Person: Solís González, María Teresa
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First Name
María Teresa
Last Name
Solís González
Affiliation
Universidad Complutense de Madrid
Faculty / Institute
Ciencias Biológicas
Department
Genética, Fisiología y Microbiología
Area
Fisiología Vegetal
Identifiers
10 results
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Item Epigenetic changes accompany developmental programmed cell death in tapetum cells(Plant and Cell Physiology, 2014) Chakrabarti, N; Testillano, PS; Risueño, MC; Cortés-Eslava, J; Rodríguez-Serrano, M; Bioggiogera, M; Corredor, E; Solís González, María TeresaThe tapetum, the nursing tissue inside anthers, undergoes cellular degradation by programmed cell death (PCD) during late stages of microspore-early pollen development. Despite the key function of tapetum, little is known about the molecular mechanisms regulating this cell death process in which profound nuclear and chromatin changes occur. Epigenetic features (DNA methylation and histone modifications) have been revealed as hallmarks that establish the functional status of chromatin domains, but no evidence on the epigenetic regulation of PCD has been reported. DNA methylation is accomplished by DNA methyltransferases, among which DNA methyl transferase 1 (MET1) constitutes one of the CG maintenance methyltransferase in plants, also showing de novo methyltransferase activity. In this work, the changes in epigenetic marks during the PCD of tapetal cells have been investigated by a multidisciplinary approach to reveal the dynamics of DNA methylation and the pattern of expression of MET1 in relation to the main cellular changes of this PCD process which have also been characterized in two species, Brassica napus and Nicotiana tabacum. The results showed that tapetum PCD progresses with the increase in global DNA methylation and MET1 expression, epigenetic changes that accompanied the reorganization of the nuclear architecture and a high chromatin condensation, activity of caspase 3-like proteases and Cyt c release. The reported data indicate a relationship between the PCD process and the DNA methylation dynamics and MET1 expression in tapetal cells, suggesting a possible new role for the epigenetic marks in the nuclear events occurring during this cell death process and providing new insights into the epigenetic control of plant PCD.Item Initiation of leaf somatic embryogenesis involves high pectin esterification, auxin accumulation and DNA demethylation in Quercus alba(Journal of Plant Physiology, 2017) Cano, V; Solís González, María Teresa; Testillano, PS; Corredoira, E; Barany, I; Rodriguez-Sanz, H; Vieitez, A; Risueño, MCSomatic embryogenesis is considered a convenient tool for investigating the regulating mechanisms of embryo formation; it is also a feasible system for in vitro regeneration procedures, with many advantages in woody species. Nevertheless, trees have shown recalcitrance to somatic embryogenesis, and its efficiency remains very low in many cases. Consequently, despite the clear potential of somatic embryogenesis in tree breeding programs, its application is limited since factors responsible for embryogenesis initiation have not yet been completely elucidated. In the present work, we investigated key cellular factors involved in the change of developmental program during leaf somatic embryogenesis initiation of white oak (Quercus alba), aiming to identify early markers of the process. The results revealed that pectin esterification, auxin accumulation and DNA demethylation were induced during embryogenesis initiation and differentially found in embryogenic cells, while they were not present in leaf cells before induction or in non-embryogenic cells after embryogenesis initiation. These three factors constitute early markers of leaf embryogenesis and represent processes that could be interconnected and involved in the regulation of cell reprogramming and embryogenesis initiation. These findings provide new insights into the mechanisms underlying plant cell reprogramming, totipotency and embryogenic competence acquisition, especially in tree species for which information is scarce, thus opening up the possibility of efficient manipulation of somatic embryogenesis induction.Item DNA methylation dynamics and MET1a-like gene expression changes during stress-induced pollen reprogramming to embryogenesis(Journal of Experimental Botany, 2012) Rodríguez-Serrano, M; Meijón, M; Cañal, MJ; Cifuentes, A; Risueño, MC; Testillano, PS; Solís González, María TeresaStress-induced plant cell