The Experts below are selected from a list of 813 Experts worldwide ranked by ideXlab platform
Miyo Terao Morita - One of the best experts on this subject based on the ideXlab platform.
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Micromanipulation of Amyloplasts with optical tweezers in Arabidopsis stems
2020Co-Authors: Yoshinori Abe, Miyo Terao Morita, Keisuke Meguriya, Takahisa Matsuzaki, Teruki Sugiyama, Hiroshi Yoshikawa, Masatsugu ToyotaAbstract:AbstractIntracellular sedimentation of highly dense, starch-filled Amyloplasts toward the gravity vector is likely a key initial step for gravity sensing in plants. However, recent live-cell imaging technology revealed that most Amyloplasts continuously exhibit dynamic, saltatory movements in the endodermal cells of Arabidopsis stems. These complicated movements led to questions about what type of Amyloplast movement triggers gravity sensing. Here we show that a confocal microscope equipped with optical tweezers can be a powerful tool to trap and manipulate Amyloplasts noninvasively, while simultaneously observing cellular responses such as vacuolar dynamics in living cells. A near-infrared (λ = 1064 nm) laser that was focused into the endodermal cells at 1 mW of laser power attracted and captured Amyloplasts at the laser focus. The optical force exerted on the Amyloplasts was theoretically estimated to be up to 1 pN. Interestingly, endosomes and trans-Golgi networks were trapped at 30 mW but not at 1 mW, which is probably due to lower refractive indices of these organelles than that of the Amyloplasts. Because Amyloplasts are in close proximity to vacuolar membranes in endodermal cells, their physical interaction could be visualized in real time. The vacuolar membranes drastically stretched and deformed in response to the manipulated movements of Amyloplasts by optical tweezers. Our new method provides deep insights into the biophysical properties of plant organelles in vivo and opens a new avenue for studying gravity-sensing mechanisms in plants.
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gravity sensing and signal conversion in plant gravitropism
Journal of Experimental Botany, 2019Co-Authors: Moritaka Nakamura, Miyo Terao Morita, Takeshi NishimuraAbstract:Plant organs control their growth orientation in response to gravity. Within gravity-sensing cells, the input (gravity sensing) and signal conversion (gravity signalling) progress sequentially. The cells contain a number of high-density, starch-accumulating Amyloplasts, which sense gravity when they reposition themselves by sedimentation to the bottom of the cell when the plant organ is re-orientated. This triggers the next step of gravity signalling, when the physical signal generated by the sedimentation of the Amyloplasts is converted into a biochemical signal, which redirects auxin transport towards the lower flank of the plant organ. This review focuses on recent advances in our knowledge of the regulatory mechanisms that underlie Amyloplast sedimentation and the system by which this is perceived, and on recent progress in characterising the factors that play significant roles in gravity signalling by which the sedimentation is linked to the regulation of directional auxin transport. Finally, we discuss the contribution of gravity signalling factors to the mechanisms that control the gravitropic set-point angle.
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live cell imaging of cytoskeletal and organelle dynamics in gravity sensing cells in plant gravitropism
Methods of Molecular Biology, 2015Co-Authors: Moritaka Nakamura, Masao Tasaka, Masatsugu Toyota, Miyo Terao MoritaAbstract:: Plants sense gravity and change their morphology/growth direction accordingly (gravitropism). The early process of gravitropism, gravity sensing, is supposed to be triggered by sedimentation of starch-filled plastids (Amyloplasts) in statocytes such as root columella cells and shoot endodermal cells. For several decades, many scientists have focused on characterizing the role of the Amyloplasts and observed their intracellular sedimentation in various plants. Recently, it has been discovered that the complex sedimentary movements of the Amyloplasts are created not only by gravity but also by cytoskeletal/organelle dynamics, such as those of actin filaments and the vacuolar membrane. Thus, to understand how plants sense gravity, we need to analyze both Amyloplast movements and their regulatory systems in statocytes. We have developed a vertical-stage confocal microscope that allows multicolor fluorescence imaging of Amyloplasts, actin filaments and vacuolar membranes in vertically oriented plant tissues. We also developed a centrifuge microscope that allows bright-field imaging of Amyloplasts during centrifugation. These microscope systems provide new insights into gravity-sensing mechanisms in Arabidopsis.
