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Samuel C Zeeman - One of the best experts on this subject based on the ideXlab platform.
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Starch synthase5 a noncanonical Starch synthase like protein promotes Starch Granule initiation in arabidopsis
The Plant Cell, 2020Co-Authors: Melanie R Abt, Barbara Pfister, Mayank Sharma, Simona Eicke, Leo Burgy, Isabel Neale, David Seung, Samuel C ZeemanAbstract:What determines the number of Starch Granules in plastids is an enigmatic aspect of Starch metabolism. Several structurally and functionally diverse proteins have been implicated in the Granule initiation process in Arabidopsis (Arabidopsis thaliana), with each protein exerting a varying degree of influence. Here, we show that a conserved Starch synthase-like protein, Starch SYNTHASE5 (SS5), regulates the number of Starch Granules that form in Arabidopsis chloroplasts. Among the Starch synthases, SS5 is most closely related to SS4, a major determinant of Granule initiation and morphology. However, unlike SS4 and the other Starch synthases, SS5 is a noncanonical isoform that lacks catalytic glycosyltransferase activity. Nevertheless, loss of SS5 reduces Starch Granule numbers that form per chloroplast in Arabidopsis, and ss5 mutant Starch Granules are larger than wild-type Granules. Like SS4, SS5 has a conserved putative surface binding site for glucans and also interacts with MYOSIN-RESEMBLING CHLOROPLAST PROTEIN, a proposed structural protein influential in Starch Granule initiation. Phenotypic analysis of a suite of double mutants lacking both SS5 and other proteins implicated in Starch Granule initiation allows us to propose how SS5 may act in this process.
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two plastidial coiled coil proteins are essential for normal Starch Granule initiation in arabidopsis
The Plant Cell, 2018Co-Authors: David Seung, Simona Eicke, Leo Burgy, Tina B Schreier, Samuel C ZeemanAbstract:The mechanism of Starch Granule initiation in chloroplasts is not fully understood. Here, we aimed to build on our recent discovery that PROTEIN TARGETING TO Starch (PTST) family members, PTST2 and PTST3, are key players in Starch Granule initiation, by identifying and characterizing additional proteins involved in the process in Arabidopsis thaliana chloroplasts. Using immunoprecipitation and mass spectrometry, we demonstrate that PTST2 interacts with two plastidial coiled-coil proteins. Surprisingly, one of the proteins is the thylakoid-associated MAR BINDING FILAMENT-LIKE PROTEIN1 (MFP1), which was proposed to bind plastid nucleoids. The other protein, MYOSIN-RESEMBLING CHLOROPLAST PROTEIN (MRC), contains long coiled coils and no known domains. Whereas wild-type chloroplasts contained multiple Starch Granules, only one large Granule was observed in most chloroplasts of the mfp1 and mrc mutants. The mfp1 mrc double mutant had a higher proportion of chloroplasts containing no visible Granule than either single mutant and accumulated ADP-glucose, the substrate for Starch synthesis. PTST2 was partially associated with the thylakoid membranes in wild-type plants, and fluorescently tagged PTST2 was located in numerous discrete patches within the chloroplast in which MFP1 was also located. In the mfp1 mutant, PTST2 was not associated with the thylakoids and formed discrete puncta, suggesting that MFP1 is necessary for normal PTST2 localization. Overall, we reveal that proper Granule initiation requires the presence of MFP1 and MRC, and the correct location of PTST2.
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degradation of glucan primers in the absence of Starch synthase 4 disrupts Starch Granule initiation in arabidopsis
Journal of Biological Chemistry, 2016Co-Authors: David Seung, Michaela Stettler, Sebastian Streb, Samuel C ZeemanAbstract:Arabidopsis leaf chloroplasts typically contain five to seven semicrystalline Starch Granules. It is not understood how the synthesis of each Granule is initiated or how Starch Granule number is determined within each chloroplast. An Arabidopsis mutant lacking the glucosyl-transferase, Starch SYNTHASE 4 (SS4) is impaired in its ability to initiate Starch Granules; its chloroplasts rarely contain more than one large Granule, and the plants have a pale appearance and reduced growth. Here we report that the chloroplastic α-amylase AMY3, a Starch-degrading enzyme, interferes with Granule initiation in the ss4 mutant background. The amy3 single mutant is similar in phenotype to the wild type under normal growth conditions, with comparable numbers of Starch Granules per chloroplast. Interestingly, the ss4 mutant displays a pleiotropic reduction in the activity of AMY3. Remarkably, complete abolition of AMY3 (in the amy3 ss4 double mutant) increases the number of Starch Granules produced in each chloroplast, suppresses the pale phenotype of ss4, and nearly restores normal growth. The amy3 mutation also restores Starch synthesis in the ss3 ss4 double mutant, which lacks Starch SYNTHASE 3 (SS3) in addition to SS4. The ss3 ss4 line is unable to initiate any Starch Granules and is thus Starchless. We suggest that SS4 plays a key role in Granule initiation, allowing it to proceed in a way that avoids premature degradation of primers by Starch hydrolases, such as AMY3.
