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If one takes a multidisciplinary, integrative perspective on historical social psychology, one sees that it is a vital thread not only in the theoretical weave of social psychology as such, but in any social science which studies the social being. The multidisciplinary character of historical social psychology is friendly to authors and ideas from other domains of knowledge. Marañón's insights suggest interesting ways of answering the main questions that arise in historical social psychology. The application of his method, as I shall try to show, can orient to us towards a social psychology concerned not only with the here and now of its object of study, but also with the way in which it has evolved through history.

Advertising in general, and television advertising in particular, is an important communication channel through which values, lifestyles, and even the socially-shared "imaginary" are transmitted. A psycho-social analysis of the imaginary contents in television advertisements is specially called-for, given that contemporary culture, strongly marked by the use of the images, constitutes what some theoreticians call "the audio-visual age". In this article we analyse advertisements from a psycho-social perspective, to identify the way in which advertisements articulate, through images, the social imaginary.

In Arabidopsis, the plastidial isoform of phosphoglucose isomerase, PGI1, mediates growth and photosynthesis, likely due to its involvement in the vascular production of cytokinins (CK). To examine this hypothesis, we characterized pgi1-2 knockout plants impaired in PGI1 and pgi1-2 plants specifically expressing PGI1 in root tips and vascular tissues. Moreover, to investigate whether the phenotype of pgi1-2 plants is due to impairments in the plastidial oxidative pentose phosphate pathway (OPPP) or the glycolytic pathway, we characterized pgl3-1 plants with reduced OPPP and pfk4pfk5 knockout plants impaired in plastidial glycolysis. Compared with wild-type (WT) leaves, pgi1-2 leaves exhibited weaker expression of photosynthesis- and 2-C-methyl-D-erythritol 4-P (MEP) pathway-related proteins, and stronger expression of oxidative stress protection-related enzymes. Consistently, pgi1-2 leaves accumulated lower levels of chlorophyll, and higher levels of tocopherols, flavonols and anthocyanins than the WT. Vascular- and root tip-specific PGI1 expression countered the reduced photosynthesis, low MEP pathway-derived CK content, dwarf phenotype and the metabolic characteristics of pgi1-2 plants, reverting them to WT-like levels. Moreover, pgl3-1, but not pfk4pfk5 plants phenocopied pgi1-2. Histochemical analyses of plants expressing GUS under the control of promoter regions of genes encoding plastidial OPPP enzymes exhibited strong GUS activity in root tips and vascular tissues. Overall, our findings show that root tip and vascular PGI1-mediated plastidial OPPP activity affects photosynthesis and growth through mechanisms involving long-distance modulation of the leaf proteome by MEP pathway-derived CKs., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

Background: Exercise has shown beneficial effects on neuronal neuroplasticity; therefore, we want to analyze the influence of high-intensity interval training (HIIT) on neuroplasticity markers in post-stroke patients. Methods: A systematic review of RCTs including studies with stroke participants was conducted using the following databases (PubMed, LILACS, ProQuest, PEDro, Web of Science). Searches lasted till (20/11/2023). Studies that used a HIIT protocol as the main treatment or as a coadjutant treatment whose outcomes were neural plasticity markers were used and compared with other exercise protocols, controls or other kinds of treatment. Studies that included other neurological illnesses, comorbidities that interfere with stroke or patients unable to complete a HIIT protocol were excluded. HIIT protocol, methods to assess intensity, neuroplasticity markers (plasmatic and neurophysiological) and other types of assessments such as cognitive scales were extracted to make a narrative synthesis. Jadad and PEDro scales were used to assess bias. Results: Eight articles were included, one included lacunar stroke (less than 3 weeks) and the rest had chronic stroke. The results found here indicate that HIIT facilitates neuronal recovery in response to an ischemic injury. This type of training increases the plasma concentrations of lactate, BDNF and VEGF, which are neurotrophic and growth factors involved in neuroplasticity. HIIT also positively regulates other neurophysiological measurements that are directly associated with a better outcome in motor learning tasks. Conclusions: We conclude that HIIT improves post-stroke recovery by increasing neuroplasticity markers. However, a limited number of studies have been found indicating that future studies are needed that assess this effect and include the analysis of the number of intervals and their duration in order to maximize this effect.

The worldwide incidence of COVID-19 has generated a pandemic of sequelae. These sequelae require multidisciplinary rehabilitative work to address the multisystemic symptoms that patients will present with now and in the future. The aim of the present systematic review is to analyze the current situation of telerehabilitation in patients with COVID-19 sequelae and its effectiveness. Searches were conducted on the following databases: PubMed, Scopus, PEDro, and Web of Science (WOS). There was no complete homogeneity among the five selected articles, so we differentiated two clinical subgroups for the clustering of outcome measures: (group one) patients with post-discharge symptoms and (group two) patients with permanent symptoms or “long COVID-19” defined as persistent symptoms > 2 months. For group one, post-discharge sequelae, improvements were obtained in cardiovascular parameters, and physical test studies in group two presented very favorable results in all the cardiorespiratory measures and physical tests evaluated. Telerehabilitation through therapeutic exercise based on mixed protocols of aerobic, respiratory, and low-load strength exercises appear to be an effective and safe strategy for the recovery of short- and long-term post-COVID-19 sequelae.

In Arabidopsis (Arabidopsis thaliana), the plastidial isoform of phosphoglucose isomerase (PGI1) mediates photosynthesis, metabolism, and development, probably due to its involvement in the synthesis of isoprenoid-derived signals in vascular tissues. Microbial volatile compounds (VCs) with molecular masses of <45 Da promote photosynthesis, growth, and starch overaccumulation in leaves through PGI1-independent mechanisms. Exposure to these compounds in leaves enhances the levels of GLUCOSE-6-PHOSPHATE/PHOSPHATE TRANSLOCATOR2 (GPT2) transcripts. We hypothesized that the PGI1-independent response to microbial volatile emissions involves GPT2 action. To test this hypothesis, we characterized the responses of wild-type (WT), GPT2-null gpt2-1, PGI1-null pgi1-2, and pgi1-2gpt2-1 plants to small fungal VCs. In addition, we characterized the responses of pgi1-2gpt2-1 plants expressing GPT2 under the control of a vascular tissue- and root tip-specific promoter to small fungal VCs. Fungal VCs promoted increases in growth, starch content, and photosynthesis in WT and gpt2-1 plants. These changes were substantially weaker in VC-exposed pgi1-2gpt2-1 plants but reverted to WT levels with vascular and root tip-specific GPT2 expression. Proteomic analyses did not detect enhanced levels of GPT2 protein in VC-exposed leaves and showed that knocking out GPT2 reduced the expression of photosynthesis-related proteins in pgi1-2 plants. Histochemical analyses of GUS activity in plants expressing GPT2-GUS under the control of the GPT2 promoter showed that GPT2 is mainly expressed in root tips and vascular tissues around hydathodes. Overall, the data indicated that the PGI1-independent response to microbial VCs involves resetting of the photosynthesis-related proteome in leaves through long-distance GPT2 action., (© The Author(s) 2022. Published by Oxford University Press on behalf of American Society of Plant Biologists.)