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a unique heat repeat containing protein shoot gravitropism6 is involved in vacuolar membrane dynamics in gravity sensing cells of arabidopsis inflorescence stem
Plant and Cell Physiology, 2014Co-Authors: Yasuko Hashiguchi, Takehide Kato, Daisuke Yano, Kiyoshi Nagafusa, Masao Tasaka, Chieko Saito, Akihiko Nakano, Takashi Ueda, Tomohiro Uemura, Miyo Terao MoritaAbstract:Plant vacuoles play critical roles in development, growth and stress responses. In mature cells, vacuolar membranes (VMs) display several types of structures, which are formed by invagination and folding of VMs into the lumenal side and can gradually move and change shape. Although such VM structures are observed in a broad range of tissue types and plant species, the molecular mechanism underlying their formation and maintenance remains unclear. Here, we report that a novel HEAT-repeat protein, SHOOT GRAVITROPISM6 (SGR6), of Arabidopsis is involved in the control of morphological changes and dynamics of VM structures in endodermal cells, which are the gravity-sensing cells in shoots. SGR6 is a membrane-associated protein that is mainly localized to the VM in stem endodermal cells. The sgr6 mutant stem exhibits a reduced gravitropic response. Higher plants utilize Amyloplast sedimentation as a means to sense gravity direction. Amyloplasts are surrounded by VMs in Arabidopsis endodermal cells, and the flexible and dynamic structure of VMs is important for Amyloplast sedimentation. We demonstrated that such dynamic features of VMs are gradually lost in sgr6 endodermal cells during a 30 min observation period. Histological analysis revealed that Amyloplast sedimentation was impaired in sgr6. Detailed live-cell imaging analyses revealed that the VM structures in sgr6 had severe defects in morphological changes and dynamics. Our results suggest that SGR6 is a novel protein involved in the formation and/or maintenance of invaginated VM structures in gravity-sensing cells.
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mechanism of higher plant gravity sensing
American Journal of Botany, 2013Co-Authors: Yasuko Hashiguchi, Masao Tasaka, Miyo Terao MoritaAbstract:Higher plants have developed statocytes, specialized tissues or cells for gravity sensing, and subsequent signal formation. Root and shoot statocytes commonly harbor a number of Amyloplasts, and Amyloplast sedimentation in the direction of gravity is a critical process in gravity sensing. However, the molecular mechanism underlying Amyloplast-dependent gravity sensing is largely unknown. In this review, we mainly describe the molecular basis for the gravity sensing mechanism, i.e., the molecules and their functions involved in Amyloplast sedimentation. Several analyses of statocyte images in living plant organs have implied differences in the regulation of Amyloplast movements between root and shoot statocytes. Amyloplasts in shoot statocytes display not only sedimentable but upward, saltatory movements, but the latter are rarely observed in root statocytes. A series of genetic studies on shoot gravitropism mutants of Arabidopsis thaliana has revealed that two intracellular components, the vacuolar membrane (VM) and actin microfilaments (AFs), within the shoot statocyte play important roles in Amyloplast dynamics. Flexible VM structures surrounding the Amyloplasts seem to allow them to freely sediment toward the bottom of cells. In contrast, long actin cables mediate the saltatory movements of Amyloplasts. Thus, Amyloplasts in shoot statocytes undergo a dynamic equilibrium of movement, and a proper intracellular environment for statocytes is essential for normal shoot gravitropism. Further analyses to identify the molecular regulators of Amyloplast dynamics, including sedimentation, may contribute to an understanding of the gravity sensing mechanism in higher plants.
Michael J. Emes - One of the best experts on this subject based on the ideXlab platform.
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analysis of protein complexes in wheat Amyloplasts reveals functional interactions among starch biosynthetic enzymes
Plant Physiology, 2008Co-Authors: Ian J. Tetlow, Fushan Liu, Robin Wait, Matthew K. Morell, Kim Gabriele Beisel, S Cameron, Amina Makhmoudova, Nicole S Bresolin, Michael J. EmesAbstract:Protein-protein interactions among enzymes of amylopectin biosynthesis were investigated in developing wheat (Triticum aestivum) endosperm. Physical interactions between starch branching enzymes (SBEs) and starch synthases (SSs) were identified from endosperm Amyloplasts during the active phase of starch deposition in the developing grain using immunoprecipitation and cross-linking strategies. Coimmunoprecipitation experiments using peptide-specific antibodies indicate that at least two distinct complexes exist containing SSI, SSIIa, and either of SBEIIa or SBEIIb. Chemical cross linking was used to identify protein complexes containing SBEs and SSs from Amyloplast extracts. Separation of extracts by gel filtration chromatography demonstrated the presence of SBE and SS forms in protein complexes of around 260 kD and that SBEII forms may also exist as homodimers. Analysis of cross-linked 260-kD aggregation products from Amyloplast lysates by mass spectrometry confirmed SSI, SSIIa, and SBEII forms as components of one or more protein complexes in Amyloplasts. In vitro phosphorylation experiments with γ-32P-ATP indicated that SSII and both forms of SBEII are phosphorylated. Treatment of the partially purified 260-kD SS-SBE complexes with alkaline phosphatase caused dissociation of the assembly into the respective monomeric proteins, indicating that formation of SS-SBE complexes is phosphorylation dependent. The 260-kD SS-SBEII protein complexes are formed around 10 to 15 d after pollination and were shown to be catalytically active with respect to both SS and SBE activities. Prior to this developmental stage, SSI, SSII, and SBEII forms were detectable only in monomeric form. High molecular weight forms of SBEII demonstrated a higher affinity for in vitro glucan substrates than monomers. These results provide direct evidence for the existence of protein complexes involved in amylopectin biosynthesis.
