@article{MalinovaKoesslerOrawetzetal.2019, author = {Malinova, Irina and K{\"o}ssler, Stella and Orawetz, Tom and Matthes, Ulrike and Orzechowski, Slawomir and Koch, Anke and Fettke, J{\"o}rg}, title = {Identification of two Arabidopsis thaliana plasma membrane transporters able to transport glucose 1-phosphate}, series = {Plant \& cell physiology}, volume = {61}, journal = {Plant \& cell physiology}, number = {2}, publisher = {Oxford University Press}, address = {Oxford}, issn = {0032-0781}, doi = {10.1093/pcp/pcz206}, pages = {381 -- 392}, year = {2019}, abstract = {Primary carbohydrate metabolism in plants includes several sugar and sugar-derivative transport processes. Over recent years, evidences have shown that in starch-related transport processes, in addition to glucose 6-phosphate, maltose, glucose and triose-phosphates, glucose 1-phosphate also plays a role and thereby increases the possible fluxes of sugar metabolites in planta. In this study, we report the characterization of two highly similar transporters, At1g34020 and At4g09810, in Arabidopsis thaliana, which allow the import of glucose 1-phosphate through the plasma membrane. Both transporters were expressed in yeast and were biochemically analyzed to reveal an antiport of glucose 1-phosphate/phosphate. Furthermore, we showed that the apoplast of Arabidopsis leaves contained glucose 1-phosphate and that the corresponding mutant of these transporters had higher glucose 1-phosphate amounts in the apoplast and alterations in starch and starch-related metabolism.}, language = {en} } @article{MalinovaMahtoBrandtetal.2018, author = {Malinova, Irina and Mahto, Harendra and Brandt, Felix and AL-Rawi, Shadha and Qasim, Hadeel and Brust, Henrike and Hejazi, Mahdi and Fettke, J{\"o}rg}, title = {EARLY STARVATION1 specifically affects the phosphorylation action of starch-related dikinases}, series = {The plant journal}, volume = {95}, journal = {The plant journal}, number = {1}, publisher = {Wiley}, address = {Hoboken}, issn = {0960-7412}, doi = {10.1111/tpj.13937}, pages = {126 -- 137}, year = {2018}, abstract = {Starch phosphorylation by starch-related dikinases glucan, water dikinase (GWD) and phosphoglucan, water dikinase (PWD) is a key step in starch degradation. Little information is known about the precise structure of the glucan substrate utilized by the dikinases and about the mechanisms by which these structures may be influenced. A 50-kDa starch-binding protein named EARLY STARVATION1 (ESV1) was analyzed regarding its impact on starch phosphorylation. In various invitro assays, the influences of the recombinant protein ESV1 on the actions of GWD and PWD on the surfaces of native starch granules were analyzed. In addition, we included starches from various sources as well as truncated forms of GWD. ESV1 preferentially binds to highly ordered, -glucans, such as starch and crystalline maltodextrins. Furthermore, ESV1 specifically influences the action of GWD and PWD at the starch granule surface. Starch phosphorylation by GWD is decreased in the presence of ESV1, whereas the action of PWD increases in the presence of ESV1. The unique alterations observed in starch phosphorylation by the two dikinases are discussed in regard to altered glucan structures at the starch granule surface.}, language = {en} } @article{MalinovaAlseekhFeiletal.2017, author = {Malinova, Irina and Alseekh, Saleh and Feil, Regina and Fernie, Alisdair R. and Baumann, Otto and Schoettler, Mark Aurel and Lunn, John Edward and Fettke, J{\"o}rg}, title = {Starch Synthase 4 and Plastidal Phosphorylase Differentially Affect Starch Granule Number and Morphology}, series = {Plant physiology : an international journal devoted to physiology, biochemistry, cellular and molecular biology, biophysics and environmental biology of plants}, volume = {174}, journal = {Plant physiology : an international journal devoted to physiology, biochemistry, cellular and molecular biology, biophysics and environmental biology of plants}, publisher = {American Society of Plant Physiologists}, address = {Rockville}, issn = {0032-0889}, doi = {10.1104/pp.16.01859}, pages = {73 -- 85}, year = {2017}, abstract = {The process of starch granule formation in leaves of Arabidopsis ( Arabidopsis thaliana) is