Microorganisms communicate with plants by exchanging chemical signals throughout the phytosphere. Before direct contact with plants occurs, beneficial microorganisms emit a plethora of volatile compounds that promote plant growth and photosynthesis as well as developmental, metabolic, transcriptional, and proteomic changes in plants. These compounds can also induce systemic drought tolerance and improve water and nutrient acquisition. Recent studies have shown that this capacity is not restricted to beneficial microbes; it also extends to phytopathogens. Plant responses to microbial volatile compounds have frequently been associated with volatile organic compounds with molecular masses ranging between ~ 45Da and 300Da. However, microorganisms also release a limited number of volatile compounds with molecular masses of less than ~45Da that react with proteins and/or act as signaling molecules. Some of these compounds promote photosynthesis and growth when exogenously applied in low concentrations. Recently, evidence has shown that small volatile compounds are important determinants of plant responses to microbial volatile emissions. However, the regulatory mechanisms involved in these responses remain poorly understood. This review summarizes current knowledge of biochemical and molecular mechanisms involved in plant growth, development, and metabolic responses to small microbial volatile compounds., (© The Author(s) 2021. Published by Oxford University Press on behalf of the Society for Experimental Biology. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Plants communicate with microorganisms by exchanging chemical signals throughout the phytosphere. Such interactions are important not only for plant productivity and fitness, but also for terrestrial ecosystem functioning. It is known that beneficial microorganisms emit diffusible substances including volatile organic compounds (VOCs) that promote growth. Consistently, soil application of cell-free culture filtrates (CF) of beneficial soil and plant-associated microorganisms enhances plant growth and yield. However, how this treatment acts in plants and whether it alters the resident soil microbiota, are largely unknown. In this work we characterized the responses of pepper ( Capsicum annuum L.) plants cultured under both greenhouse and open field conditions and of soil microbiota to soil application of CFs of beneficial and phytopathogenic fungi. To evaluate the contribution of VOCs occurring in the CFs to these responses, we characterized the responses of plants and of soil microbiota to application of distillates (DE) of the fungal CFs. CFs and their respective DEs contained the same potentially biogenic VOCs, and application of these extracts enhanced root growth and fruit yield, and altered the nutritional characteristics of fruits. High-throughput amplicon sequencing of bacterial 16S and fungal ITS rRNA genes of the soil microbiota revealed that the CF and DE treatments altered the microbial community compositions, and led to strong enrichment of the populations of the same beneficial bacterial and fungal taxa. Our findings show that CFs of both beneficial and phytopathogenic fungi can be used as biostimulants, and provide evidence that VOCs occurring in the fungal CFs act as mediators of the plants' responses to soil application of fungal CFs through stimulation of the beneficial soil microbiota., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Baroja-Fernández, Almagro, Sánchez-López, Bahaji, Gámez-Arcas, De Diego, Dolezal, Muñoz, Climent Sanz and Pozueta-Romero.)

Microorganisms produce volatile compounds (VCs) with molecular masses of less than 300 Da that promote plant growth and photosynthesis. Recently, we have shown that small VCs of less than 45 Da other than CO 2 are major determinants of plant responses to fungal volatile emissions. However, the regulatory mechanisms involved in the plants' responses to small microbial VCs remain unclear. In Arabidopsis thaliana plants exposed to small fungal VCs, growth promotion is accompanied by reduction of the thiol redox of Calvin-Benson cycle (CBC) enzymes and changes in the levels of shikimate and 2-C-methyl-D-erythritol 4-phosphate (MEP) pathway-related compounds. We hypothesized that plants' responses to small microbial VCs involve post-translational modulation of enzymes of the MEP and shikimate pathways via mechanisms involving redox-activated photosynthesis signaling. To test this hypothesis, we compared the responses of wild-type (WT) plants and a cfbp1 mutant defective in a redox-regulated isoform of the CBC enzyme fructose-1,6-bisphosphatase to small VCs emitted by the fungal phytopathogen Alternaria alternata. Fungal VC-promoted growth and photosynthesis, as well as metabolic and proteomic changes, were substantially weaker in cfbp1 plants than in WT plants. In WT plants, but not in cfbp1 plants, small fungal VCs reduced the levels of both transcripts and proteins of the stromal Clp protease system and enhanced those of plastidial chaperonins and co-chaperonins. Consistently, small fungal VCs promoted the accumulation of putative Clp protease clients including MEP and shikimate pathway enzymes. clpr1-2 and clpc1 mutants with disrupted plastidial protein homeostasis responded weakly to small fungal VCs, strongly indicating that plant responses to microbial volatile emissions require a finely regulated plastidial protein quality control system. Our findings provide strong evidence that plant responses to fungal VCs involve chloroplast-to-nucleus retrograde signaling of redox-activated photosynthesis leading to proteostatic regulation of the MEP and shikimate pathways., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Ameztoy, Sánchez-López, Muñoz, Bahaji, Almagro, Baroja-Fernández, Gámez-Arcas, De Diego, Doležal, Novák, Pěnčík, Alpízar, Rodríguez-Concepción and Pozueta-Romero.)

Volatile compounds (VCs) emitted by the fungal phytopathogen Penicillium aurantiogriseum promote root growth and developmental changes in Arabidopsis. Here we characterised the metabolic and molecular responses of roots to fungal volatiles. Proteomic analyses revealed that these compounds reduce the levels of aquaporins, the iron carrier IRT1 and apoplastic peroxidases. Fungal VCs also increased the levels of enzymes involved in the production of mevalonate (MVA)-derived isoprenoids, nitrogen assimilation and conversion of methionine to ethylene and cyanide. Consistently, fungal VC-treated roots accumulated high levels of hydrogen peroxide (H 2 O 2 ), MVA-derived cytokinins, ethylene, cyanide and long-distance nitrogen transport amino acids. qRT-PCR analyses showed that many proteins differentially expressed by fungal VCs are encoded by VC non-responsive genes. Expression patterns of hormone reporters and developmental characterisation of mutants provided evidence for the involvement of cyanide scavenging and enhanced auxin, ethylene, cytokinin and H 2 O 2 signalling in the root architecture changes promoted by fungal VCs. Our findings show that VCs from P. aurantiogriseum modify root metabolism and architecture, and improve nutrient and water use efficiencies through transcriptionally and non-transcriptionally regulated proteome resetting mechanisms. Some of these mechanisms are subject to long-distance regulation by photosynthesis and differ from those triggered by VCs emitted by beneficial microorganisms., (© 2020 John Wiley & Sons Ltd.)

