Artículos (Bioquímica Vegetal y Biología Molecular)

URI permanente para esta colecciónhttps://hdl.handle.net/11441/10824

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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.
APE1 is involved in the stabilization/formation of the RC47 complex of PSII in oxygenic prototrophic organisms
(Oxford University Press, 2025-12-17) Torres Romero, Diego; Muro-Pastor, María Isabel; Arana, Francisco José; Hornero-Méndez, Dámaso; Serrato, Antonio Jesús; González García, María de la Cruz; Sahrawy, Mariam; Florencio Bellido, Francisco Javier; Mérida, Ángel; Bioquímica Vegetal y Biología Molecular; Ministerio de Ciencia, Innovación y Universidades (MICIU). España; Agencia Estatal de Investigación. España
Acclimation of Photosynthesis to Environment 1 (APE1) has been identified as a protein involved in the adaptation of plants to high illumination. Several studies indicate its association with PSII, but its specific function in photosynthesis has not yet been determined. In this study, we used biochemical and molecular biology approaches to identify the function of APE1 in Arabidopsis (Arabidopsis thaliana) and the cyanobacterium Synechocystis sp. PCC 6803. APE1 elimination in the plant altered various, seemingly unrelated, photosynthetic processes, such as maximum quantum efficiency of PSII, non-photochemical quenching (NPQ), and state transitions. The levels of PSII core proteins were reduced in the ape1 mutant. Our results indicated that APE1 is involved in the formation or stabilization of the RC47 complex of PSII, in both plants and cyanobacteria. The alterations in NPQ and state transitions of the plant were a consequence of the lesion in PSII. These alterations were observed only when the mutant was stressed by high-light (HL) treatment, confirming that APE1 is necessary for optimal PSII performance under HL stress.
A complex and dynamic redox network regulates oxygen reduction at photosystem I in Arabidopsis
(Oxford University Press, 2024-09-26) Hani, Umama; Naranjo Río-Miranda, Belén; Shimakawa, Ginga; Espinasse, Christophe; Vanacker, Helene; Setif, Pierre; Rintamaki, Eevi; Issakidis-Bourguet, Emmanuelle; Krieger-Liszkay, Anja; Bioquímica Vegetal y Biología Molecular
Thiol-dependent redox regulation of enzyme activities plays a central role in regulating photosynthesis. Besides the regulation of metabolic pathways, alternative electron transport is subjected to thiol-dependent regulation. We investigated the regulation of O2 reduction at photosystem I. The level of O2 reduction in leaves and isolated thylakoid membranes depends on the photoperiod in which plants are grown. We used a set of Arabidopsis (Arabidopsis thaliana) mutant plants affected in the stromal, membrane, and lumenal thiol network to study the redox protein partners involved in regulating O2 reduction. Light-dependent O2 reduction was determined in leaves and thylakoids of plants grown in short-day and long-day conditions using a spin-trapping electron paramagnetic resonance assay. In wild-type samples from short-day conditions, reactive oxygen species generation was double that of samples from long-day conditions, while this difference was abolished in several redoxin mutants. An in vitro reconstitution assay showed that thioredoxin m, NADPH-thioredoxin reductase C, and NADPH are required for high O2-reduction levels in thylakoids from plants grown in long-day conditions. Using isolated photosystem I, we also showed that reduction of a photosystem I protein is responsible for the increase in O2 reduction. Furthermore, differences in the membrane localization of m-type thioredoxins and 2-Cys peroxiredoxin were detected between thylakoids of short-day and long-day plants. Overall, we propose a model of redox regulation of O2 reduction according to the reduction power of the stroma and the ability of different thiol-containing proteins to form a network of redox interactions.
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.