obscure. Besides STARCH SYNTHASE4 (SS4), the PLASTIDIAL PHOSPHORYLASE (PHS1) also seems to be involved, since dpe2-1/phs1a double mutants lacking both PHS1 and the cytosolic DISPROPORTIONATING ENZYME2 (DPE2) displayed only one starch granule per chloroplast under normal growth conditions. For further studies, a dpe2-1/phs1a/ss4 triple mutant and various combinations of double mutants were generated and metabolically analyzed with a focus on starch metabolism. The dpe2-1/phs1a/ ss4 mutant revealed a massive starch excess phenotype. Furthermore, these plants grown under 12 h of light/12 h of dark harbored a single large and spherical starch granule per plastid. The number of starch granules was constant when the light/dark regime was altered, but this was not observed in the parental lines. With regard to growth, photosynthetic parameters, and metabolic analyses, the triple mutant additionally displayed alterations in comparison with ss4 and dpe21/phs1a. The results clearly illustrate that PHS1 and SS4 are differently involved in starch granule formation and do not act in series. However, SS4 appears to exert a stronger influence. In connection with the characterized double mutants, we discuss the generation of starch granules and the observed formation of spherical starch granules.}, language = {en} } @article{MalinovaFettke2017, author = {Malinova, Irina and Fettke, J{\"o}rg}, title = {Reduced starch granule number per chloroplast in the dpe2/phs1 mutant is dependent on initiation of starch degradation}, series = {PLoS one}, volume = {12}, journal = {PLoS one}, publisher = {PLoS}, address = {San Fransisco}, issn = {1932-6203}, doi = {10.1371/journal.pone.0187985}, pages = {11}, year = {2017}, abstract = {An Arabidopsis double knock-out mutant lacking cytosolic disproportionating enzyme 2 (DPE2) and the plastidial phosphorylase (PHS1) revealed a dwarf-growth phenotype, reduced starch content, an uneven distribution of starch within the plant rosette, and a reduced number of starch granules per chloroplast under standard growth conditions. In contrast, the wild type contained 5-7 starch granules per chloroplast. Mature and old leaves of the double mutant were essentially starch free and showed plastidial disintegration. Several analyses revealed that the number of starch granules per chloroplast was affected by the dark phase. So far, it was unclear if it was the dark phase per se or starch degradation in the dark that was connected to the observed decrease in the number of starch granules per chloroplast. Therefore, in the background of the double mutant dpe2/phs1, a triple mutant was generated lacking the initial starch degrading enzyme glucan, water dikinase (GWD). The triple mutant showed improved plant growth, a starch-excess phenotype, and a homogeneous starch distribution. Furthermore, the number of starch granules per chloroplast was increased and was similar to wild type. However, starch granule morphology was only slightly affected by the lack of GWD as in the triple mutant and, like in dpe2/phs1, more spherical starch granules were observed. The characterized triple mutant was discussed in the context of the generation of starch granules and the formation of starch granule morphology.}, language = {en} } @article{OrawetzMalinovaOrzechowskietal.2016, author = {Orawetz, Tom and Malinova, Irina and Orzechowski, Slawomir and Fettke, J{\"o}rg}, title = {Reduction of the plastidial phosphorylase in potato (Solanum tuberosum L.) reveals impact on storage starch structure during growth at low temperature}, series = {Plant physiology and biochemistry : an official journal of the Federation of European Societies of Plant Physiology}, volume = {100}, journal = {Plant physiology and biochemistry : an official journal of the Federation of European Societies of Plant Physiology}, publisher = {Elsevier}, address = {Paris}, issn = {0981-9428}, doi = {10.1016/j.plaphy.2016.01.013}, pages = {141 -- 149}, year = {2016}, abstract = {Tubers of potato (Solanum tuberosum L.), one of the most important crops, are a prominent example for an efficient production of storage starch. Nevertheless, the synthesis of this storage starch is not completely understood. The plastidial phosphorylase (Phol; EC 2.4.11) catalyzes the