Microorganisms produce volatile compounds (VCs) that promote plant growth and photosynthesis through complex mechanisms involving cytokinin (CK) and abscisic acid (ABA). We hypothesized that plants' responses to microbial VCs involve posttranslational modifications of the thiol redox proteome through action of plastidial NADPH-dependent thioredoxin reductase C (NTRC), which regulates chloroplast redox status via its functional relationship with 2-Cys peroxiredoxins. To test this hypothesis, we analysed developmental, metabolic, hormonal, genetic, and redox proteomic responses of wild-type (WT) plants and a NTRC knockout mutant (ntrc) to VCs emitted by the phytopathogen Alternaria alternata. Fungal VC-promoted growth, changes in root architecture, shifts in expression of VC-responsive CK- and ABA-regulated genes, and increases in photosynthetic capacity were substantially weaker in ntrc plants than in WT plants. As in WT plants, fungal VCs strongly promoted growth, chlorophyll accumulation, and photosynthesis in ntrc-Δ2cp plants with reduced 2-Cys peroxiredoxin expression. OxiTRAQ-based quantitative and site-specific redox proteomic analyses revealed that VCs promote global reduction of the thiol redox proteome (especially of photosynthesis-related proteins) of WT leaves but its oxidation in ntrc leaves. Our findings show that NTRC is an important mediator of plant responses to microbial VCs through mechanisms involving global thiol redox proteome changes that affect photosynthesis., (© 2019 John Wiley & Sons Ltd.)

A "box-in-box" cocultivation system was used to investigate plant responses to microbial volatile compounds (VCs) and to evaluate the contributions of organic and inorganic VCs (VOCs and VICs, respectively) to these responses. Arabidopsis plants were exposed to VCs emitted by adjacent Alternaria alternata and Penicillium aurantiogriseum cultures, with and without charcoal filtration. No VOCs were detected in the headspace of growth chambers containing fungal cultures with charcoal filters. However, these growth chambers exhibited elevated CO 2 and bioactive CO and NO headspace concentrations. Independently of charcoal filtration, VCs from both fungal phytopathogens promoted growth and distinct developmental changes. Plants cultured at CO 2 levels observed in growth boxes containing fungal cultures were identical to those cultured at ambient CO 2 . Plants exposed to charcoal-filtered fungal VCs, nonfiltered VCs, or superelevated CO 2 levels exhibited transcriptional changes resembling those induced by increased irradiance. Thus, in the "box-in-box" system, (a) fungal VICs other than CO 2 and/or VOCs not detected by our analytical systems strongly influence the plants' responses to fungal VCs, (b) different microorganisms release VCs with distinct action potentials, (c) transcriptional changes in VC-exposed plants are mainly due to enhanced photosynthesis signaling, and (d) regulation of some plant responses to fungal VCs is primarily posttranscriptional., (© 2018 John Wiley & Sons Ltd.)

Zea mays Brittle1-1 (ZmBT1-1) is an essential component of the starch biosynthetic machinery in maize endosperms, enabling ADPglucose transport from cytosol to amyloplast in exchange for AMP or ADP. Although ZmBT1-1 has been long considered to be an amyloplast-specific marker, evidence has been provided that ZmBT1-1 is dually localized to plastids and mitochondria (Bahaji et al., 2011b). The mitochondrial localization of ZmBT1-1 suggested that this protein may have as-yet unidentified function(s). To understand the mitochondrial ZmBT1-1 function(s), we produced and characterized transgenic Zmbt1-1 plants expressing ZmBT1-1 delivered specifically to mitochondria. Metabolic and differential proteomic analyses showed down-regulation of sucrose synthase (SuSy)-mediated channeling of sucrose into starch metabolism, and up-regulation of the conversion of sucrose breakdown products generated by cell wall invertase (CWI) into ethanol and alanine, in Zmbt1-1 endosperms compared to wild-type. Electron microscopic analyses of Zmbt1-1 endosperm cells showed gross alterations in the mitochondrial ultrastructure. Notably, the protein expression pattern, metabolic profile, and aberrant mitochondrial ultrastructure of Zmbt1-1 endosperms were rescued by delivering ZmBT1-1 specifically to mitochondria. Results presented here provide evidence that the reduced starch content in Zmbt1-1 endosperms is at least partly due to (i) mitochondrial dysfunction, (ii) enhanced CWI-mediated channeling of sucrose into ethanol and alanine metabolism, and (iii) reduced SuSy-mediated channeling of sucrose into starch metabolism due to the lack of mitochondrial ZmBT1-1. Our results also strongly indicate that (a) mitochondrial ZmBT1-1 is an important determinant of the metabolic fate of sucrose entering the endosperm cells, and (b) plastidic ZmBT1-1 is not the sole ADPglucose transporter in maize endosperm amyloplasts. The possible involvement of mitochondrial ZmBT1-1 in exchange between intramitochondrial AMP and cytosolic ADP is discussed.

ADP-glucose is the precursor of glycogen biosynthesis in bacteria, and a compound abundant in the starchy plant organs ingested by many mammals. Here we show that the enteric species Escherichia coli is capable of scavenging exogenous ADP-glucose for use as a glycosyl donor in glycogen biosynthesis and feed the adenine nucleotide pool. To unravel the molecular mechanisms involved in this process, we screened the E. coli single-gene deletion mutants of the Keio collection for glycogen content in ADP-glucose-containing culture medium. In comparison to wild-type (WT) cells, individual ∆nupC and ∆nupG mutants lacking the cAMP/CRP responsive inner-membrane nucleoside transporters NupC and NupG displayed reduced glycogen contents and slow ADP-glucose incorporation. In concordance, ∆cya and ∆crp mutants accumulated low levels of glycogen and slowly incorporated ADP-glucose. Two-thirds of the glycogen-excess mutants identified during screening lacked functions that underlie envelope biogenesis and integrity, including the RpoE specific RseA anti-sigma factor. These mutants exhibited higher ADP-glucose uptake than WT cells. The incorporation of either ∆crp, ∆nupG or ∆nupC null alleles sharply reduced the ADP-glucose incorporation and glycogen content initially witnessed in ∆rseA cells. Overall, the data showed that E. coli incorporates extracellular ADP-glucose through a cAMP/CRP-regulated process involving the NupC and NupG nucleoside transporters that is facilitated under envelope stress conditions.