H263A and SCAN1/H493R mutant TDP1 block TOP1-induced double-strand break repair during gene transcription in quiescent cells and promote cell death
(Springer Nature, 2025-11) Rubio Contreras, Diana; Hidalgo-García, Daniel; Angulo-Jiménez, Carmen; Granado Calle, Esperanza; Sabio-Bonilla, Margarita; Ruiz Pérez, José Francisco; Gómez Herreros, Fernando; Bioquímica Vegetal y Biología Molecular; Genética; Ministerio de Ciencia, Innovación y Universidades (MICIU). España; Agencia Estatal de Investigación. España
DNA single-strand break (SSB) repair defects lead to hereditary neurological syndromes. Spinocerebellar ataxia with axonal neuropathy type 1 (SCAN1), is caused by the homozygous H493R mutation in tyrosyl-DNA phosphodiesterase 1 (TDP1), an enzyme that initiates the repair of DNA topoisomerase 1 (TOP1)-induced SSBs by unlinking the TOP1 peptide from the break. Although TDP1 also initiates the repair of TOP1-induced DNA double-strand breaks (DSBs) associated with transcription, the role of TOP1-induced DSBs in SCAN1 pathology remains unclear. Here, we have addressed the impact of the SCAN1/H493R mutation on the repair of TOP1-induced DSBs. We demonstrate that while TDP1 loss delays the repair of these breaks, SCAN1/H493R completely blocks it in RPE-1 quiescent cells. This blockage is specific to DSBs and is accompanied by a prolonged trapping of mutated TDP1 on DNA, but not of TOP1 cleavage complexes (TOP1cc). Intriguingly, the H263A inactivating mutation of TDP1, which accumulates TOP1cc, also blocks TOP1-induced DSB repair. Importantly, both SCAN1/H493R and H263A mutations exhibit genome instability and cell death. Moreover, we demonstrate that tyrosyl-DNA phosphodiesterase 2 (TDP2) can compensate for TDP1 loss in RPE-1 quiescent cells. Collectively, our data support the potential role of TOP1-induced DSBs as a main contributor to certain hereditary neurological syndromes.
ATG3 is subjected to redox regulation to quarantee ATG8 lipidation under ROS-generating stresses
(Taylor & Francis Group, 2024) Mallén Ponce, Manuel J.; Pérez-Pérez, María Esther; Bioquímica Vegetal y Biología Molecular; Ministerio de Ciencia e Innovación
The conjugation of ATG8 (autophagy-related 8) proteins to the lipid phospha-tidylethanolamine (PE) is the result of the coordinated and highly regulated action of several ATG core proteins, including ATG4 proteases and the E1 (ATG7)- and E2 (ATG3)- activating enzymes. Although it has been stablished that ROS signaling plays an important role in autophagy activation, the molecular mechanisms underlying the redox control of ATG proteins remain largely unclear. We have recently shown that ATG3 activity in Chlamydomonas rein- hardtii is subjected to reversible redox regulation to ensure ATG8 lipidation and autophagy progression under ROS-linked stress conditions.
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.
Enhanced non-enzymatic H2S generation extends lifespan and healthspan in male mice
(Elsevier, 2025-12-19) Cáliz-Molina, María Ángeles; López-Fernández-Sobrino, Raúl; Pino-Pérez, Inmaculada; Panadero-Morón, Concepción; Vilches-Pérez, María del Carmen; Camacho-Cabrera, María; García-Ruiz, Almudena; Pérez-Rosendo, Leopoldo; Espadas, Isabel; Venegas Calerón, Mónica; Aroca Aguilar, Ángeles; González Prieto, Román; Bernabeu Wittel, Máximo; Martín-Montalvo, Alejandro; Fisiología Médica y Biofísica; Bioquímica Vegetal y Biología Molecular; Biología Celular; Medicina; Ministerio de Ciencia e Innovación (MICIN). España; Junta de Andalucía; Sociedad Española de Diabetes (SED); Instituto de Salud Carlos III
Hydrogen sulfide is a gasotransmitter with biological functions, including roles in antioxidant defenses, mitochondrial bioenergetics, and cellular signaling via cysteine persulfidation. Several longevity-promoting interventions enhance endogenous hydrogen sulfide generation. However, whether enhanced hydrogen sulfide generation extends healthspan and lifespan in mammals remains unknown. Here, we investigated the in vivo effects of the non-enzymatic hydrogen sulfide generation promoted by natural diallyl sulforated compounds. Diallyl sulforated compounds extended lifespan and improved the main aspects of healthspan, including glucoregulation, locomotor function, and neurocognition in wild-type male mice across their lifespan. At the histological and molecular levels, we observed reductions in hepatic lipid-droplet size, attenuation of transcriptional and proteomic signatures associated with mTOR and immune-related pathways, and increased cysteine persulfidation in proteins. In humans, greater protein persulfidation in individuals with polypathological conditions was associated with increased muscle strength and lower triglyceride levels, supporting its physiological relevance. Our findings uncover the potential of enhanced hydrogen sulfide generation to promote healthy aging.