reprogramming involves changes in global genome organization, being the epigenetic modifications key factors in the regulation of genome flexibility. DNA methylation, accomplished by DNA methyltransferases, constitutes a prominent epigenetic modification of the chromatin fibre which is locked in a transcriptionally inactive conformation. Changes in DNA methylation accompany the reorganization of the nuclear architecture during plant cell differentiation and proliferation. After a stress treatment, in vitro-cultured microspores are reprogrammed and change their gametophytic developmental pathway towards embryogenesis, the process constituting a useful system of reprogramming in isolated cells for applied and basic research. Gene expression driven by developmental and stress cues often depends on DNA methylation; however, global DNA methylation and genome-wide expression patterns relationship is still poorly understood. In this work, the dynamics of DNA methylation patterns in relation to nuclear architecture and the expression of BnMET1a-like DNA methyltransferase genes have been analysed during pollen development and pollen reprogramming to embryogenesis in Brassica napus L. by a multidisciplinary approach. Results showed an epigenetic reprogramming after microspore embryogenesis induction which involved a decrease of global DNA methylation and its nuclear redistribution with the change of developmental programme and the activation of cell proliferation, while DNA methylation increases with pollen and embryo differentiation in a cell-type-specific manner. Changes in the presence, abundance, and distribution of BnMET1a-like transcripts highly correlated with variations in DNA methylation. Mature zygotic and pollen embryos presented analogous patterns of DNA methylation and MET1a-like expression, providing new evidence of the similarities between both developmental embryogenic programmes.Item Stress-Induced Microspore Embryogenesis Requires Endogenous Auxin Synthesis and Polar Transport in Barley.(Frontiers in Plant Science, 2019) Pérez-Pérez, Yolanda; Solís González, María Teresa; Solís, María Teresa; Testillano, PS; Risueño, Maía del Carmen; El-Tantawy, AhmedStress-induced microspore embryogenesis is a model in vitro system of cell reprogramming, totipotency acquisition, and embryo development. After induction, responsive microspores abandon their developmental program to follow an embryogenic pathway, leading to in vitro embryo formation. This process is widely used to produce doubled-haploid lines, essential players to create new materials in modern breeding programs, particularly in cereals, although its efficiency is still low in many crop species, because the regulating mechanisms are still elusive. Stress signaling and endogenous hormones, mainly auxin, have been proposed as determinant factors of microspore embryogenesis induction in some eudicot species; however, much less information is available in monocot plants. In this study, we have analyzed the dynamics and possible role of endogenous auxin during stress-induced microspore embryogenesis in the monocot Hordeum vulgare, barley. The results showed auxin accumulation in early proembryo cells, from embryogenesis initiation and a further increase with embryo development and differentiation, correlating with the induction and expression pattern of the auxin biosynthesis gene HvTAR2-like. Pharmacological treatments with kynurenine, inhibitor of auxin biosynthesis, and α-(p-chlorophenoxy)-isobutyric acid (PCIB), auxin antagonist, impaired embryogenesis initiation and development, indicating that de novo auxin synthesis and its activity were required for the process. Efflux carrier gene HvPIN1-like was also induced with embryogenesis initiation and progression; auxin transport inhibition by N-1-naphthylphthalamic acid significantly reduced embryo development at early and advanced stages. The results indicate activation of auxin biosynthesis with microspore embryogenesis initiation and progression, in parallel with the activation of polar auxin transport, and reveal a central role of auxin in the process in a monocot species. The findings give new insights into the complex regulation of stress-induced microspore embryogenesis, particularly in monocot plants for which information is still scarce, and suggest that manipulation of endogenous auxin content could be a target to improve in vitro embryo production.Item Pectin de-methylesterification and AGP increase promote cell wall remodeling and are required during somatic embryogenesis of Quercus suber.