The plastid-localized phosphoglucose isomerase isoform PGI1 is an important determinant of growth in Arabidopsis thaliana , likely due to its involvement in the biosynthesis of plastidial isoprenoid-derived hormones. Here, we investigated whether PGI1 also influences seed yields. PGI1 is strongly expressed in maturing seed embryos and vascular tissues. PGI1- null pgi1-2 plants had ∼60% lower seed yields than wild-type plants, with reduced numbers of inflorescences and thus fewer siliques and seeds per plant. These traits were associated with low bioactive gibberellin (GA) contents. Accordingly, wild-type phenotypes were restored by exogenous GA application. pgi1-2 seeds were lighter and accumulated ∼50% less fatty acids (FAs) and ∼35% less protein than wild-type seeds. Seeds of cytokinin-deficient plants overexpressing CYTOKININ OXIDASE/DEHYDROGENASE1 (35S:AtCKX1) and GA-deficient ga20ox1 ga20ox2 mutants did not accumulate low levels of FAs, and exogenous application of the cytokinin 6-benzylaminopurine and GAs did not rescue the reduced weight and FA content of pgi1-2 seeds. Seeds from reciprocal crosses between pgi1-2 and wild-type plants accumulated wild-type levels of FAs and proteins. Therefore, PGI1 is an important determinant of Arabidopsis seed yield due to its involvement in two processes: GA-mediated reproductive development and the metabolic conversion of plastidial glucose-6-phosphate to storage reserves in the embryo., (© 2018 American Society of Plant Biologists. All rights reserved.)

Airway infection by nontypeable Haemophilus influenzae (NTHi) associates to chronic obstructive pulmonary disease (COPD) exacerbation and asthma neutrophilic airway inflammation. Lipids are key inflammatory mediators in these disease conditions and consequently, NTHi may encounter free fatty acids during airway persistence. However, molecular information on the interplay NTHi-free fatty acids is limited, and we lack evidence on the importance of such interaction to infection. Maintenance of the outer membrane lipid asymmetry may play an essential role in NTHi barrier function and interaction with hydrophobic molecules. VacJ/MlaA-MlaBCDEF prevents phospholipid accumulation at the bacterial surface, being the only system involved in maintaining membrane asymmetry identified in NTHi. We assessed the relationship among the NTHi VacJ/MlaA outer membrane lipoprotein, bacterial and exogenous fatty acids, and respiratory infection. The vacJ/mlaA gene inactivation increased NTHi fatty acid and phospholipid global content and fatty acyl specific species, which in turn increased bacterial susceptibility to hydrophobic antimicrobials, decreased NTHi epithelial infection, and increased clearance during pulmonary infection in mice with both normal lung function and emphysema, maybe related to their shared lung fatty acid profiles. Altogether, we provide evidence for VacJ/MlaA as a key bacterial factor modulating NTHi survival at the human airway upon exposure to hydrophobic molecules.

Although there is a great wealth of data supporting the occurrence of simultaneous synthesis and breakdown of storage carbohydrate in many organisms, previous 13CO2 pulse-chase based studies indicated that starch degradation does not operate in illuminated Arabidopsis leaves. Here we show that leaves of gwd, sex4, bam4, bam1/bam3 and amy3/isa3/lda starch breakdown mutants accumulate higher levels of starch than wild type (WT) leaves when cultured under continuous light (CL) conditions. We also show that leaves of CL grown dpe1 plants impaired in the plastidic disproportionating enzyme accumulate higher levels of maltotriose than WT leaves, the overall data providing evidence for the occurrence of extensive starch degradation in illuminated leaves. Moreover, we show that leaves of CL grown mex1/pglct plants impaired in the chloroplastic maltose and glucose transporters display a severe dwarf phenotype and accumulate high levels of maltose, strongly indicating that the MEX1 and pGlcT transporters are involved in the export of starch breakdown products to the cytosol to support growth during illumination. To investigate whether starch breakdown products can be recycled back to starch during illumination through a mechanism involving ADP-glucose pyrophosphorylase (AGP) we conducted kinetic analyses of the stable isotope carbon composition (δ13C) in starch of leaves of 13CO2 pulsed-chased WT and AGP lacking aps1 plants. Notably, the rate of increase of δ13C in starch of aps1 leaves during the pulse was exceedingly higher than that of WT leaves. Furthermore, δ13C decline in starch of aps1 leaves during the chase was much faster than that of WT leaves, which provides strong evidence for the occurrence of AGP-mediated cycling of starch breakdown products in illuminated Arabidopsis leaves., Competing Interests: The authors have declared that no competing interests exist.

It is known that volatile emissions from some beneficial rhizosphere microorganisms promote plant growth. Here we show that volatile compounds (VCs) emitted by phylogenetically diverse rhizosphere and non-rhizhosphere bacteria and fungi (including plant pathogens and microbes that do not normally interact mutualistically with plants) promote growth and flowering of various plant species, including crops. In Arabidopsis plants exposed to VCs emitted by the phytopathogen Alternaria alternata, changes included enhancement of photosynthesis and accumulation of high levels of cytokinins (CKs) and sugars. Evidence obtained using transgenic Arabidopsis plants with altered CK status show that CKs play essential roles in this phenomenon, because growth and flowering responses to the VCs were reduced in mutants with CK-deficiency (35S:AtCKX1) or low receptor sensitivity (ahk2/3). Further, we demonstrate that the plant responses to fungal VCs are light-dependent. Transcriptomic analyses of Arabidopsis leaves exposed to A. alternata VCs revealed changes in the expression of light- and CK-responsive genes involved in photosynthesis, growth and flowering. Notably, many genes differentially expressed in plants treated with fungal VCs were also differentially expressed in plants exposed to VCs emitted by the plant growth promoting rhizobacterium Bacillus subtilis GB03, suggesting that plants react to microbial VCs through highly conserved regulatory mechanisms., (© 2016 John Wiley & Sons Ltd.)