reversible transfer of glucosyl residues from glucose-1-phosphate to the non-reducing end of alpha-glucans with the release of orthophosphate. Thus, the enzyme is in principle able to act during starch synthesis. However, so far under normal growth conditions no alterations in tuber starch metabolism were observed. Based on analyses of other species and also from in vitro experiments with potato tuber slices it was supposed, that Phol has a stronger impact on starch metabolism, when plants grow under low temperature conditions. Therefore, we analyzed the starch content, granule size, as well as the internal structure of starch granules isolated from potato plants grown under low temperatures. Besides wild type, transgenic potato plants with a strong reduction in the Phol activity were analyzed. No significant alterations in starch content and granule size were detected. In contrast, when plants were cultivated at low temperatures the chain length distributions of the starch granules were altered. Thus, the granules contained more short glucan chains. That was not observed in the transgenic plants, revealing that Pho1 in wild type is involved in the formation of the short glucan chains, at least at low temperatures. (C) 2016 Elsevier Masson SAS. All rights reserved.}, language = {en} } @article{MalinovaQasimBrustetal.2018, author = {Malinova, Irina and Qasim, Hadeel M. and Brust, Henrike and Fettke, J{\"o}rg}, title = {Parameters of Starch Granule Genesis in Chloroplasts of Arabidopsis thaliana}, series = {Frontiers in Plant Science}, volume = {9}, journal = {Frontiers in Plant Science}, publisher = {Frontiers Media}, address = {Lausanne}, issn = {1664-462X}, doi = {10.3389/fpls.2018.00761}, pages = {1 -- 7}, year = {2018}, abstract = {Starch is the primary storage carbohydrate in most photosynthetic organisms and allows the accumulation of carbon and energy in form of an insoluble and semi-crystalline particle. In the last decades large progress, especially in the model plant Arabidopsis thaliana, was made in understanding the structure and metabolism of starch and its conjunction. The process underlying the initiation of starch granules remains obscure, although this is a fundamental process and seems to be strongly regulated, as in Arabidopsis leaves the starch granule number per chloroplast is fixed with 5-7. Several single, double, and triple mutants were reported in the last years that showed massively alterations in the starch granule number per chloroplast and allowed further insights in this important process. This mini review provides an overview of the current knowledge of processes involved in the initiation and formation of starch granules. We discuss the central role of starch synthase 4 and further proteins for starch genesis and affecting metabolic factors.}, language = {en} } @article{MalinovaMahlowAlseekhetal.2014, author = {Malinova, Irina and Mahlow, Sebastian and Alseekh, Saleh and Orawetz, Tom and Fernie, Alisdair R. and Baumann, Otto and Steup, Martin and Fettke, J{\"o}rg}, title = {Double knockout mutants of arabidopsis grown under normal conditions reveal that the plastidial phosphorylase isozyme participates in transitory starch metabolism}, series = {Plant physiology : an international journal devoted to physiology, biochemistry, cellular and molecular biology, biophysics and environmental biology of plants}, volume = {164}, journal = {Plant physiology : an international journal devoted to physiology, biochemistry, cellular and molecular biology, biophysics and environmental biology of plants}, number = {2}, publisher = {American Society of Plant Physiologists}, address = {Rockville}, issn = {0032-0889}, doi = {10.1104/pp.113.227843}, pages = {907 -- 921}, year = {2014}, abstract = {In leaves of two starch-related single-knockout lines lacking either the cytosolic transglucosidase (also designated as disproportionating enzyme 2, DPE2) or the maltose transporter (MEX1), the activity of the plastidial phosphorylase isozyme (PHS1) is increased. In both mutants, metabolism of starch-derived maltose is impaired but inhibition is effective at different subcellular sites. Two constitutive double knockout mutants were generated (designated as dpe2-1 x phs1a and mex1 x phs1b) both lacking