(Frontiers in Plant Science, 2019) Pérez-Pérez, Yolanda; Carneros, Elena; Berenguer, Eduardo; Solís González, María Teresa; Bárány, Ivett; Pintos López, Beatriz; Gómez Garay, María Aranzazu; Risueño, María del Carmen; Testillano, PSSomatic embryogenesis is a reliable system for in vitro plant regeneration, with biotechnological applications in trees, but the regulating mechanisms are largely unknown. Changes in cell wall mechanics controlled by methylesterification of pectins, mediated by pectin methylesterases (PMEs) and pectin methyl esterase inhibitors (PMEIs) underlie many developmental processes. Arabinogalactan proteins (AGPs) are highly glycosylated proteins located at the surface of plasma membranes, in cell walls, and in extracellular secretions, with key roles in a range of different processes. In this study, we have investigated changes in two cell wall components, pectins and AGPs, during somatic embryogenesis in Quercus suber, a forest tree of high economic and ecologic value. At early embryogenesis stages, cells of proembryogenic masses showed high levels of esterified pectins and expression of QsPME and QsPMEI genes encoding a PME and a putative PMEI, respectively. At advanced stages, differentiating cells of heart, torpedo and cotyledonary embryos exhibited walls rich in de-esterified pectins, while QsPME gene expression and PME activity progressively increased. AGPs were detected in cell walls of proembryogenic masses and somatic embryos. QsLys-rich-AGP18, QsLys-rich-AGP17, and QsAGP16L1 gene expression increased with embryogenesis progression, as did the level of total AGPs, detected by dot blot with β-glucosyl Yariv reagent. Immuno dot blot, immunofluorescence assays and confocal analysis using monoclonal antibodies to high- (JIM7, LM20) and low- (JIM5, LM19) methylesterified pectins, and to certain AGP epitopes (LM6, LM2) showed changes in the amount and distribution pattern of esterified/de-esterified pectins and AGP epitopes, that were associated with proliferation and differentiation and correlated with expression of the PME and AGP genes analyzed. Pharmacological treatments with catechin, an inhibitor of PME activity, and Yariv reagent, which blocks AGPs, impaired the progression of embryogenesis, with pectin de-esterification and an increase in AGP levels being necessary for embryo development. Findings indicate a role for pectins and AGPs during somatic embryogenesis of cork oak, promoting the cell wall remodeling during the process. They also provide new insights into the regulating mechanisms of somatic embryogenesis in woody species, for which information is still scarce, opening up new possibilities to improve in vitro embryo production in tree breeding.Item Auxin biosynthesis, accumulation, action and transport are involved in stress-induced microspore embryogenesis initiation and progression in Brassica napus.(Plant and Cell physiology, 2015) Rodriguez-Sanz, H; Solís González, María Teresa; Testillano, PS; Lopez, MF; Gomez Cadenas, A; Risueño, MCIsolated microspores are reprogrammed in vitro by stress, becoming totipotent cells and producing embryos and plants via a process known as microspore embryogenesis. Despite the abundance of data on auxin involvement in plant development and embryogenesis, no data are available regarding the dynamics of auxin concentration, cellular localization and the expression of biosynthesis genes during microspore embryogenesis. This work involved the analysis of auxin concentration and cellular accumulation; expression of TAA1 and NIT2 encoding enzymes of two auxin biosynthetic pathways; expression of the PIN1-like efflux carrier; and the effects of inhibition of auxin transport and action by N-1-naphthylphthalamic acid (NPA) and α-(p-chlorophenoxy) isobutyric acid (PCIB) during Brassica napus microspore embryogenesis. The results indicated de novo auxin synthesis after stress-induced microspore reprogramming and embryogenesis initiation, accompanying the first cell divisions. The progressive increase of auxin concentration during progression of embryogenesis correlated with the expression patterns of TAA1 and NIT2 genes of auxin biosynthetic pathways. Auxin was evenly distributed in early embryos, whereas in heart/torpedo embryos auxin was accumulated in apical and basal embryo regions. Auxin efflux carrier PIN1-like gene expression was induced in early multicellular embryos and increased at the globular/torpedo embryo stages. Inhibition of polar auxin transport (PAT) and action, by NPA and PCIB, impaired embryo development, indicating that PAT and auxin action are required for microspore embryo progression. NPA also modified auxin embryo accumulation patterns. These findings indicate that endogenous auxin biosynthesis, action and polar transport are required in stress-induced microspore reprogramming, embryogenesis initiation and progression.Item Inhibition of histone H3K9 methylation by BIX-01294 promotes stress-induced microspore totipotency and enhances embryogenesis initiation.