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Starch synthase 4 is essential for coordination of Starch Granule formation with chloroplast division during arabidopsis leaf expansion
New Phytologist, 2013Co-Authors: Matilda Crumptontaylor, Christopher M Hylton, Regina Feil, John E Lunn, Simona Eicke, Samuel C Zeeman, Marilyn J Pike, Alison M. SmithAbstract:Arabidopsis thaliana mutants lacking the SS4 isoform of Starch synthase have strongly reduced numbers of Starch Granules per chloroplast, suggesting that SS4 is necessary for the normal generation of Starch Granules. To establish whether it plays a direct role in this process, we investigated the circumstances in which Granules are formed in ss4 mutants. Starch Granule numbers and distribution and the accumulation of Starch synthase substrates and products were investigated during ss4 leaf development, and in ss4 mutants carrying mutations or transgenes that affect Starch turnover or chloroplast volume. We found that immature ss4 leaves have no Starch Granules, but accumulate high concentrations of the Starch synthase substrate ADPglucose. Granule numbers are partially restored by elevating the capacity for glucan synthesis (via expression of bacterial glycogen synthase) or by increasing the volumes of individual chloroplasts (via introduction of arc mutations). However, these Granules are abnormal in distribution, size and shape. SS4 is an essential component of a mechanism that coordinates Granule formation with chloroplast division during leaf expansion and determines the abundance and the flattened, discoid shape of leaf Starch Granules. Keywords: ADPglucose, Arabidopsis thaliana, chloroplast, leaf expansion, Starch Granule, Starch synthase, Starch synthesis Introduction The process by which Starch Granules arise is not known. Suggestions range from largely physico-chemical mechanisms (Doi, 1965; Geddes & Greenwood, 1969; Ziegler et al., 2005) to the existence of specific protein primers analogous to the glycogenins of fungi and animals (e.g. Rothschild & Tandecarz, 1994; Singh et al., 1995; Langeveld et al., 2002; Chatterjee et al., 2005). Recent attention has focussed on the role of one isoform of soluble Starch synthase, Starch synthase 4 (SS4, At4 g18240). Although SS4 contributes little to total Starch synthase activity, ss4 mutants of Arabidopsis have at most one or two Starch Granules per chloroplast (Roldan et al., 2007) rather than the normal five or six (Crumpton-Taylor et al., 2012). No other Starch synthase is individually necessary for normal Granule numbers (Roldan et al., 2007), thus SS4 may have a specific function in Granule formation. However, other isoforms of Starch synthase may partially substitute for this function. The additional loss of SS3 further reduces Starch Granule numbers in the ss4 mutant background (Szydlowski et al., 2009; Merida & D′Hulst, 2012). The importance of SS4 for Starch Granule formation remains to be established. First, it is not known whether SS4 is required primarily for maintenance of Starch Granule numbers in mature leaves, or whether it also has a role in immature leaves where new Granules arise in concert with chloroplast division (Crumpton-Taylor et al., 2012). Second, it is not clear whether the reduction in Starch Granule numbers in ss4 mutants is a direct or an indirect consequence of the loss of SS4. Mutants have several additional phenotypes including reduced growth rates, altered Starch Granule anatomy and morphology and a reduction in the extent of diel Starch turnover (Roldan et al., 2007). It remains possible that the reduction in Granule numbers in ss4 mutants is an indirect consequence of one of these alterations. Third, a recent study suggests that SS4 may be limiting for Starch synthesis in wild-type plants. Its overexpression reportedly results in higher concentrations of Starch at the end of the day and accelerated plant growth (Gamez-Arjona et al., 2011). These results have important implications for the control of Starch turnover and are of biotechnological interest, but the relationship between Starch concentrations and Starch Granule numbers and sizes in plants with elevated SS4 was not reported. The aim of our work was to establish whether SS4 has a direct or an indirect role in Starch Granule formation, and to shed further light on where and when its actions are required for the establishment of normal Granule numbers. To this end we examined the phenotype of the ss4 mutant through leaf development, and investigated the impact of loss of SS4 in mutant and transgenic backgrounds in which Starch metabolism is altered or chloroplast volumes are abnormally large. Our results indicate that SS4 is directly and specifically required for the establishment of normal numbers and distributions of Starch Granules during leaf expansion, and that it is also necessary for the normal flattened, discoid shape of leaf Starch Granules.