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Characterization of ADP-glucose transport across the cereal endosperm Amyloplast envelope
Journal of experimental botany, 2007Co-Authors: Caroline G. Bowsher, Michael J. Emes, Edward F. A. L. Scrase-field, Sergio Esposito, Ian J. TetlowAbstract:Most of the carbon used for starch biosynthesis in cereal endosperms is derived from ADP-glucose (ADP-Glc) synthesized by extra-plastidial AGPase activity, and imported directly across the Amyloplast envelope. The properties of the wheat endosperm Amyloplast ADP-Glc transporter were analysed with respect to substrate kinetics and specificities using reconstituted Amyloplast envelope proteins in a proteoliposome-based assay system, as well as with isolated intact organelles. Experiments with liposomes showed that ADP-Glc transport was dependent on counter-exchange with other adenylates. Rates of ADP-Glc transport were highest with ADP and AMP as counter-exchange substrates, and kinetic analysis revealed that the transport system has a similar affinity for ADP and AMP. Measurement of ADP and AMP efflux from intact Amyloplasts showed that, under conditions of ADP-Glc-dependent starch biosynthesis, ADP is exported from the plastid at a rate equal to that of ADP-Glc utilization by starch synthases. Photo-affinity labelling of Amyloplast membranes with the substrate analogue 8-azido-[alpha-32P]ADP-Glc showed that the polypeptide involved in substrate binding is an integral membrane protein of 38 kDa. This study shows that the ADP-Glc transporter in cereal endosperm Amyloplasts imports ADP-Glc in exchange for ADP which is produced as a by-product of the starch synthase reaction inside the plastid.
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protein phosphorylation in Amyloplasts regulates starch branching enzyme activity and protein protein interactions
The Plant Cell, 2004Co-Authors: Ian J. Tetlow, Caroline G. Bowsher, Sergio Esposito, Robin Wait, Rut Akkasaeng, Matthew K. Morell, Behjat Kosarhashemi, Michael J. EmesAbstract:Protein phosphorylation in Amyloplasts and chloroplasts of Triticum aestivum (wheat) was investigated after the incubation of intact plastids with γ-32P-ATP. Among the soluble phosphoproteins detected in plastids, three forms of starch branching enzyme (SBE) were phosphorylated in Amyloplasts (SBEI, SBEIIa, and SBEIIb), and both forms of SBE in chloroplasts (SBEI and SBEIIa) were shown to be phosphorylated after sequencing of the immunoprecipitated 32P-labeled phosphoproteins using quadrupole-orthogonal acceleration time of flight mass spectrometry. Phosphoamino acid analysis of the phosphorylated SBE forms indicated that the proteins are all phosphorylated on Ser residues. Analysis of starch granule–associated phosphoproteins after incubation of intact Amyloplasts with γ-32P-ATP indicated that the granule-associated forms of SBEII and two granule-associated forms of starch synthase (SS) are phosphorylated, including SSIIa. Measurement of SBE activity in Amyloplasts and chloroplasts showed that phosphorylation activated SBEIIa (and SBEIIb in Amyloplasts), whereas dephosphorylation using alkaline phosphatase reduced the catalytic activity of both enzymes. Phosphorylation and dephosphorylation had no effect on the measurable activity of SBEI in Amyloplasts and chloroplasts, and the activities of both granule-bound forms of SBEII in Amyloplasts were unaffected by dephosphorylation. Immunoprecipitation experiments using peptide-specific anti-SBE antibodies showed that SBEIIb and starch phosphorylase each coimmunoprecipitated with SBEI in a phosphorylation-dependent manner, suggesting that these enzymes may form protein complexes within the Amyloplast in vivo. Conversely, dephosphorylation of immunoprecipitated protein complex led to its disassembly. This article reports direct evidence that enzymes of starch metabolism (amylopectin synthesis) are regulated by protein phosphorylation and indicate a wider role for protein phosphorylation and protein–protein interactions in the control of starch anabolism and catabolism.