(Frontiers in Plant Science, 2017) Berenguer, Eduardo; Solís González, María Teresa; Testillano, PS; Perez-Perez, YMicrospore embryogenesis is a process of cell reprogramming, totipotency acquisition and embryogenesis initiation, induced in vitro by stress treatments and widely used in plant breeding for rapid production of doubled-haploids, but its regulating mechanisms are still largely unknown. Increasing evidence has revealed epigenetic reprogramming during microspore embryogenesis, through DNA methylation, but less is known about the involvement of histone modifications. In this study, we have analyzed the dynamics and possible role of histone H3K9 methylation, a major repressive modification, as well as the effects on microspore embryogenesis initiation of BIX-01294, an inhibitor of histone methylation, tested for the first time in plants, in Brassica napus and Hordeum vulgare. Results revealed that microspore reprogramming and initiation of embryogenesis involved a low level of H3K9 methylation. With the progression of embryogenesis, methylation of H3K9 increased, correlating with gene expression profiles of BnHKMT SUVR4-like and BnLSD1-like (writer and eraser enzymes of H3K9me2). At early stages, BIX-01294 promoted cell reprogramming, totipotency and embryogenesis induction, while diminishing bulk H3K9 methylation. DNA methylation was also reduced by short-term BIX-01294 treatment. By contrast, long BIX-01294 treatments hindered embryogenesis progression, indicating that H3K9 methylation is required for embryo differentiation. These findings open up new possibilities to enhance microspore embryogenesis efficiency in recalcitrant species through pharmacological modulation of histone methylation by using BIX-01294.Item Arabinogalactan proteins profiles and distribution patterns during pollen development and pollen embryogenesis of Brassica napus(Plant Reproduction, 2013) Solís González, María Teresa; El Tantawy, A; Da costa, ML; Testillano, PS; Coimbra, Silvia; Risueño, MCArabinogalactan proteins (AGPs), present in cell walls, plasma membranes and extracellular secretions, are massively glycosylated hydroxyproline-rich proteins that play a key role in several plant developmental processes. After stress treatment, microspores cultured in vitro can reprogramme and change their gametophytic developmental pathways towards embryogenesis, thereby producing embryos which can further give rise to haploid and double haploid plants, important biotechnological tools in plant breeding. Microspore embryogenesis constitutes a convenient system for studying the mechanisms underlying cell reprogramming and embryo formation. In this work, the dynamics of both AGP presence and distribution were studied during pollen development and microspore embryogenesis in Brassica napus, by employing a multidisciplinary approach using monoclonal antibodies for AGPs (LM2, LM6, JIM13, JIM14, MAC207) and analysing the expression pattern of the BnAGP Sta 39–4 gene. Results showed the developmental regulation and defined localization of the studied AGP epitopes during the two microspore developmental pathways, revealing different distribution patterns for AGPs with different antigenic reactivity. AGPs recognized by JIM13, JIM14 and MAC207 antibodies were related to pollen maturation, whereas AGPs labelled by LM2 and LM6 were associated with embryo development. Interestingly, the AGPs labelled by JIM13 and JIM14 were induced with the change of microspore fate. Increases in the expression of the Sta 39–4 gene, JIM13 and JIM14 epitopes found specifically in 2–4 cell stage embryo cell walls, suggested that AGPs are early molecular markers of microspore embryogenesis. Later, LM2 and LM6 antigens increased progressively with embryo development and localized on cell walls and cytoplasmic spots, suggesting an active production and secretion of AGPs during in vitro embryo formation. These results give new insights into the involvement of AGPs as potential regulating/signalling molecules in microspore reprogramming and embryogenesis.Item Proteases with caspase 3-like activity participate in cell death during stress-induced microspore embryogenesis of «Brassica napus.