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loss of Starch Granule initiation has a deleterious effect on the growth of arabidopsis plants due to an accumulation of adp glucose
Plant Physiology, 2013Co-Authors: Paula Ragel, Regina Feil, John E Lunn, Samuel C Zeeman, Sebastian Streb, Mariam Sahrawy, Maria Grazia Annunziata, Angel MeridaAbstract:Starch SYNTHASE4 (SS4) is required for proper Starch Granule initiation in Arabidopsis (Arabidopsis thaliana), although SS3 can partially replace its function. Unlike other Starch-deficient mutants, ss4 and ss3/ss4 mutants grow poorly even under long-day conditions. They have less chlorophyll and carotenoids than the wild type and lower maximal rates of photosynthesis. There is evidence of photooxidative damage of the photosynthetic apparatus in the mutants from chlorophyll a fluorescence parameters and their high levels of malondialdehyde. Metabolite profiling revealed that ss3/ss4 accumulates over 170 times more ADP-glucose (Glc) than wild-type plants. Restricting ADP-Glc synthesis, by introducing mutations in the plastidial phosphoglucomutase (pgm1) or the small subunit of ADP-Glc pyrophosphorylase (aps1), largely restored photosynthetic capacity and growth in pgm1/ss3/ss4 and aps1/ss3/ss4 triple mutants. It is proposed that the accumulation of ADP-Glc in the ss3/ss4 mutant sequesters a large part of the plastidial pools of adenine nucleotides, which limits photophosphorylation, leading to photooxidative stress, causing the chlorotic and stunted growth phenotypes of the plants.
Bicheng Wu - One of the best experts on this subject based on the ideXlab platform.
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structural design approaches for creating fat droplet and Starch Granule mimetics
Food & Function, 2017Co-Authors: David Julian Mcclements, Cheryl Chung, Bicheng WuAbstract:This article focuses on hydrogel-based strategies for creating reduced calorie foods with desirable physicochemical, sensory, and nutritional properties. Initially, the role of fat droplets and Starch Granules in foods is discussed, and then different methods for fabricating hydrogel beads are reviewed, including phase separation, antisolvent precipitation, injection, and emulsion template methods. Finally, the potential application of hydrogel beads as fat droplet and Starch Granule replacements is discussed. There is still a need for large-scale, high-throughout, and economical methods of fabricating hydrogel beads suitable for utilization within the food industry.
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microstructure rheology of mixed colloidal dispersions influence of ph induced droplet aggregation on Starch Granule fat droplet mixtures
Journal of Food Engineering, 2013Co-Authors: Bicheng Wu, Brian Degner, David Julian McclementsAbstract:Abstract The creation of high quality reduced-fat food products is challenging because the removal of fat adversely affects quality attributes, such as appearance, texture, and flavor. This study investigated the impact of pH-induced droplet aggregation on the properties of model food systems consisting of fat droplets and Starch Granules. Oil-in-water emulsions (2 wt.% oil) containing whey-protein coated lipid droplets aggregated extensively when heated (90 °C, 5 min) at pH values around their isoelectric point (pH 5) but not at lower (pH 3.5) or higher (pH 7) values, which was attributed to changes in electrostatic repulsion. The physicochemical properties of mixed lipid droplet–Starch dispersions (2 wt.% oil, 4 wt.% Starch) prepared under similar conditions (pH 3.5, 5, and 7; 90 °C for 5 min) were also measured. At pH 5, extensive lipid droplet aggregation was observed in mixed systems, which led to a large increase in their yield stress and apparent viscosity when compared to mixed systems at pH 3.5 and 7. These results show that the rheological properties of mixed fat droplet–Starch Granule suspensions can be modulated by controlling the electrostatic interactions between the fat droplets so as to change their flocculation state. This study has important implications for fabricating high quality reduced-fat products with desirable sensory attributes.