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starch synthesis and carbohydrate oxidation in Amyloplasts from developing wheat endosperm
Planta, 1994Co-Authors: Ian J. Tetlow, Kerry J Blissett, Michael J. EmesAbstract:The rates of incorporation of various metabolites into starch by isolated Amyloplasts from developing endosperm of spring wheat (Triticum aestivum L. cv. Axona) were examined. Of the metabolites tested that were likely to be present in the cytosol at concentrations sufficient to sustain starch synthesis, only glucose 1-phosphate (Glc1P) supported physiologically relevant rates of starch synthesis. Incorporation of Glc1P into starch was both dependent on the presence of ATP and intact organelles. The rate of incorporation of hexose into starch became saturated at a Glc1P concentration of less than 1 mol·m-3 in the presence of 1 mol·m-3 ATP. Starch synthesis from 5 mol · m-3 ADP-glucose supplied to the organelles occurred at rates 15-fold higher than from similar concentrations of Glc1P, but it is argued that this is probably of little physiological relevance. The net incorporation of hexose units into starch from GlclP was inhibited 50% by 100 mmol.m-3 carboxyatractyloside. Carbohydrate oxidation in the Amyloplast was stimulated by the addition of 2-oxoglutarate and glutamine, and in such circumstances incorporation of14C-labelled metabolites into starch was reduced. Glucose 6-phosphate proved to be a better substrate for oxidative pathways than Glc1P. Our results suggest that Glc1P is the primary substrate for starch synthesis in developing wheat endosperm, and that ATP required for starch synthesis is imported via an adenylate translocator.
Ian J. Tetlow - One of the best experts on this subject based on the ideXlab platform.
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analysis of protein complexes in wheat Amyloplasts reveals functional interactions among starch biosynthetic enzymes
Plant Physiology, 2008Co-Authors: Ian J. Tetlow, Fushan Liu, Robin Wait, Matthew K. Morell, Kim Gabriele Beisel, S Cameron, Amina Makhmoudova, Nicole S Bresolin, Michael J. EmesAbstract:Protein-protein interactions among enzymes of amylopectin biosynthesis were investigated in developing wheat (Triticum aestivum) endosperm. Physical interactions between starch branching enzymes (SBEs) and starch synthases (SSs) were identified from endosperm Amyloplasts during the active phase of starch deposition in the developing grain using immunoprecipitation and cross-linking strategies. Coimmunoprecipitation experiments using peptide-specific antibodies indicate that at least two distinct complexes exist containing SSI, SSIIa, and either of SBEIIa or SBEIIb. Chemical cross linking was used to identify protein complexes containing SBEs and SSs from Amyloplast extracts. Separation of extracts by gel filtration chromatography demonstrated the presence of SBE and SS forms in protein complexes of around 260 kD and that SBEII forms may also exist as homodimers. Analysis of cross-linked 260-kD aggregation products from Amyloplast lysates by mass spectrometry confirmed SSI, SSIIa, and SBEII forms as components of one or more protein complexes in Amyloplasts. In vitro phosphorylation experiments with γ-32P-ATP indicated that SSII and both forms of SBEII are phosphorylated. Treatment of the partially purified 260-kD SS-SBE complexes with alkaline phosphatase caused dissociation of the assembly into the respective monomeric proteins, indicating that formation of SS-SBE complexes is phosphorylation dependent. The 260-kD SS-SBEII protein complexes are formed around 10 to 15 d after pollination and were shown to be catalytically active with respect to both SS and SBE activities. Prior to this developmental stage, SSI, SSII, and SBEII forms were detectable only in monomeric form. High molecular weight forms of SBEII demonstrated a higher affinity for in vitro glucan substrates than monomers. These results provide direct evidence for the existence of protein complexes involved in amylopectin biosynthesis.