(The EuroBiotech Journal., 2019) Berenguer, Eduardo; Solís González, María Teresa; Pérez-Perez, Y; Testillano, PSMicrospore embryogenesis is a model system of plant cell reprogramming, totipotency acquisition, stress response and embryogenesis initiation. This in vitro system constitutes an important biotechnological tool for haploid and doubled-haploid plant production, very useful for crop breeding. In this process, microspores (cells that produce pollen grains in planta) are reprogrammed toward embryogenesis by specific stress treatment, but many microspores die after the stress. The occurrence of cell death is a serious limiting problem that greatly reduces microspore embryogenesis yield. In animals, increasing evidence has revealed caspase proteolytic activities as essential executioners of programmed cell death (PCD) processes, however, less is known in plants. Although plant genomes do not contain caspase homologues, caspase-like proteolytic activities have been detected in many plant PCD processes. In the present study, we have analysed caspase 3-like activity and its involvement in stress-induced cell death during initial stages of microspore embryogenesis of Brassica napus. After stress treatment to induce embryogenesis, isolated microspore cultures showed high levels of cell death and caspase 3-like proteolytic activity was induced. Treatments with specific inhibitor of caspase 3-like activity reduced cell death and increased embryogenesis induction efficiency. Our findings indicate the involvement of proteases with caspase 3-like activity in the initiation and/or execution of cell death at early microspore embryogenesis in B. napus, giving new insights into the pathways of stress-induced cell death in plants and opening a new way to improve in vitro embryogenesis efficiency by using chemical modulators of cell death proteases.Item BnPME is progressively induced after microspore reprogramming to embryogenesis, correlating with pectin de-esterification and cell differentiation in Brassica napus(BMC Plant Biology, 2016) Solís González, María Teresa; Testillano, PS; Risueño, MC; Berenguer, EBackground: Pectins are one of the main components of plant cell walls. They are secreted to the wall as highly methylesterified forms that can be de-esterified by pectin methylesterases (PMEs). The degree of methylesterification of pectins changes during development, PMEs are involved in the cell wall remodeling that occurs during diverse plant developmental processes. Nevertheless, the functional meaning of pectin-related wall remodeling in different cell types and processes remains unclear. In vivo, the microspore follows the gametophytic pathway and differentiates to form the pollen grain. In vitro, the microspore can be reprogrammed by stress treatments becoming a totipotent cell that starts to proliferate and follows the embryogenic pathway, a process known as microspore embryogenesis. Results: To investigate if the change of developmental programme of the microspore towards embryogenesis involves changes in pectin esterification levels, which would cause the cell wall remodeling during the process, in the present study, dynamics of PME expression and degrees of pectin esterification have been analysed during microspore embryogenesis and compared with the gametophytic development, in Brassica napus. A multidisciplinary approach has been adopted including BnPME gene expression analysis by quantitative RT-PCR, fluorescence in situ hybridization, immuno-dot-blot and immunofluorescence with JIM5 and JIM7 antibodies to reveal low and highly-methylesterified pectins. The results showed that cell differentiation at advanced developmental stages involved induction of BnPME expression and pectin de-esterification, processes that were also detected in zygotic embryos, providing additional evidence that microspore embryogenesis mimics zygotic embryogenesis. By contrast, early microspore embryogenesis, totipotency and proliferation were associated with low expression of BnPME and high levels of esterified pectins. Conclusions: The results show that the change of developmental programme of the microspore involves changes in pectin esterification associated with proliferation and differentiation events, which may cause the cell wall remodeling during the process. The findings indicate pectin-related modifications in the cell wall during microspore embryogenesis, providing new insights into the role of pectin esterification and cell wall configuration in microspore totipotency, embryogenesis induction and progression.