the partners acquire, share, or compete for bicarbonate necessary for their photosynthesis is unknown. The genomes of both endobionts (ReuHH01 and RintRC01, respectively) contain genes encoding SulP-family proteins, which are oxyanion transporters. To study the possible involvement of these transporters in bicarbonate uptake, we used complementation in a Synechocystis sp. PCC 6803 mutant that is unable to grow at air levels of CO2 because all five of its inorganic carbon uptake systems have been inactivated. Of the five genes tested, only one (RintRC_3892) from the periplasmic endobiont complemented the mutant to grow with air levels of CO2 or at low bicarbonate concentrations. The complemented strain showed strong sodium-dependent and low-affinity bicarbonate uptake that was consistent with bicarbonate concentrations expected in the diatom periplasm. Additionally, all the amino acids involved in the bicarbonate binding site of BicA from Synechocystis sp. PCC 6803 are conserved in RintRC_3892. Finally, the importance of the RintRC_3892 protein was confirmed by the consistent detection of its transcripts in wild Richelia populations from three different oceans. Combined our results showed no evidence for a bicarbonate transporter in the cytoplasmic endobiont, whereas the periplasmic endobiont has retained a SulP-type bicarbonate transporter for its own photosynthesis.
Two opposing redox signals mediated by 2-cys peroxiredoxin shape the redox proteome during photosynthetic induction
(Elsevier, 2025-08-05) Doron, Shani; Lampl, Nardy; Savidor, Alon; Pri-Or, Amir; Katina, Corine; Cejudo Fernández, Francisco Javier; Levin, Yishai; Rosenwasser, Shilo; Bioquímica Vegetal y Biología Molecular; European Research Council (ERC); Israel Science Foundation
Photosynthetic induction, characterized by the lag in CO2 assimilation rates during transition from darkness to light, has traditionally been attributed to Rubisco activase activity and stomatal opening. Yet, the faster induction of photosynthesis in the 2-Cys peroxiredoxins (Prxs) mutant (2cpab) suggested a role for oxidative signals in regulating photosynthetic rates, although the underlying molecular mechanism remains unclear. SPEAR, a redox proteomics approach, was used to systematically map redox changes occurring during photosynthesis induction and to unravel the role of 2-Cys Prxs in shaping these redox alterations. No significant difference was observed in protein expression levels between WT and 2cpab plants, suggesting that protein abundance does not account for the 2cpab phenotype. During the transition from dark to low light, 82 and 54 cysteine-containing peptides were reduced or oxidized, respectively, in WT plants. Most redox-regulated cysteines in photosynthetic proteins were found oxidized in the dark and became reduced in response to light. A reverse pattern was observed among redox-regulated cysteines in proteins involved in starch degradation and chloroplast glycolysis, which shifted from a reduced to an oxidized state in response to light. These findings demonstrate the initiation of two opposing redox responses, affecting distinct sets of metabolic proteins during the induction phase. Remarkably, a significantly lower number of cysteines were reduced or oxidized in 2cpab plants, highlighting the crucial role 2-Cys Prxs play in shaping both signals. Taken together, rotational shifts between metabolic pathways during the photosynthesis induction phase are regulated by two opposing redox signals mediated by 2-Cys Prx activity.
Redox regulation of membrane-associated processes mediated by chloroplastic thioredoxins
(Elsevier, 2026-02) Vargas, Paola; Torres Romero, Diego; Mérida, Ángel; Sahrawy, Mariam; Serrato, Antonio J.; Bioquímica Vegetal y Biología Molecular; Ministerio de Ciencia, Innovación y Universidades (MICIU). España; Ministerio de Ciencia e Innovación (MICIN). España
Plant chloroplasts are complex organelles that house a plethora of redox-controlled metabolic processes. However, our understanding of membrane-level redox signalling remains limited. In order to expand our knowledge of redox regulation in these photosynthetic subcellular compartments, we carried out in vitro and in vivo experimental approaches focused on the analysis of processes taking place at the membrane level. In addition to the classic stromal localization, these approaches have revealed that chloroplastic thioredoxins (TRXs) from Arabidopsis thaliana are also membrane-associated proteins, having a network of non-stromal interactors. We have identified 185 putative chloroplastic targets, with 80 % predicted to be located at the envelope or thylakoid membranes, and classified into 18 functional categories, with the most prevalent one being related to photosynthesis and, notably, metabolite/ion transport, a novel finding in redox regulation. Direct in vivo interactions between TRXs m and three integral proteins involved in protein import and metabolite transport, as well as one thylakoid membrane-bound protein that regulates proteolytic processes, were confirmed. Moreover, the role of TRXs m appears to extend beyond the regulation of the primary process of photosynthesis, such as during the establishment of greening cotyledons and the protection against high-light intensities. These findings provide a novel perspective on the function of TRXs as multifaceted regulators. The present study aims to address a current knowledge gap by exploring redox signalling in the membranes of plant chloroplasts.

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