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Characterization of ADP-glucose transport across the cereal endosperm Amyloplast envelope
Journal of experimental botany, 2007Co-Authors: Caroline G. Bowsher, Michael J. Emes, Edward F. A. L. Scrase-field, Sergio Esposito, Ian J. TetlowAbstract:Most of the carbon used for starch biosynthesis in cereal endosperms is derived from ADP-glucose (ADP-Glc) synthesized by extra-plastidial AGPase activity, and imported directly across the Amyloplast envelope. The properties of the wheat endosperm Amyloplast ADP-Glc transporter were analysed with respect to substrate kinetics and specificities using reconstituted Amyloplast envelope proteins in a proteoliposome-based assay system, as well as with isolated intact organelles. Experiments with liposomes showed that ADP-Glc transport was dependent on counter-exchange with other adenylates. Rates of ADP-Glc transport were highest with ADP and AMP as counter-exchange substrates, and kinetic analysis revealed that the transport system has a similar affinity for ADP and AMP. Measurement of ADP and AMP efflux from intact Amyloplasts showed that, under conditions of ADP-Glc-dependent starch biosynthesis, ADP is exported from the plastid at a rate equal to that of ADP-Glc utilization by starch synthases. Photo-affinity labelling of Amyloplast membranes with the substrate analogue 8-azido-[alpha-32P]ADP-Glc showed that the polypeptide involved in substrate binding is an integral membrane protein of 38 kDa. This study shows that the ADP-Glc transporter in cereal endosperm Amyloplasts imports ADP-Glc in exchange for ADP which is produced as a by-product of the starch synthase reaction inside the plastid.
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protein phosphorylation in Amyloplasts regulates starch branching enzyme activity and protein protein interactions
The Plant Cell, 2004Co-Authors: Ian J. Tetlow, Caroline G. Bowsher, Sergio Esposito, Robin Wait, Rut Akkasaeng, Matthew K. Morell, Behjat Kosarhashemi, Michael J. EmesAbstract:Protein phosphorylation in Amyloplasts and chloroplasts of Triticum aestivum (wheat) was investigated after the incubation of intact plastids with γ-32P-ATP. Among the soluble phosphoproteins detected in plastids, three forms of starch branching enzyme (SBE) were phosphorylated in Amyloplasts (SBEI, SBEIIa, and SBEIIb), and both forms of SBE in chloroplasts (SBEI and SBEIIa) were shown to be phosphorylated after sequencing of the immunoprecipitated 32P-labeled phosphoproteins using quadrupole-orthogonal acceleration time of flight mass spectrometry. Phosphoamino acid analysis of the phosphorylated SBE forms indicated that the proteins are all phosphorylated on Ser residues. Analysis of starch granule–associated phosphoproteins after incubation of intact Amyloplasts with γ-32P-ATP indicated that the granule-associated forms of SBEII and two granule-associated forms of starch synthase (SS) are phosphorylated, including SSIIa. Measurement of SBE activity in Amyloplasts and chloroplasts showed that phosphorylation activated SBEIIa (and SBEIIb in Amyloplasts), whereas dephosphorylation using alkaline phosphatase reduced the catalytic activity of both enzymes. Phosphorylation and dephosphorylation had no effect on the measurable activity of SBEI in Amyloplasts and chloroplasts, and the activities of both granule-bound forms of SBEII in Amyloplasts were unaffected by dephosphorylation. Immunoprecipitation experiments using peptide-specific anti-SBE antibodies showed that SBEIIb and starch phosphorylase each coimmunoprecipitated with SBEI in a phosphorylation-dependent manner, suggesting that these enzymes may form protein complexes within the Amyloplast in vivo. Conversely, dephosphorylation of immunoprecipitated protein complex led to its disassembly. This article reports direct evidence that enzymes of starch metabolism (amylopectin synthesis) are regulated by protein phosphorylation and indicate a wider role for protein phosphorylation and protein–protein interactions in the control of starch anabolism and catabolism.
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starch synthesis and carbohydrate oxidation in Amyloplasts from developing wheat endosperm
Planta, 1994Co-Authors: Ian J. Tetlow, Kerry J Blissett, Michael J. EmesAbstract:The rates of incorporation of various metabolites into starch by isolated Amyloplasts from developing endosperm of spring wheat (Triticum aestivum L. cv. Axona) were examined. Of the metabolites tested that were likely to be present in the cytosol at concentrations sufficient to sustain starch synthesis, only glucose 1-phosphate (Glc1P) supported physiologically relevant rates of starch synthesis. Incorporation of Glc1P into starch was both dependent on the presence of ATP and intact organelles. The rate of incorporation of hexose into starch became saturated at a Glc1P concentration of less than 1 mol·m-3 in the presence of 1 mol·m-3 ATP. Starch synthesis from 5 mol · m-3 ADP-glucose supplied to the organelles occurred at rates 15-fold higher than from similar concentrations of Glc1P, but it is argued that this is probably of little physiological relevance. The net incorporation of hexose units into starch from GlclP was inhibited 50% by 100 mmol.m-3 carboxyatractyloside. Carbohydrate oxidation in the Amyloplast was stimulated by the addition of 2-oxoglutarate and glutamine, and in such circumstances incorporation of14C-labelled metabolites into starch was reduced. Glucose 6-phosphate proved to be a better substrate for oxidative pathways than Glc1P. Our results suggest that Glc1P is the primary substrate for starch synthesis in developing wheat endosperm, and that ATP required for starch synthesis is imported via an adenylate translocator.