Volatile compounds (VCs) emitted by phylogenetically diverse microorganisms (including plant pathogens and microbes that do not normally interact mutualistically with plants) promote photosynthesis, growth, and the accumulation of high levels of starch in leaves through cytokinin (CK)-regulated processes. In Arabidopsis (Arabidopsis thaliana) plants not exposed to VCs, plastidic phosphoglucose isomerase (pPGI) acts as an important determinant of photosynthesis and growth, likely as a consequence of its involvement in the synthesis of plastidic CKs in roots. Moreover, this enzyme plays an important role in connecting the Calvin-Benson cycle with the starch biosynthetic pathway in leaves. To elucidate the mechanisms involved in the responses of plants to microbial VCs and to investigate the extent of pPGI involvement, we characterized pPGI-null pgi1-2 Arabidopsis plants cultured in the presence or absence of VCs emitted by Alternaria alternata We found that volatile emissions from this fungal phytopathogen promote growth, photosynthesis, and the accumulation of plastidic CKs in pgi1-2 leaves. Notably, the mesophyll cells of pgi1-2 leaves accumulated exceptionally high levels of starch following VC exposure. Proteomic analyses revealed that VCs promote global changes in the expression of proteins involved in photosynthesis, starch metabolism, and growth that can account for the observed responses in pgi1-2 plants. The overall data show that Arabidopsis plants can respond to VCs emitted by phytopathogenic microorganisms by triggering pPGI-independent mechanisms., (© 2016 American Society of Plant Biologists. All Rights Reserved.)

We characterized multiple knock-out mutants of the four Arabidopsis sucrose phosphate synthase (SPSA1, SPSA2, SPSB and SPSC) isoforms. Despite their reduced SPS activity, spsa1/spsa2, spsa1/spsb, spsa2/spsb, spsa2/spsc, spsb/spsc, spsa1/spsa2/spsb and spsa2/spsb/spsc mutants displayed wild type (WT) vegetative and reproductive morphology, and showed WT photosynthetic capacity and respiration. In contrast, growth of rosettes, flowers and siliques of the spsa1/spsc and spsa1/spsa2/spsc mutants was reduced compared with WT plants. Furthermore, these plants displayed a high dark respiration phenotype. spsa1/spsb/spsc and spsa1/spsa2/spsb/spsc seeds poorly germinated and produced aberrant and sterile plants. Leaves of all viable sps mutants, except spsa1/spsc and spsa1/spsa2/spsc, accumulated WT levels of nonstructural carbohydrates. spsa1/spsc leaves possessed high levels of metabolic intermediates and activities of enzymes of the glycolytic and tricarboxylic acid cycle pathways, and accumulated high levels of metabolic intermediates of the nocturnal starch-to-sucrose conversion process, even under continuous light conditions. Results presented in this work show that SPS is essential for plant viability, reveal redundant functions of the four SPS isoforms in processes that are important for plant growth and nonstructural carbohydrate metabolism, and strongly indicate that accelerated starch turnover and enhanced respiration can alleviate the blockage of sucrose biosynthesis in spsa1/spsc leaves., (Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.)

Phosphoglucose isomerase (PGI) catalyzes the reversible isomerization of glucose-6-phosphate and fructose-6-phosphate. It is involved in glycolysis and in the regeneration of glucose-6-P molecules in the oxidative pentose phosphate pathway (OPPP). In chloroplasts of illuminated mesophyll cells PGI also connects the Calvin-Benson cycle with the starch biosynthetic pathway. In this work we isolated pgi1-3, a mutant totally lacking pPGI activity as a consequence of aberrant intron splicing of the pPGI encoding gene, PGI1. Starch content in pgi1-3 source leaves was ca. 10-15% of that of wild type (WT) leaves, which was similar to that of leaves of pgi1-2, a T-DNA insertion pPGI null mutant. Starch deficiency of pgi1 leaves could be reverted by the introduction of a sex1 null mutation impeding β-amylolytic starch breakdown. Although previous studies showed that starch granules of pgi1-2 leaves are restricted to both bundle sheath cells adjacent to the mesophyll and stomata guard cells, microscopy analyses carried out in this work revealed the presence of starch granules in the chloroplasts of pgi1-2 and pgi1-3 mesophyll cells. RT-PCR analyses showed high expression levels of plastidic and extra-plastidic β-amylase encoding genes in pgi1 leaves, which was accompanied by increased β-amylase activity. Both pgi1-2 and pgi1-3 mutants displayed slow growth and reduced photosynthetic capacity phenotypes even under continuous light conditions. Metabolic analyses revealed that the adenylate energy charge and the NAD(P)H/NAD(P) ratios in pgi1 leaves were lower than those of WT leaves. These analyses also revealed that the content of plastidic 2-C-methyl-D-erythritol 4-phosphate (MEP)-pathway derived cytokinins (CKs) in pgi1 leaves were exceedingly lower than in WT leaves. Noteworthy, exogenous application of CKs largely reverted the low starch content phenotype of pgi1 leaves. The overall data show that pPGI is an important determinant of photosynthesis, energy status, growth and starch accumulation in mesophyll cells likely as a consequence of its involvement in the production of OPPP/glycolysis intermediates necessary for the synthesis of plastidic MEP-pathway derived hormones such as CKs.

In Escherichia coli, ppGpp is a major determinant of growth and glycogen accumulation. Levels of this signaling nucleotide are controlled by the balanced activities of the ppGpp RelA synthetase and the dual-function hydrolase/synthetase SpoT. Here we report the construction of spoT null (ΔspoT) mutants obtained by transducing a ΔspoT allele from ΔrelAΔspoT double mutants into relA+ cells. Iodine staining of randomly selected transductants cultured on a rich complex medium revealed differences in glycogen content among them. Sequence and biochemical analyses of 8 ΔspoT clones displaying glycogen-deficient phenotypes revealed different inactivating mutations in relA and no detectable ppGpp when cells were cultured on a rich complex medium. Remarkably, although the co-existence of ΔspoT with relA proficient alleles has generally been considered synthetically lethal, we found that 11 ΔspoT clones displaying high glycogen phenotypes possessed relA mutant alleles with non-inactivating mutations that encoded stable RelA proteins and ppGpp contents reaching 45-85% of those of wild type cells. None of the ΔspoT clones, however, could grow on M9-glucose minimal medium. Both Sanger sequencing of specific genes and high-throughput genome sequencing of the ΔspoT clones revealed that suppressor mutations were restricted to the relA locus. The overall results (a) defined in around 4 nmoles ppGpp/g dry weight the threshold cellular levels that suffice to trigger net glycogen accumulation, (b) showed that mutations in relA, but not necessarily inactivating mutations, can be selected to compensate total SpoT function(s) loss, and (c) provided useful tools for studies of the in vivo regulation of E. coli RelA ppGpp synthetase.