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Microstructure & rheology of mixed colloidal dispersions: Influence of pH-induced droplet aggregation on Starch Granule–fat droplet mixtures
Journal of Food Engineering, 2013Co-Authors: Bicheng Wu, Brian Degner, David Julian McclementsAbstract:Abstract The creation of high quality reduced-fat food products is challenging because the removal of fat adversely affects quality attributes, such as appearance, texture, and flavor. This study investigated the impact of pH-induced droplet aggregation on the properties of model food systems consisting of fat droplets and Starch Granules. Oil-in-water emulsions (2 wt.% oil) containing whey-protein coated lipid droplets aggregated extensively when heated (90 °C, 5 min) at pH values around their isoelectric point (pH 5) but not at lower (pH 3.5) or higher (pH 7) values, which was attributed to changes in electrostatic repulsion. The physicochemical properties of mixed lipid droplet–Starch dispersions (2 wt.% oil, 4 wt.% Starch) prepared under similar conditions (pH 3.5, 5, and 7; 90 °C for 5 min) were also measured. At pH 5, extensive lipid droplet aggregation was observed in mixed systems, which led to a large increase in their yield stress and apparent viscosity when compared to mixed systems at pH 3.5 and 7. These results show that the rheological properties of mixed fat droplet–Starch Granule suspensions can be modulated by controlling the electrostatic interactions between the fat droplets so as to change their flocculation state. This study has important implications for fabricating high quality reduced-fat products with desirable sensory attributes.
David Julian Mcclements - One of the best experts on this subject based on the ideXlab platform.
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structural design approaches for creating fat droplet and Starch Granule mimetics
Food & Function, 2017Co-Authors: David Julian Mcclements, Cheryl Chung, Bicheng WuAbstract:This article focuses on hydrogel-based strategies for creating reduced calorie foods with desirable physicochemical, sensory, and nutritional properties. Initially, the role of fat droplets and Starch Granules in foods is discussed, and then different methods for fabricating hydrogel beads are reviewed, including phase separation, antisolvent precipitation, injection, and emulsion template methods. Finally, the potential application of hydrogel beads as fat droplet and Starch Granule replacements is discussed. There is still a need for large-scale, high-throughout, and economical methods of fabricating hydrogel beads suitable for utilization within the food industry.
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microstructure rheology of mixed colloidal dispersions influence of ph induced droplet aggregation on Starch Granule fat droplet mixtures
Journal of Food Engineering, 2013Co-Authors: Bicheng Wu, Brian Degner, David Julian McclementsAbstract:Abstract The creation of high quality reduced-fat food products is challenging because the removal of fat adversely affects quality attributes, such as appearance, texture, and flavor. This study investigated the impact of pH-induced droplet aggregation on the properties of model food systems consisting of fat droplets and Starch Granules. Oil-in-water emulsions (2 wt.% oil) containing whey-protein coated lipid droplets aggregated extensively when heated (90 °C, 5 min) at pH values around their isoelectric point (pH 5) but not at lower (pH 3.5) or higher (pH 7) values, which was attributed to changes in electrostatic repulsion. The physicochemical properties of mixed lipid droplet–Starch dispersions (2 wt.% oil, 4 wt.% Starch) prepared under similar conditions (pH 3.5, 5, and 7; 90 °C for 5 min) were also measured. At pH 5, extensive lipid droplet aggregation was observed in mixed systems, which led to a large increase in their yield stress and apparent viscosity when compared to mixed systems at pH 3.5 and 7. These results show that the rheological properties of mixed fat droplet–Starch Granule suspensions can be modulated by controlling the electrostatic interactions between the fat droplets so as to change their flocculation state. This study has important implications for fabricating high quality reduced-fat products with desirable sensory attributes.
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Microstructure & rheology of mixed colloidal dispersions: Influence of pH-induced droplet aggregation on Starch Granule–fat droplet mixtures
Journal of Food Engineering, 2013Co-Authors: Bicheng Wu, Brian Degner, David Julian McclementsAbstract:Abstract The creation of high quality reduced-fat food products is challenging because the removal of fat adversely affects quality attributes, such as appearance, texture, and flavor. This study investigated the impact of pH-induced droplet aggregation on the properties of model food systems consisting of fat droplets and Starch Granules. Oil-in-water emulsions (2 wt.% oil) containing whey-protein coated lipid droplets aggregated extensively when heated (90 °C, 5 min) at pH values around their isoelectric point (pH 5) but not at lower (pH 3.5) or higher (pH 7) values, which was attributed to changes in electrostatic repulsion. The physicochemical properties of mixed lipid droplet–Starch dispersions (2 wt.% oil, 4 wt.% Starch) prepared under similar conditions (pH 3.5, 5, and 7; 90 °C for 5 min) were also measured. At pH 5, extensive lipid droplet aggregation was observed in mixed systems, which led to a large increase in their yield stress and apparent viscosity when compared to mixed systems at pH 3.5 and 7. These results show that the rheological properties of mixed fat droplet–Starch Granule suspensions can be modulated by controlling the electrostatic interactions between the fat droplets so as to change their flocculation state. This study has important implications for fabricating high quality reduced-fat products with desirable sensory attributes.
Alison M. Smith - One of the best experts on this subject based on the ideXlab platform.