Masao Tasaka - One of the best experts on this subject based on the ideXlab platform.
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live cell imaging of cytoskeletal and organelle dynamics in gravity sensing cells in plant gravitropism
Methods of Molecular Biology, 2015Co-Authors: Moritaka Nakamura, Masao Tasaka, Masatsugu Toyota, Miyo Terao MoritaAbstract:: Plants sense gravity and change their morphology/growth direction accordingly (gravitropism). The early process of gravitropism, gravity sensing, is supposed to be triggered by sedimentation of starch-filled plastids (Amyloplasts) in statocytes such as root columella cells and shoot endodermal cells. For several decades, many scientists have focused on characterizing the role of the Amyloplasts and observed their intracellular sedimentation in various plants. Recently, it has been discovered that the complex sedimentary movements of the Amyloplasts are created not only by gravity but also by cytoskeletal/organelle dynamics, such as those of actin filaments and the vacuolar membrane. Thus, to understand how plants sense gravity, we need to analyze both Amyloplast movements and their regulatory systems in statocytes. We have developed a vertical-stage confocal microscope that allows multicolor fluorescence imaging of Amyloplasts, actin filaments and vacuolar membranes in vertically oriented plant tissues. We also developed a centrifuge microscope that allows bright-field imaging of Amyloplasts during centrifugation. These microscope systems provide new insights into gravity-sensing mechanisms in Arabidopsis.
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a unique heat repeat containing protein shoot gravitropism6 is involved in vacuolar membrane dynamics in gravity sensing cells of arabidopsis inflorescence stem
Plant and Cell Physiology, 2014Co-Authors: Yasuko Hashiguchi, Takehide Kato, Daisuke Yano, Kiyoshi Nagafusa, Masao Tasaka, Chieko Saito, Akihiko Nakano, Takashi Ueda, Tomohiro Uemura, Miyo Terao MoritaAbstract:Plant vacuoles play critical roles in development, growth and stress responses. In mature cells, vacuolar membranes (VMs) display several types of structures, which are formed by invagination and folding of VMs into the lumenal side and can gradually move and change shape. Although such VM structures are observed in a broad range of tissue types and plant species, the molecular mechanism underlying their formation and maintenance remains unclear. Here, we report that a novel HEAT-repeat protein, SHOOT GRAVITROPISM6 (SGR6), of Arabidopsis is involved in the control of morphological changes and dynamics of VM structures in endodermal cells, which are the gravity-sensing cells in shoots. SGR6 is a membrane-associated protein that is mainly localized to the VM in stem endodermal cells. The sgr6 mutant stem exhibits a reduced gravitropic response. Higher plants utilize Amyloplast sedimentation as a means to sense gravity direction. Amyloplasts are surrounded by VMs in Arabidopsis endodermal cells, and the flexible and dynamic structure of VMs is important for Amyloplast sedimentation. We demonstrated that such dynamic features of VMs are gradually lost in sgr6 endodermal cells during a 30 min observation period. Histological analysis revealed that Amyloplast sedimentation was impaired in sgr6. Detailed live-cell imaging analyses revealed that the VM structures in sgr6 had severe defects in morphological changes and dynamics. Our results suggest that SGR6 is a novel protein involved in the formation and/or maintenance of invaginated VM structures in gravity-sensing cells.
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mechanism of higher plant gravity sensing
American Journal of Botany, 2013Co-Authors: Yasuko Hashiguchi, Masao Tasaka, Miyo Terao MoritaAbstract:Higher plants have developed statocytes, specialized tissues or cells for gravity sensing, and subsequent signal formation. Root and shoot statocytes commonly harbor a number of Amyloplasts, and Amyloplast sedimentation in the direction of gravity is a critical process in gravity sensing. However, the molecular mechanism underlying Amyloplast-dependent gravity sensing is largely unknown. In this review, we mainly describe the molecular basis for the gravity sensing mechanism, i.e., the molecules and their functions involved in Amyloplast sedimentation. Several analyses of statocyte images in living plant organs have implied differences in the regulation of Amyloplast movements between root and shoot statocytes. Amyloplasts in shoot statocytes display not only sedimentable but upward, saltatory movements, but the latter are rarely observed in root statocytes. A series of genetic studies on shoot gravitropism mutants of Arabidopsis thaliana has revealed that two intracellular components, the vacuolar membrane (VM) and actin microfilaments (AFs), within the shoot statocyte play important roles in Amyloplast dynamics. Flexible VM structures surrounding the Amyloplasts seem to allow them to freely sediment toward the bottom of cells. In contrast, long actin cables mediate the saltatory movements of Amyloplasts. Thus, Amyloplasts in shoot statocytes undergo a dynamic equilibrium of movement, and a proper intracellular environment for statocytes is essential for normal shoot gravitropism. Further analyses to identify the molecular regulators of Amyloplast dynamics, including sedimentation, may contribute to an understanding of the gravity sensing mechanism in higher plants.