A quick-to-implement and optimized CRISPR-Cas9 protocol to obtain insertional and small indel mutants in Chlamydomonas reinhardtii
(Elsevier, 2025-12) De Silvio, Mariano A.; Sánchez-Retuerta, Camila; Ruiz Sola, María del Águila; Baidukova, Olga; Monte, Elena; Bioquímica Vegetal y Biología Molecular; Ministerio de Ciencia, Innovación y Universidades (MICIU). España; Agencia Estatal de Investigación
Chlamydomonas reinhardtii is a leading model organism in algal research, widely used to study photosynthesis, chloroplast and cilia biology, and more recently, metabolism, light signaling, the cell cycle, and algal biotechnology. Its sequenced genome has significantly accelerated research in the field, while improved genome-editing tools are key to advancing reverse genetics and genetic engineering. Building on previous advances, we present a streamlined and efficient CRISPR-Cas9 protocol for generating knockout mutants in Chlamydomonas via non-homologous end joining (NHEJ), using only commercially available reagents. Additionally, we introduce a cost-effective, PCR-based screening method capable of detecting mutants with large insertions as well as short indels -as small as one base pair- thereby enhancing overall CRISPR efficiency. • This protocol is easy to setup and can be fully executed using commercially available reagents. • This protocol allows for quick implementation and generation of mutants: 5 weeks from design to sequencing of candidate mutants. • This protocol describes a novel PCR-based strategy to identify mutants containing short indels. Screening is designed to identify large insertion mutants and the often overlooked small indel mutants.
Glucose Uncouples Nitrogen Sensing From Chlorosis via a Photosynthetic Checkpoint in Synechocystis sp. PCC 6803
(Wiley, 2025-11-21) Ortega Martínez, Pablo; Giner Lamia, Joaquín; Wey, Laura T.; Muro Pastor, María Isabel; Florencio Bellido, Francisco Javier; Díaz Troya, Sandra; Bioquímica Vegetal y Biología Molecular; Ministerio de Ciencia, Innovación y Universidades (MICIU). España; Novo Nordisk Foundation
Cyanobacteria adapt to nitrogen starvation by undergoing chlorosis, a regulated bleaching process that involves the degradation of phycobilisomes, the light-harvesting antennae complexes, and accumulation of glycogen. While this response is well characterized under photoautotrophic conditions, its modulation by external organic carbon sources such as glucose remains poorly characterized. Here, we investigated how glucose affects the response to nitrogen deprivation in the model cyanobacterium Synechocystis sp. PCC 6803. Using an integrative approach combining physiological assays, targeted metabolomics, RNA sequencing, chlorophyll fluorescence and absorbance spectroscopy, we studied the underlying regulatory mechanisms, focusing on photosynthetic electron transport. Glucose supplementation prevented bleaching, even when added after nitrogen deprivation symptoms had begun. This effect was associated with excess glycogen accumulation, disrupted carbon partitioning, and buildup of metabolic intermediates, indicating a metabolic overflow. Despite these physiological differences, transcriptomic responses to nitrogen deprivation were largely similar regardless of glucose supplementation, suggesting regulation at the post-transcriptional or metabolic level. Glucose also impaired photosynthetic electron transport by creating a redox bottleneck at the photosystem II (PSII) acceptor side, leading to decreased electron transport to photosystem I (PSI) and oxidation of the P700 pool. These findings suggest that reduction of the P700 acceptor side is required to trigger chlorosis. Our results demonstrate that glucose uncouples nitrogen sensing from the bleaching process by altering photosynthetic electron flow. We propose the existence of a redox-sensitive checkpoint that integrates metabolic state with photosynthetic performance, offering new insights into stress adaptation in cyanobacteria.
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.
Halotolerant black yeast Neophaeotheca triangularis as a source of melanin
(Elsevier, 2026-02) Peña, Mercedes; Gómez Villegas, Patricia; Prados, Jose; Melguizo, Consolación; Carvalho, Carla C.C.R.; Bioquímica Vegetal y Biología Molecular; European Union (UE); Fundação para a Ciência e a Tecnologia (FCT). Portugal; Ministerio de Ciencia, Innovación y Universidades (MICIU). España
Archaea and bacteria are the most studied extremophiles, but fungi also demonstrate remarkable tolerance, particularly in hypersaline environments such as solar salterns. Among salt-adapted fungi, black yeasts have been shown to be adapted to such environments, having developed defense mechanisms such as the production of melanin, a pigment that plays a crucial role in environmental stress protection. Melanin is a complex, high-molecular-weight polymer widely found across biological kingdoms. During the isolation of microorganisms from samples collected in crystallization ponds in a saltern, a black yeast was found able to produce significant amounts of melanin. The yeast was identified as belonging to the species Neophaeotheca triangularis. The aim of this study was to optimize the cultivation conditions of the strain, to characterize the produced melanin, and to assess its biological activities, including its antitumor and antioxidant properties.

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