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Starch synthase 4 is essential for coordination of Starch Granule formation with chloroplast division during arabidopsis leaf expansion
New Phytologist, 2013Co-Authors: Matilda Crumptontaylor, Christopher M Hylton, Regina Feil, John E Lunn, Simona Eicke, Samuel C Zeeman, Marilyn J Pike, Alison M. SmithAbstract:Arabidopsis thaliana mutants lacking the SS4 isoform of Starch synthase have strongly reduced numbers of Starch Granules per chloroplast, suggesting that SS4 is necessary for the normal generation of Starch Granules. To establish whether it plays a direct role in this process, we investigated the circumstances in which Granules are formed in ss4 mutants. Starch Granule numbers and distribution and the accumulation of Starch synthase substrates and products were investigated during ss4 leaf development, and in ss4 mutants carrying mutations or transgenes that affect Starch turnover or chloroplast volume. We found that immature ss4 leaves have no Starch Granules, but accumulate high concentrations of the Starch synthase substrate ADPglucose. Granule numbers are partially restored by elevating the capacity for glucan synthesis (via expression of bacterial glycogen synthase) or by increasing the volumes of individual chloroplasts (via introduction of arc mutations). However, these Granules are abnormal in distribution, size and shape. SS4 is an essential component of a mechanism that coordinates Granule formation with chloroplast division during leaf expansion and determines the abundance and the flattened, discoid shape of leaf Starch Granules. Keywords: ADPglucose, Arabidopsis thaliana, chloroplast, leaf expansion, Starch Granule, Starch synthase, Starch synthesis Introduction The process by which Starch Granules arise is not known. Suggestions range from largely physico-chemical mechanisms (Doi, 1965; Geddes & Greenwood, 1969; Ziegler et al., 2005) to the existence of specific protein primers analogous to the glycogenins of fungi and animals (e.g. Rothschild & Tandecarz, 1994; Singh et al., 1995; Langeveld et al., 2002; Chatterjee et al., 2005). Recent attention has focussed on the role of one isoform of soluble Starch synthase, Starch synthase 4 (SS4, At4 g18240). Although SS4 contributes little to total Starch synthase activity, ss4 mutants of Arabidopsis have at most one or two Starch Granules per chloroplast (Roldan et al., 2007) rather than the normal five or six (Crumpton-Taylor et al., 2012). No other Starch synthase is individually necessary for normal Granule numbers (Roldan et al., 2007), thus SS4 may have a specific function in Granule formation. However, other isoforms of Starch synthase may partially substitute for this function. The additional loss of SS3 further reduces Starch Granule numbers in the ss4 mutant background (Szydlowski et al., 2009; Merida & D′Hulst, 2012). The importance of SS4 for Starch Granule formation remains to be established. First, it is not known whether SS4 is required primarily for maintenance of Starch Granule numbers in mature leaves, or whether it also has a role in immature leaves where new Granules arise in concert with chloroplast division (Crumpton-Taylor et al., 2012). Second, it is not clear whether the reduction in Starch Granule numbers in ss4 mutants is a direct or an indirect consequence of the loss of SS4. Mutants have several additional phenotypes including reduced growth rates, altered Starch Granule anatomy and morphology and a reduction in the extent of diel Starch turnover (Roldan et al., 2007). It remains possible that the reduction in Granule numbers in ss4 mutants is an indirect consequence of one of these alterations. Third, a recent study suggests that SS4 may be limiting for Starch synthesis in wild-type plants. Its overexpression reportedly results in higher concentrations of Starch at the end of the day and accelerated plant growth (Gamez-Arjona et al., 2011). These results have important implications for the control of Starch turnover and are of biotechnological interest, but the relationship between Starch concentrations and Starch Granule numbers and sizes in plants with elevated SS4 was not reported. The aim of our work was to establish whether SS4 has a direct or an indirect role in Starch Granule formation, and to shed further light on where and when its actions are required for the establishment of normal Granule numbers. To this end we examined the phenotype of the ss4 mutant through leaf development, and investigated the impact of loss of SS4 in mutant and transgenic backgrounds in which Starch metabolism is altered or chloroplast volumes are abnormally large. Our results indicate that SS4 is directly and specifically required for the establishment of normal numbers and distributions of Starch Granules during leaf expansion, and that it is also necessary for the normal flattened, discoid shape of leaf Starch Granules.