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an arabidopsis e3 ligase shoot gravitropism9 modulates the interaction between statoliths and f actin in gravity sensing
The Plant Cell, 2011Co-Authors: Moritaka Nakamura, Masao Tasaka, Masatsugu Toyota, Miyo Terao MoritaAbstract:Higher plants use the sedimentation of Amyloplasts in statocytes as statolith to sense the direction of gravity during gravitropism. In Arabidopsis thaliana inflorescence stem statocyte, Amyloplasts are in complex movement; some show jumping-like saltatory movement and some tend to sediment toward the gravity direction. Here, we report that a RING-type E3 ligase SHOOT GRAVITROPISM9 (SGR9) localized to Amyloplasts modulates Amyloplast dynamics. In the sgr9 mutant, which exhibits reduced gravitropism, Amyloplasts did not sediment but exhibited increased saltatory movement. Amyloplasts sometimes formed a cluster that is abnormally entangled with actin filaments (AFs) in sgr9. By contrast, in the fiz1 mutant, an ACT8 semidominant mutant that induces fragmentation of AFs, Amyloplasts, lost saltatory movement and sedimented with nearly statically. Both treatment with Latrunculin B, an inhibitor of AF polymerization, and the fiz1 mutation rescued the gravitropic defect of sgr9. In addition, fiz1 decreased saltatory movement and induced Amyloplast sedimentation even in sgr9. Our results suggest that Amyloplasts are in equilibrium between sedimentation and saltatory movement in wild-type endodermal cells. Furthermore, this equilibrium is the result of the interaction between Amyloplasts and AFs modulated by the SGR9. SGR9 may promote detachment of Amyloplasts from AFs, allowing the Amyloplasts to sediment in the AFs-dependent equilibrium of Amyloplast dynamics.
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a c2h2 type zinc finger protein sgr5 is involved in early events of gravitropism in arabidopsis inflorescence stems
Plant Journal, 2006Co-Authors: Miyo Terao Morita, Takehide Kato, Keitaro Sakaguchi, Shinichiro Kiyose, Kensuke Taira, Moritaka Nakamura, Masao TasakaAbstract:: Plants can sense the direction of gravity and change the growth orientation of their organs. To elucidate the molecular mechanisms of gravity perception and the signal transduction of gravitropism, we have characterized a number of shoot gravitropism (sgr) mutants of Arabidopsis. The sgr5-1 mutant shows reduced gravitropism in the inflorescence stem but its root and hypocotyl have normal gravitropism. SGR5 encodes a zinc finger protein with a coiled-coil motif. The SGR5-GFP fusion protein is localized in the nucleus of Arabidopsis protoplasts, suggesting that SGR5 may act as a transcription factor. Analysis of GUS expression under the control of the SGR5 promoter revealed that SGR5 is mainly expressed in the endodermis, the gravity-sensing tissue in inflorescence stems. Furthermore, the observation that endodermis-specific expression of SGR5 using the SCR promoter in the sgr5-1 mutant restores shoot gravitropism indicates that it could function in the gravity-sensing endodermal cell layer. In contrast to other sgr mutants reported previously, almost all Amyloplasts in the endodermal cells of the sgr5-1 mutant sedimented in the direction of gravity. Taken together, our results suggest that SGR5 may be involved in an early event in shoot gravitropism such as gravity perception and/or a signaling process subsequent to Amyloplast sedimentation as a putative transcription factor in gravity-perceptive cells.
Bob B. Buchanan - One of the best experts on this subject based on the ideXlab platform.