In leaves, it is widely assumed that starch is the end-product of a metabolic pathway exclusively taking place in the chloroplast that (a) involves plastidic phosphoglucomutase (pPGM), ADPglucose (ADPG) pyrophosphorylase (AGP) and starch synthase (SS), and (b) is linked to the Calvin-Benson cycle by means of the plastidic phosphoglucose isomerase (pPGI). This view also implies that AGP is the sole enzyme producing the starch precursor molecule, ADPG. However, mounting evidence has been compiled pointing to the occurrence of important sources, other than the pPGI-pPGM-AGP pathway, of ADPG. To further explore this possibility, in this work two independent laboratories have carried out HPLC-MS/MS analyses of ADPG content in leaves of the near-starchless pgm and aps1 mutants impaired in pPGM and AGP, respectively, and in leaves of double aps1/pgm mutants grown under two different culture conditions. We also measured the ADPG content in wild type (WT) and aps1 leaves expressing in the plastid two different ADPG cleaving enzymes, and in aps1 leaves expressing in the plastid GlgC, a bacterial AGP. Furthermore, we measured the ADPG content in ss3/ss4/aps1 mutants impaired in starch granule initiation and chloroplastic ADPG synthesis. We found that, irrespective of their starch contents, pgm and aps1 leaves, WT and aps1 leaves expressing in the plastid ADPG cleaving enzymes, and aps1 leaves expressing in the plastid GlgC accumulate WT ADPG content. In clear contrast, ss3/ss4/aps1 leaves accumulated ca. 300 fold-more ADPG than WT leaves. The overall data showed that, in Arabidopsis leaves, (a) there are important ADPG biosynthetic pathways, other than the pPGI-pPGM-AGP pathway, (b) pPGM and AGP are not major determinants of intracellular ADPG content, and (c) the contribution of the chloroplastic ADPG pool to the total ADPG pool is low.

Structurally composed of the glucose homopolymers amylose and amylopectin, starch is the main storage carbohydrate in vascular plants, and is synthesized in the plastids of both photosynthetic and non-photosynthetic cells. Its abundance as a naturally occurring organic compound is surpassed only by cellulose, and represents both a cornerstone for human and animal nutrition and a feedstock for many non-food industrial applications including production of adhesives, biodegradable materials, and first-generation bioethanol. This review provides an update on the different proposed pathways of starch biosynthesis occurring in both autotrophic and heterotrophic organs, and provides emerging information about the networks regulating them and their interactions with the environment. Special emphasis is given to recent findings showing that volatile compounds emitted by microorganisms promote both growth and the accumulation of exceptionally high levels of starch in mono- and dicotyledonous plants. We also review how plant biotechnologists have attempted to use basic knowledge on starch metabolism for the rational design of genetic engineering traits aimed at increasing starch in annual crop species. Finally we present some potential biotechnological strategies for enhancing starch content., (© 2013 Elsevier Inc. All rights reserved.)

Escherichia coli glycogen metabolism involves the regulation of glgBXCAP operon expression and allosteric control of the GlgC [ADPG (ADP-glucose) pyrophosphorylase]-mediated catalysis of ATP and G1P (glucose-1-phosphate) to ADPG linked to glycogen biosynthesis. E. coli glycogen metabolism is also affected by glgS. Though the precise function of the protein it encodes is unknown, its deficiency causes both reduced glycogen content and enhanced levels of the GlgC-negative allosteric regulator AMP. The transcriptomic analyses carried out in the present study revealed that, compared with their isogenic BW25113 wild-type strain, glgS-null (ΔglgS) mutants have increased expression of the operons involved in the synthesis of type 1 fimbriae adhesins, flagella and nucleotides. In agreement, ΔglgS cells were hyperflagellated and hyperfimbriated, and displayed elevated swarming motility; these phenotypes all reverted to the wild-type by ectopic glgS expression. Also, ΔglgS cells accumulated high colanic acid content and displayed increased ability to form biofilms on polystyrene surfaces. F-driven conjugation based on large-scale interaction studies of glgS with all the non-essential genes of E. coli showed that deletion of purine biosynthesis genes complement the glycogen-deficient, high motility and high biofilm content phenotypes of ΔglgS cells. Overall the results of the present study indicate that glycogen deficiency in ΔglgS cells can be ascribed to high flagellar propulsion and high exopolysaccharide and purine nucleotides biosynthetic activities competing with GlgC for the same ATP and G1P pools. Supporting this proposal, glycogen-less ΔglgC cells displayed an elevated swarming motility, and accumulated high levels of colanic acid and biofilm. Furthermore, glgC overexpression reverted the glycogen-deficient, high swarming motility, high colanic acid and high biofilm content phenotypes of ΔglgS cells to the wild-type. As on the basis of the present study GlgS has emerged as a major determinant of E. coli surface composition and because its effect on glycogen metabolism appears to be only indirect, we propose to rename it as ScoR (surface composition regulator).

Sucrose synthase (SuSy) is a highly regulated cytosolic enzyme that catalyzes the conversion of sucrose and a nucleoside diphosphate into the corresponding nucleoside diphosphate glucose and fructose. In cereal endosperms, it is widely assumed that the stepwise reactions of SuSy, UDPglucose pyrophosphorylase and ADPglucose (ADPG) pyrophosphorylase (AGP) take place in the cytosol to convert sucrose into ADPG necessary for starch biosynthesis, although it has also been suggested that SuSy may participate in the direct conversion of sucrose into ADPG. In this study, the levels of the major primary carbon metabolites, and the activities of starch metabolism-related enzymes were assessed in endosperms of transgenic maize plants ectopically expressing StSUS4, which encodes a potato SuSy isoform. A total of 29 fertile lines transformed with StSUS4 were obtained, five of them containing a single copy of the transgene that was still functional after five generations. The number of seeds per ear of the five transgenic lines containing a single StSUS4 copy was comparable with that of wild-type (WT) control seeds. However, transgenic seeds accumulated 10-15% more starch at the mature stage, and contained a higher amylose/amylopectin balance than WT seeds. Endosperms of developing StSUS4-expressing seeds exhibited a significant increase in SuSy activity, and in starch and ADPG contents when compared with WT endosperms. No significant changes could be detected in the transgenic seeds in the content of soluble sugars, and in activities of starch metabolism-related enzymes when compared with WT seeds. A suggested metabolic model is presented wherein both AGP and SuSy are involved in the production of ADPG linked to starch biosynthesis in maize endosperm cells.