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control of Starch Granule numbers in arabidopsis chloroplasts
Plant Physiology, 2012Co-Authors: Matilda Crumptontaylor, Scott Grandison, Kenneth M Y Png, A J Bushby, Alison M. SmithAbstract:The aim of this work was to investigate Starch Granule numbers in Arabidopsis (Arabidopsis thaliana) leaves. Lack of quantitative information on the extent of genetic, temporal, developmental, and environmental variation in Granule numbers is an important limitation in understanding control of Starch degradation and the mechanism of Granule initiation. Two methods were developed for reliable estimation of numbers of Granules per chloroplast. First, direct measurements were made on large series of consecutive sections of mesophyll tissue obtained by focused ion beam-scanning electron microscopy. Second, average numbers were calculated from the Starch contents of leaves and chloroplasts and estimates of Granule mass based on Granule dimensions. Examination of wild-type plants and accumulation and regulation of chloroplast (arc) mutants with few, large chloroplasts provided the following new insights. There is wide variation in chloroplast volumes in cells of wild-type leaves. Granule numbers per chloroplast are correlated with chloroplast volume, i.e. large chloroplasts have more Granules than small chloroplasts. Mature leaves of wild-type plants and arc mutants have approximately the same number of Granules per unit volume of stroma, regardless of the size and number of chloroplasts per cell. Granule numbers per unit volume of stroma are also relatively constant in immature leaves but are greater than in mature leaves. Granule initiation occurs as chloroplasts divide in immature leaves, but relatively little initiation occurs in mature leaves. Changes in leaf Starch content over the diurnal cycle are largely brought about by changes in the volume of a fixed number of Granules.
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Starch Granule initiation is controlled by a heteromultimeric isoamylase in potato tubers
Proceedings of the National Academy of Sciences of the United States of America, 2004Co-Authors: Regla Bustos, Robert Seale, Anne Edwards, Christopher M Hylton, Brendan Fahy, Miranda N Nebane, Cathie Martin, Alison M. SmithAbstract:Starch Granule initiation is not understood, but recent evidence implicates a Starch debranching enzyme, isoamylase, in the control of this process. Potato tubers contain isoamylase activity attributable to a heteromultimeric protein containing Stisa1 and Stisa2, the products of two of the three isoamylase genes of potato. To discover whether this enzyme is involved in Starch Granule initiation, activity was reduced by expression of antisense RNA for Stisa1 or Stisa2. Transgenic tubers accumulated a small amount of a soluble glucan, similar in structure to the phytoglycogen of cereal, Arabidopsis, and Chlamydomonas mutants lacking isoamylase. The major effect, however, was on the number of Starch Granules. Transgenic tubers accumulated large numbers of tiny Granules not seen in normal tubers. These data indicate that the heteromultimeric isoamylase functions during Starch synthesis to suppress the initiation of glucan molecules in the plastid stroma that would otherwise crystallize to nucleate new Starch Granules.
Simona Eicke - One of the best experts on this subject based on the ideXlab platform.
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Starch synthase5 a noncanonical Starch synthase like protein promotes Starch Granule initiation in arabidopsis
The Plant Cell, 2020Co-Authors: Melanie R Abt, Barbara Pfister, Mayank Sharma, Simona Eicke, Leo Burgy, Isabel Neale, David Seung, Samuel C ZeemanAbstract:What determines the number of Starch Granules in plastids is an enigmatic aspect of Starch metabolism. Several structurally and functionally diverse proteins have been implicated in the Granule initiation process in Arabidopsis (Arabidopsis thaliana), with each protein exerting a varying degree of influence. Here, we show that a conserved Starch synthase-like protein, Starch SYNTHASE5 (SS5), regulates the number of Starch Granules that form in Arabidopsis chloroplasts. Among the Starch synthases, SS5 is most closely related to SS4, a major determinant of Granule initiation and morphology. However, unlike SS4 and the other Starch synthases, SS5 is a noncanonical isoform that lacks catalytic glycosyltransferase activity. Nevertheless, loss of SS5 reduces Starch Granule numbers that form per chloroplast in Arabidopsis, and ss5 mutant Starch Granules are larger than wild-type Granules. Like SS4, SS5 has a conserved putative surface binding site for glucans and also interacts with MYOSIN-RESEMBLING CHLOROPLAST PROTEIN, a proposed structural protein influential in Starch Granule initiation. Phenotypic analysis of a suite of double mutants lacking both SS5 and other proteins implicated in Starch Granule initiation allows us to propose how SS5 may act in this process.