functional PHS1. They reveal that in normally grown plants, the plastidial phosphorylase isozyme participates in transitory starch degradation and that the central carbon metabolism is closely integrated into the entire cell biology. All plants were grown either under continuous illumination or in a light-dark regime. Both double mutants were compromised in growth and, compared with the single knockout plants, possess less average leaf starch when grown in a light-dark regime. Starch and chlorophyll contents decline with leaf age. As revealed by transmission electron microscopy, mesophyll cells degrade chloroplasts, but degradation is not observed in plants grown under continuous illumination. The two double mutants possess similar but not identical phenotypes. When grown in a light-dark regime, mesophyll chloroplasts of dpe2-1 x phs1a contain a single starch granule but under continuous illumination more granules per chloroplast are formed. The other double mutant synthesizes more granules under either growth condition. In continuous light, growth of both double mutants is similar to that of the parental single knockout lines. Metabolite profiles and oligoglucan patterns differ largely in the two double mutants.}, language = {en} } @article{KunzZamaniNourHaeusleretal.2014, author = {Kunz, Hans-Henning and Zamani-Nour, Shirin and Haeusler, Rainer E. and Ludewig, Katja and Schroeder, Julian I. and Malinova, Irina and Fettke, J{\"o}rg and Fluegge, Ulf-Ingo and Gierth, Markus}, title = {Loss of cytosolic phosphoglucose isomerase affects carbohydrate metabolism in leaves and is essential for fertility of arabidopsis}, series = {Plant physiology : an international journal devoted to physiology, biochemistry, cellular and molecular biology, biophysics and environmental biology of plants}, volume = {166}, journal = {Plant physiology : an international journal devoted to physiology, biochemistry, cellular and molecular biology, biophysics and environmental biology of plants}, number = {2}, publisher = {American Society of Plant Physiologists}, address = {Rockville}, issn = {0032-0889}, doi = {10.1104/pp.114.241091}, pages = {753 -- U960}, year = {2014}, abstract = {Carbohydrate metabolism in plants is tightly linked to photosynthesis and is essential for energy and carbon skeleton supply of the entire organism. Thus, the hexose phosphate pools of the cytosol and the chloroplast represent important metabolic resources that are maintained through action of phosphoglucose isomerase (PGI) and phosphoglucose mutase interconverting glucose 6-phosphate, fructose 6-phosphate, and glucose 1-phosphate. Here, we investigated the impact of disrupted cytosolic PGI (cPGI) function on plant viability and metabolism. Overexpressing an artificial microRNA targeted against cPGI (amiR-cpgi) resulted in adult plants with vegetative tissue essentially free of cPGI activity. These plants displayed diminished growth compared with the wild type and accumulated excess starch in chloroplasts but maintained low sucrose content in leaves at the end of the night. Moreover, amiR-cpgi plants exhibited increased nonphotochemical chlorophyll a quenching during photosynthesis. In contrast to amiR-cpgi plants, viable transfer DNA insertion mutants disrupted in cPGI function could only be identified as heterozygous individuals. However, homozygous transfer DNA insertion mutants could be isolated among plants ectopically expressing cPGI. Intriguingly, these plants were only fertile when expression was driven by the ubiquitin10 promoter but sterile when the seed-specific unknown seed protein promoter or the Cauliflower mosaic virus 35S promoter were employed. These data show that metabolism is apparently able to compensate for missing cPGI activity in adult amiR-cpgi plants and indicate an essential function for cPGI in plant reproduction. Moreover, our data suggest a feedback regulation in amiR-cpgi plants that fine-tunes cytosolic sucrose metabolism with plastidic starch turnover.