Synthesized by glycogen synthase and starch synthases (SS) using ADP-glucose as the sugar donor molecule, glycogen and starch accumulate as predominant storage carbohydrates in most bacteria and plants, respectively. We have recently shown that the so-called "starch-less" Arabidopsis thaliana adg1-1 and aps1 mutants impaired in ADP-glucose pyrophosphorylase do indeed accumulate low starch content in normal growth conditions, and relatively high starch content when plants were cultured in the presence of microbial volatiles. Our results were strongly supported by data obtained using a highly sensitive method for confocal fluorescence microscopic visualization of iodine stained starch granules. Using Arabidopsis leaves from WT plants, aps1 plants, ss3/ss4 plants lacking both class III and class IV SS, gbss plants lacking the granule-bound SS, and sus1/sus2/sus3/sus4 plants lacking four genes that code for proteins with sucrose synthase activity, in this work we precisely describe the method for preparation of plant samples for starch microscopic examination. Furthermore, we show that this method can be used to visualize glycogen in bacteria, and pure starch granules, amylose and amylopectin.

Sucrose synthase (SuSy) catalyzes the reversible conversion of sucrose and NDP into the corresponding nucleotide-sugars and fructose. The Arabidopsis genome possesses six SUS genes (AtSUS1-6) that code for proteins with SuSy activity. As a first step to investigate optimum fructose and UDP-glucose (UDPG) concentrations necessary to measure maximum sucrose-producing SuSy activity in crude extracts of Arabidopsis, in this work we performed kinetic analyses of recombinant AtSUS1 in two steps: (1) SuSy reaction at pH 7.5, and (2) chromatographic measurement of sucrose produced in step 1. These analyses revealed a typical Michaelis-Menten behavior with respect to both UDPG and fructose, with Km values of 50 μM and 25 mM, respectively. Unlike earlier studies showing the occurrence of substrate inhibition of UDP-producing AtSUS1 by fructose and UDP-glucose, these analyses also revealed no substrate inhibition of AtSUS1 at any UDPG and fructose concentration. By including 200 mM fructose and 1 mM UDPG in the SuSy reaction assay mixture, we found that sucrose-producing SuSy activity in leaves and stems of Arabidopsis were exceedingly higher than previously reported activities. Furthermore, we found that SuSy activities in organs of the sus1/sus2/sus3/sus4 mutant were ca. 80-90% of those found in WT plants.

ADP-glucose pyrophosphorylase (AGP) is a heterotetrameric enzyme comprising two small and two large subunits that catalyze the production of ADP-glucose linked to starch biosynthesis. The current paradigm on leaf starch metabolism assumes that post-translational redox modification of AGP in response to light is a major determinant of fine regulation of transitory starch accumulation. According to this view, under oxidizing conditions occurring during the night the two AGP small subunits (APS1) are covalently linked via an intermolecular disulfide bridge that inactivates the protein, whereas under reducing conditions occurring during the day NADP-thioredoxin reductase C (NTRC)-dependent reductive monomerization of APS1 activates the enzyme. In this work we have analyzed changes in the redox status of APS1 during dark-light transition in leaves of plants cultured under different light intensities. Furthermore, we have carried out time-course analyses of starch content in ntrc mutants, and in aps1 mutants expressing the Escherichia coli redox-insensitive AGP (GlgC) in the chloroplast. We also characterized aps1 plants expressing a redox-insensitive, mutated APS1 (APS1mut) form in which the highly conserved Cys81 residue involved in the formation of the intermolecular disulfide bridge has been replaced by serine. We found that a very moderate, NTRC-dependent APS1 monomerization process in response to light occurred only when plants were cultured under photo-oxidative conditions. We also found that starch accumulation rates during the light in leaves of both ntrc mutants and GlgC-expressing aps1 mutants were similar to those of wild-type leaves. Furthermore, the pattern of starch accumulation during illumination in leaves of APS1mut-expressing aps1 mutants was similar to that of APS1-expressing aps1 mutants at any light intensity. The overall data demonstrate that post-translational redox modification of AGP in response to light is not a major determinant of fine regulation of transitory starch accumulation in Arabidopsis.

Sucrose synthase (SUS) catalyzes the reversible conversion of sucrose and a nucleoside diphosphate into the corresponding nucleoside diphosphate-glucose and fructose. In Arabidopsis, a multigene family encodes six SUS (SUS1-6) isoforms. The involvement of SUS in the synthesis of UDP-glucose and ADP-glucose linked to Arabidopsis cellulose and starch biosynthesis, respectively, has been questioned by Barratt et al. [(2009) Proc Natl Acad Sci USA 106:13124-13129], who showed that (i) SUS activity in wild type (WT) leaves is too low to account for normal rate of starch accumulation in Arabidopsis, and (ii) different organs of the sus1/sus2/sus3/sus4 SUS mutant impaired in SUS activity accumulate WT levels of ADP-glucose, UDP-glucose, cellulose and starch. However, these authors assayed SUS activity under unfavorable pH conditions for the reaction. By using favorable pH conditions for assaying SUS activity, in this work we show that SUS activity in the cleavage direction is sufficient to support normal rate of starch accumulation in WT leaves. We also demonstrate that sus1/sus2/sus3/sus4 leaves display WT SUS5 and SUS6 expression levels, whereas leaves of the sus5/sus6 mutant display WT SUS1-4 expression levels. Furthermore, we show that SUS activity in leaves and stems of the sus1/sus2/sus3/sus4 and sus5/sus6 plants is ∼85% of that of WT leaves, which can support normal cellulose and starch biosynthesis. The overall data disprove Barratt et al. (2009) claims, and are consistent with the possible involvement of SUS in cellulose and starch biosynthesis in Arabidopsis.

It has been shown that homozygous AtBT1::T-DNA Arabidopsis mutants display an aberrant growth and sterility phenotype, and that AtBT1 is a carrier that is exclusively localized to the inner plastidial envelope and is required for export of newly synthesized adenylates into the cytosol. However, a recent demonstration that AtBT1 is localized to both plastids and mitochondria suggested that plastidic AtBT1 is not necessary for normal growth and fertility of Arabidopsis. To test this hypothesis, we produced and characterized homozygous AtBT1::T-DNA mutants stably expressing either dually localized AtBT1 or AtBT1 specifically localized to the mitochondrial compartment. These analyses revealed that the aberrant growth and sterility phenotype of homozygous AtBT1::T-DNA mutants was complemented when expressing both the dual-targeted AtBT1 and AtBT1 specifically delivered to mitochondria. These data confirm that (i) plastidic AtBT1 is not strictly required for normal growth and fertility of the plant, and (ii) specific delivery of AtBT1 to mitochondria is enough to complement the aberrant growth and sterility phenotype of homozygous AtBT1::T-DNA mutants. Furthermore, data presented here question the idea that the requirement for AtBT1 is due to its involvement in transport of newly synthesized adenylates from the plastid to the cytosol, and suggest that the protein may play as yet unidentified functions in plastids and mitochondria., (© 2011 The Authors. The Plant Journal © 2011 Blackwell Publishing Ltd.)