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proteome of Amyloplasts isolated from developing wheat endosperm presents evidence of broad metabolic capability
Journal of Experimental Botany, 2006Co-Authors: Yves Balmer, William H. Vensel, Bob B. Buchanan, Frances M. Dupont, William J HurkmanAbstract:By contrast to chloroplasts, our knowledge of Amyloplasts—organelles that synthesize and store starch in heterotrophic plant tissues—is in a formative stage. While our understanding of what is considered their primary function, i.e. the biosynthesis and degradation of starch, has increased dramatically in recent years, relatively little is known about other biochemical processes taking place in these organelles. To help fill this gap, a proteomic analysis of Amyloplasts isolated from the starchy endosperm of wheat seeds (10 d postanthesis) has been conducted. The study has led to the identification of 289 proteins that function in a range of processes, including carbohydrate metabolism, cytoskeleton/plastid division, energetics, nitrogen and sulphur metabolism, nucleic acid-related reactions, synthesis of various building blocks, proteinrelated reactions, transport, signalling, stress, and a variety of other activities grouped under ‘miscellaneous’. The function of 12% of the proteins was unknown. The results highlight the role of the Amyloplast as a starch-storing organelle that fulfills a spectrum of biosynthetic needs of the parent tissue. When compared with a recent proteomic analysis of whole endosperm, the current study demonstrates the advantage of using isolated organelles in proteomic studies.
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a complete ferredoxin thioredoxin system regulates fundamental processes in Amyloplasts
Proceedings of the National Academy of Sciences of the United States of America, 2006Co-Authors: Yves Balmer, Wanda Manieri, William H. Vensel, Peter Schürmann, William J Hurkman, Nick Cai, Bob B. BuchananAbstract:†‡ A growing number of processes throughout biology are regulated by redox via thiol– disulfide exchange. This mechanism is particularly widespread in plants, where almost 200 proteins have been linked to thioredoxin (Trx), a widely distributed small regulatory disulfide protein. The current study extends regulation by Trx to Amyloplasts, organelles prevalent in heterotrophic plant tissues that, among other biosynthetic activities, catalyze the synthesis and storage of copious amounts of starch. Using proteomics and immunological methods, we identified the components of the ferredoxinTrx system (ferredoxin, ferredoxin–Trx reductase, and Trx), originally described for chloroplasts, in Amyloplasts isolated from wheat starchy endosperm. Ferredoxin is reduced not by light, as in chloroplasts, but by metabolically generated NADPH via ferredoxin–NADP reductase. However, once reduced, ferredoxin appears to act as established for chloroplasts, i.e., via ferredoxin– Trx reductase and a Trx (m-type). A proteomics approach in combination with affinity chromatography and a fluorescent thiol probe led to the identification of 42 potential Trx target proteins, 13 not previously recognized, including a major membrane transporter (Brittle-1 or ADP-glucose transporter). The proteins function in a range of processes in addition to starch metabolism: biosynthesis of lipids, amino acids, and nucleotides; protein folding; and several miscellaneous reactions. The results suggest a mechanism whereby light is initially recognized as a thiol signal in chloroplasts, then as a sugar during transit to the sink, where it is converted again to a thiol signal. In this way, Amyloplast reactions in the grain can be coordinated with photosynthesis taking place in leaves. redox regulation target proteins ferredoxin–thioredoxin reductase
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A complete ferredoxin/thioredoxin system regulates fundamental processes in Amyloplasts
Proceedings of the National Academy of Sciences of the United States of America, 2006Co-Authors: Yves Balmer, Wanda Manieri, William H. Vensel, Peter Schürmann, William J Hurkman, Nick Cai, Bob B. BuchananAbstract:A growing number of processes throughout biology are regulated by redox via thiol-disulfide exchange. This mechanism is particularly widespread in plants, where almost 200 proteins have been linked to thioredoxin (Trx), a widely distributed small regulatory disulfide protein. The current study extends regulation by Trx to Amyloplasts, organelles prevalent in heterotrophic plant tissues that, among other biosynthetic activities, catalyze the synthesis and storage of copious amounts of starch. Using proteomics and immunological methods, we identified the components of the ferredoxin/Trx system (ferredoxin, ferredoxin-Trx reductase, and Trx), originally described for chloroplasts, in Amyloplasts isolated from wheat starchy endosperm. Ferredoxin is reduced not by light, as in chloroplasts, but by metabolically generated NADPH via ferredoxin-NADP reductase. However, once reduced, ferredoxin appears to act as established for chloroplasts, i.e., via ferredoxin-Trx reductase and a Trx (m-type). A proteomics approach in combination with affinity chromatography and a fluorescent thiol probe led to the identification of 42 potential Trx target proteins, 13 not previously recognized, including a major membrane transporter (Brittle-1 or ADP-glucose transporter). The proteins function in a range of processes in addition to starch metabolism: biosynthesis of lipids, amino acids, and nucleotides; protein folding; and several miscellaneous reactions. The results suggest a mechanism whereby light is initially recognized as a thiol signal in chloroplasts, then as a sugar during transit to the sink, where it is converted again to a thiol signal. In this way, Amyloplast reactions in the grain can be coordinated with photosynthesis taking place in leaves.