}, language = {en} } @article{MalinovaKunzAlseekhetal.2014, author = {Malinova, Irina and Kunz, Hans-Henning and Alseekh, Saleh and Herbst, Karoline and Fernie, Alisdair R. and Gierth, Markus and Fettke, J{\"o}rg}, title = {Reduction of the cytosolic phosphoglucomutase in arabidopsis reveals impact on plant growth, seed and root development, and carbohydrate partitioning}, series = {PLoS one}, volume = {9}, journal = {PLoS one}, number = {11}, publisher = {PLoS}, address = {San Fransisco}, issn = {1932-6203}, doi = {10.1371/journal.pone.0112468}, pages = {11}, year = {2014}, abstract = {Phosphoglucomutase (PGM) catalyses the interconversion of glucose 1-phosphate (G1P) and glucose 6-phosphate (G6P) and exists as plastidial (pPGM) and cytosolic (cPGM) isoforms. The plastidial isoform is essential for transitory starch synthesis in chloroplasts of leaves, whereas the cytosolic counterpart is essential for glucose phosphate partitioning and, therefore, for syntheses of sucrose and cell wall components. In Arabidopsis two cytosolic isoforms (PGM2 and PGM3) exist. Both PGM2 and PGM3 are redundant in function as single mutants reveal only small or no alterations compared to wild type with respect to plant primary metabolism. So far, there are no reports of Arabidopsis plants lacking the entire cPGM or total PGM activity, respectively. Therefore, amiRNA transgenic plants were generated and used for analyses of various parameters such as growth, development, and starch metabolism. The lack of the entire cPGM activity resulted in a strongly reduced growth revealed by decreased rosette fresh weight, shorter roots, and reduced seed production compared to wild type. By contrast content of starch, sucrose, maltose and cell wall components were significantly increased. The lack of both cPGM and pPGM activities in Arabidopsis resulted in dwarf growth, prematurely die off, and inability to develop a functional inflorescence. The combined results are discussed in comparison to potato, the only described mutant with lack of total PGM activity.}, language = {en} } @article{FettkeMalinovaAlbrechtetal.2011, author = {Fettke, J{\"o}rg and Malinova, Irina and Albrecht, Tanja and Hejazi, Mahdi and Steup, Martin}, title = {Glucose-1-Phosphate transport into protoplasts and chloroplasts from leaves of arabidopsis}, series = {Plant physiology : an international journal devoted to physiology, biochemistry, cellular and molecular biology, biophysics and environmental biology of plants}, volume = {155}, journal = {Plant physiology : an international journal devoted to physiology, biochemistry, cellular and molecular biology, biophysics and environmental biology of plants}, number = {4}, publisher = {American Society of Plant Physiologists}, address = {Rockville}, issn = {0032-0889}, doi = {10.1104/pp.110.168716}, pages = {1723 -- 1734}, year = {2011}, abstract = {Almost all glucosyl transfer reactions rely on glucose-1-phosphate (Glc-1-P) that either immediately acts as glucosyl donor or as substrate for the synthesis of the more widely used Glc dinucleotides, ADPglucose or UDPglucose. In this communication, we have analyzed two Glc-1-P-related processes: the carbon flux from externally supplied Glc-1-P to starch by either mesophyll protoplasts or intact chloroplasts from Arabidopsis (Arabidopsis thaliana). When intact protoplasts or chloroplasts are incubated with [U-C-14]Glc-1-P, starch is rapidly labeled. Incorporation into starch is unaffected by the addition of unlabeled Glc-6-P or Glc, indicating a selective flux from Glc-1-P to starch. However, illuminated protoplasts incorporate less C-14 into starch when unlabeled bicarbonate is supplied in addition to the C-14-labeled Glc-1-P. Mesophyll protoplasts incubated with [U-C-14] Glc-1-P incorporate C-14 into the plastidial pool of adenosine diphosphoglucose. Protoplasts prepared from leaves of mutants of Arabidopsis that lack either the plastidial phosphorylase or the phosphoglucomutase isozyme incorporate C-14 derived from external Glc-1-P into starch, but incorporation into starch is insignificant when protoplasts from a mutant possessing a highly reduced ADPglucose pyrophosphorylase activity are studied. Thus, the path of assimilatory starch biosynthesis initiated by extraplastidial Glc-1-P leads to the plastidial pool of adenosine diphosphoglucose, and at this intermediate it is fused with the Calvin cycle-driven route. Mutants lacking the plastidial phosphoglucomutase contain a small yet significant amount of transitory starch.}, language = {en} }