Microbial volatiles promote the accumulation of exceptionally high levels of starch in leaves. Time-course analyses of starch accumulation in Arabidopsis leaves exposed to fungal volatiles (FV) emitted by Alternaria alternata revealed that a microbial volatile-induced starch accumulation process (MIVOISAP) is due to stimulation of starch biosynthesis during illumination. The increase of starch content in illuminated leaves of FV-treated hy1/cry1, hy1/cry2, and hy1/cry1/cry2 Arabidopsis mutants was many-fold lower than that of wild-type (WT) leaves, indicating that MIVOISAP is subjected to photoreceptor-mediated control. This phenomenon was inhibited by cordycepin and accompanied by drastic changes in the Arabidopsis transcriptome. MIVOISAP was also accompanied by enhancement of the total 3-phosphoglycerate/Pi ratio, and a two- to threefold increase of the levels of the reduced form of ADP-glucose pyrophosphorylase. Using different Arabidopsis knockout mutants, we investigated the impact in MIVOISAP of downregulation of genes directly or indirectly related to starch metabolism. These analyses revealed that the magnitude of the FV-induced starch accumulation was low in mutants impaired in starch synthase (SS) classes III and IV and plastidial NADP-thioredoxin reductase C (NTRC). Thus, the overall data showed that Arabidopsis MIVOISAP involves a photocontrolled, transcriptionally and post-translationally regulated network wherein photoreceptor-, SSIII-, SSIV-, and NTRC-mediated changes in redox status of plastidial enzymes play important roles.

It is widely considered that ADP-glucose pyrophosphorylase (AGP) is the sole source of ADP-glucose linked to bacterial glycogen and plant starch biosynthesis. Genetic evidence that bacterial glycogen biosynthesis occurs solely by the AGP pathway has been obtained with glgC⁻ AGP mutants. However, recent studies have shown that (i) these mutants can accumulate high levels of ADP-glucose and glycogen, and (ii) there are sources other than GlgC, of ADP-glucose linked to glycogen biosynthesis. In Arabidopsis, evidence showing that starch biosynthesis occurs solely by the AGP pathway has been obtained with the starchless adg1-1 and aps1 AGP mutants. However, mounting evidence has been compiled previewing the occurrence of more than one important ADP-glucose source in plants. In attempting to solve this 20-year-old controversy, in this work we carried out a judicious characterization of both adg1-1 and aps1. Both mutants accumulated wild-type (WT) ADP-glucose and approximately 2% of WT starch, as further confirmed by confocal fluorescence microscopic observation of iodine-stained leaves and of leaves expressing granule-bound starch synthase fused with GFP. Introduction of the sex1 mutation affecting starch breakdown into adg1-1 and aps1 increased the starch content to 8-10% of the WT starch. Furthermore, aps1 leaves exposed to microbial volatiles for 10 h accumulated approximately 60% of the WT starch. aps1 plants expressing the bacterial ADP-glucose hydrolase EcASPP in the plastid accumulated normal ADP-glucose and reduced starch when compared with aps1 plants, whereas aps1 plants expressing EcASPP in the cytosol showed reduced ADP-glucose and starch. Moreover, aps1 plants expressing bacterial AGP in the plastid accumulated WT starch and ADP-glucose. The overall data show that (i) there occur important source(s), other than AGP, of ADP-glucose linked to starch biosynthesis, and (ii) AGP is a major determinant of starch accumulation but not of intracellular ADP-glucose content in Arabidopsis.

Although it is generally accepted that Escherichia coli glycogen genes are organized in two tandemly arranged, differentially regulated glgBX and glgCAP operons, RT (reverse transcriptase)-PCR analyses carried out in the present study showed that E. coli cells possess transcripts comprising the five glgBXCAP genes. glg::lacZY expression analyses in cells lacking the region immediately upstream of the glgB gene revealed an almost total abolishment of glgB, glgX and glgC expression, but only a 50-60% reduction of the wild-type glgA and glgP expression levels. Furthermore, similar analyses showed that glgA and glgP expression was almost totally abolished in cells lacking glgA upstream sequences, including glgC, glgB and the asd-glgB intergenic region upstream of glgB. These results indicate that E. coli glgBXCAP genes are organized in a single transcriptional unit controlled by promoter sequences occurring upstream of glgB, and that an alternative suboperonic promoter is located within glgC, driving expression of the glgA and glgP genes. Computer searches for consensus promoters, and analyses of glgB::lacZY and glgA::lacZY expression in cells containing deletions of glgB and glgA upstream sequences identified regions directing glgBXCAP and glgAP expression. 5' RACE (rapid amplification of cDNA ends) analyses located a glgBXCAP transcription start site 155 bp upstream of the glgB initiation codon, and a glgAP transcription start site 359 bp upstream of the glgA initiation codon. Finally, glg::lacZY expression analyses on cells lacking the relA or phoP regulatory genes indicated that both the glgBXCAP operon and the suboperonic promoter driving glgAP expression form part of both the RelA and PhoP-PhoQ regulons.

Using the Keio collection of gene-disrupted mutants of Escherichia coli, we have recently carried out a genome-wide screening of the genes affecting glycogen metabolism. Among the mutants identified in the study, Delta mgtA, Delta phoP and Delta phoQ cells, all lacking genes that are induced under low extracellular Mg2+ conditions, displayed glycogen-deficient phenotypes. In this work we show that these mutants accumulated normal glycogen levels when the culture medium was supplemented with submillimolar Mg2+ concentrations. Expression analyses conducted in wild-type, Delta phoP and Delta phoQ cells showed that the glgCAP operon is under PhoP-PhoQ control in the submillimolar Mg2+ concentration range. Subsequent screening of the Keio collection under non-limiting Mg2+ allowed the identification of 183 knock-out mutants with altered glycogen levels. The stringent and general stress responses, end-turnover of tRNA, intracellular AMP levels, and metabolism of amino acids, iron, carbon and sulfur were major determinants of glycogen levels. glgC::lacZY expression analyses using mutants representing different functional categories revealed that the glgCAP operon belongs to the RelA regulon. We propose an integrated metabolic model wherein glycogen metabolism is (a) tightly controlled by the energy and nutritional status of the cell and (b) finely regulated by changes in environmental Mg2+ occurring at the submillimolar concentration range.