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two plastidial coiled coil proteins are essential for normal Starch Granule initiation in arabidopsis
The Plant Cell, 2018Co-Authors: David Seung, Simona Eicke, Leo Burgy, Tina B Schreier, Samuel C ZeemanAbstract:The mechanism of Starch Granule initiation in chloroplasts is not fully understood. Here, we aimed to build on our recent discovery that PROTEIN TARGETING TO Starch (PTST) family members, PTST2 and PTST3, are key players in Starch Granule initiation, by identifying and characterizing additional proteins involved in the process in Arabidopsis thaliana chloroplasts. Using immunoprecipitation and mass spectrometry, we demonstrate that PTST2 interacts with two plastidial coiled-coil proteins. Surprisingly, one of the proteins is the thylakoid-associated MAR BINDING FILAMENT-LIKE PROTEIN1 (MFP1), which was proposed to bind plastid nucleoids. The other protein, MYOSIN-RESEMBLING CHLOROPLAST PROTEIN (MRC), contains long coiled coils and no known domains. Whereas wild-type chloroplasts contained multiple Starch Granules, only one large Granule was observed in most chloroplasts of the mfp1 and mrc mutants. The mfp1 mrc double mutant had a higher proportion of chloroplasts containing no visible Granule than either single mutant and accumulated ADP-glucose, the substrate for Starch synthesis. PTST2 was partially associated with the thylakoid membranes in wild-type plants, and fluorescently tagged PTST2 was located in numerous discrete patches within the chloroplast in which MFP1 was also located. In the mfp1 mutant, PTST2 was not associated with the thylakoids and formed discrete puncta, suggesting that MFP1 is necessary for normal PTST2 localization. Overall, we reveal that proper Granule initiation requires the presence of MFP1 and MRC, and the correct location of PTST2.
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the Starch Granule associated protein early starvation1 is required for the control of Starch degradation in arabidopsis thaliana leaves
The Plant Cell, 2016Co-Authors: Doreen Feike, Simona Eicke, David Seung, Alexander Graf, Sylvain Bischof, Tamaryn Ellick, Mario Coiro, Sebastian Soyk, Tabea Mettleraltmann, Martin TrickAbstract:To uncover components of the mechanism that adjusts the rate of leaf Starch degradation to the length of the night, we devised a screen for mutant Arabidopsis thaliana plants in which Starch reserves are prematurely exhausted. The mutation in one such mutant, named early starvation1 (esv1), eliminates a previously uncharacterized protein. Starch in mutant leaves is degraded rapidly and in a nonlinear fashion, so that reserves are exhausted 2 h prior to dawn. The ESV1 protein and a similar uncharacterized Arabidopsis protein (named Like ESV1 [LESV]) are located in the chloroplast stroma and are also bound into Starch Granules. The region of highest similarity between the two proteins contains a series of near-repeated motifs rich in tryptophan. Both proteins are conserved throughout Starch-synthesizing organisms, from angiosperms and monocots to green algae. Analysis of transgenic plants lacking or overexpressing ESV1 or LESV, and of double mutants lacking ESV1 and another protein necessary for Starch degradation, leads us to propose that these proteins function in the organization of the Starch Granule matrix. We argue that their misexpression affects Starch degradation indirectly, by altering matrix organization and, thus, accessibility of Starch polymers to Starch-degrading enzymes.
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Starch synthase 4 is essential for coordination of Starch Granule formation with chloroplast division during arabidopsis leaf expansion
New Phytologist, 2013Co-Authors: Matilda Crumptontaylor, Christopher M Hylton, Regina Feil, John E Lunn, Simona Eicke, Samuel C Zeeman, Marilyn J Pike, Alison M. SmithAbstract:Arabidopsis thaliana mutants lacking the SS4 isoform of Starch synthase have strongly reduced numbers of Starch Granules per chloroplast, suggesting that SS4 is necessary for the normal generation of Starch Granules. To establish whether it plays a direct role in this process, we investigated the circumstances in which Granules are formed in ss4 mutants. Starch Granule numbers and distribution and the accumulation of Starch synthase substrates and products were investigated during ss4 leaf development, and in ss4 mutants carrying mutations or transgenes that affect Starch turnover or chloroplast volume. We found that immature ss4 leaves have no Starch Granules, but accumulate high concentrations of the Starch synthase substrate ADPglucose. Granule numbers are partially restored by elevating the capacity for glucan synthesis (via expression of bacterial glycogen synthase) or by increasing the volumes of individual chloroplasts (via introduction of arc mutations). However, these Granules are abnormal in distribution, size and shape. SS4 is an essential component of a mechanism that coordinates Granule formation with chloroplast division during leaf expansion and determines the abundance and the flattened, discoid shape of leaf Starch Granules. Keywords: ADPglucose, Arabidopsis thaliana, chloroplast, leaf expansion, Starch Granule, Starch synthase, Starch synthesis Introduction The process by which Starch Granules arise is not known. Suggestions range from largely physico-chemical mechanisms (Doi, 1965; Geddes & Greenwood, 1969; Ziegler et al., 2005) to the existence of specific protein primers analogous to the glycogenins of fungi and animals (e.g. Rothschild & Tandecarz, 1994; Singh et al., 1995; Langeveld et al., 2002; Chatterjee et al., 2005). Recent attention has focussed on the role of one isoform of soluble Starch synthase, Starch synthase 4 (SS4, At4 g18240). Although SS4 contributes little to total Starch synthase activity, ss4 mutants of Arabidopsis have at most one or two Starch Granules per chloroplast (Roldan et al., 2007) rather than the normal five or six (Crumpton-Taylor et al., 2012). No other Starch synthase is individually necessary for normal Granule numbers (Roldan et al., 2007), thus SS4 may have a specific function in Granule formation. However, other isoforms of Starch synthase may partially substitute for this function. The additional loss of SS3 further reduces Starch Granule numbers in the ss4 mutant background (Szydlowski et al., 2009; Merida & D′Hulst, 2012). The importance of SS4 for Starch Granule formation remains to be established. First, it is not known whether SS4 is required primarily for maintenance of Starch Granule numbers in mature leaves, or whether it also has a role in immature leaves where new Granules arise in concert with chloroplast division (Crumpton-Taylor et al., 2012). Second, it is not clear whether the reduction in Starch Granule numbers in ss4 mutants is a direct or an indirect consequence of the loss of SS4. Mutants have several additional phenotypes including reduced growth rates, altered Starch Granule anatomy and morphology and a reduction in the extent of diel Starch turnover (Roldan et al., 2007). It remains possible that the reduction in Granule numbers in ss4 mutants is an indirect consequence of one of these alterations. Third, a recent study suggests that SS4 may be limiting for Starch synthesis in wild-type plants. Its overexpression reportedly results in higher concentrations of Starch at the end of the day and accelerated plant growth (Gamez-Arjona et al., 2011). These results have important implications for the control of Starch turnover and are of biotechnological interest, but the relationship between Starch concentrations and Starch Granule numbers and sizes in plants with elevated SS4 was not reported. The aim of our work was to establish whether SS4 has a direct or an indirect role in Starch Granule formation, and to shed further light on where and when its actions are required for the establishment of normal Granule numbers. To this end we examined the phenotype of the ss4 mutant through leaf development, and investigated the impact of loss of SS4 in mutant and transgenic backgrounds in which Starch metabolism is altered or chloroplast volumes are abnormally large. Our results indicate that SS4 is directly and specifically required for the establishment of normal numbers and distributions of Starch Granules during leaf expansion, and that it is also necessary for the normal flattened, discoid shape of leaf Starch Granules.
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evidence for distinct mechanisms of Starch Granule breakdown in plants
Journal of Biological Chemistry, 2006Co-Authors: Simona Eicke, Thierry Delatte, Martin Umhang, Martine Trevisan, David Thorneycroft, Steven M Smith, Samuel C ZeemanAbstract:The aim of this work was to understand the initial steps of Starch breakdown inside chloroplasts. In the non-living endosperm of germinating cereal grains, Starch breakdown is initiated by -amylase secreted from surrounding cells. However, loss of -amylase from Arabidopsis does not prevent chloroplastic Starch breakdown (Yu, T.-S., Zeeman, S. C., Thorneycroft, D., Fulton, D. C., Dunstan, H., Lue, W.-L., Hegemann, B., Tung, S.-Y., Umemoto, T., Chapple, A., Tsai, D.-L., Wang, S.-M, Smith, A. M., Chen, J., and Smith, S. M. (2005) J. Biol. Chem. 280, 9773–9779), implying that other enzymes must attack the Starch Granule. Here, we present evidence that the debranching enzyme isoamylase 3 (ISA3) acts at the surface of the Starch Granule. Atisa3 mutants have more leaf Starch and a slower rate of Starch breakdown than wild-type plants. The amylopectin of Atisa3 contains many very short branches and ISA3-GFP localizes to Granule-like structures inside chloroplasts. We suggest that ISA3 removes short branches from the Granule surface. To understand how some Starch is still degraded in Atisa3 mutants we eliminated a second debranching enzyme, limit dextrinase (pullulanase-type). Atlda mutants are indistinguishable from the wild type. However, the Atisa3/Atlda double mutant has a more severe Starch-excess phenotype and a slower rate of Starch breakdown than Atisa3 single mutants. The double mutant accumulates soluble branched oligosaccharides (limit dextrins) that are undetectable in the wild-type and the single mutants. Together these results suggest that glucan debranching occurs primarily at the Granule surface via ISA3, but in its absence soluble branched glucans are debranched in the stroma via limit dextrinase. Consistent with this model, chloroplastic -amylase AtAMY3, which could release soluble branched glucans, is induced in Atisa3 and in the Atisa3/Atlda double mutant.