The role of GDP-L-galactose phosphorylase in the control of
ascorbate biosynthesis

J 2021

The role of GDP-L-galactose phosphorylase in the control of ascorbate biosynthesis

FENECH, M., V. AMORIM-SILVA, A.E. DEL VALLE, Dominique ARNAUD, N. RUIZ-LOPEZ et. al.

Základní údaje

Originální název

The role of GDP-L-galactose phosphorylase in the control of ascorbate biosynthesis

Autoři

FENECH, M., V. AMORIM-SILVA, A.E. DEL VALLE, Dominique ARNAUD (250 Francie, garant, domácí), N. RUIZ-LOPEZ, A.G. CASTILLO, N. SMIRNOFF a M.A. BOTELLA

Vydání

Plant Physiology, ROCKVILLE, American Society of Plant Physiologists, 2021, 0032-0889

Další údaje

Jazyk

angličtina

Typ výsledku

Článek v odborném periodiku

Obor

10611 Plant sciences, botany

Stát vydavatele

Spojené státy

Utajení

není předmětem státního či obchodního tajemství

Odkazy

URL

Impakt faktor

Impact factor: 8.005

Kód RIV

RIV/00216224:14740/21:00124197

Organizační jednotka

Středoevropský technologický institut

DOI

http://dx.doi.org/10.1093/plphys/kiab010

UT WoS

000649356200029

Klíčová slova anglicky

Arabidopsis; Ascorbic Acid; Galactose; Gene Expression Regulation; Plant; Genetic Variation; Genotype; Guanosine Diphosphate; Mutation; Phosphorylases; Genetically Modified; Tobacco

Štítky

rivok

Příznaky

Mezinárodní význam, Recenzováno

Změněno: 16. 2. 2022 14:46, Mgr. Pavla Foltynová, Ph.D.

Anotace

V originále

The enzymes involved in l-ascorbate biosynthesis in photosynthetic organisms (the Smirnoff-Wheeler [SW] pathway) are well established. Here, we analyzed their subcellular localizations and potential physical interactions and assessed their role in the control of ascorbate synthesis. Transient expression of C terminal-tagged fusions of SW genes in Nicotiana benthamiana and Arabidopsis thaliana mutants complemented with genomic constructs showed that while GDP-D-mannose epimerase is cytosolic, all the enzymes from GDP-D-mannose pyrophosphorylase (GMP) to L-galactose dehydrogenase (L-GalDH) show a dual cytosolic/nuclear localization. All transgenic lines expressing functional SW protein green fluorescent protein fusions driven by their endogenous promoters showed a high accumulation of the fusion proteins, with the exception of those lines expressing GDP-L-galactose phosphorylase (GGP) protein, which had very low abundance. Transient expression of individual or combinations of SW pathway enzymes in N. benthamiana only increased ascorbate concentration if GGP was included. Although we did not detect direct interaction between the different enzymes of the pathway using yeast-two hybrid analysis, consecutive SW enzymes, as well as the first and last enzymes (GMP and L-GalDH) associated in coimmunoprecipitation studies. This association was supported by gel filtration chromatography, showing the presence of SW proteins in high-molecular weight fractions. Finally, metabolic control analysis incorporating known kinetic characteristics showed that previously reported feedback repression at the GGP step, combined with its relatively low abundance, confers a high-flux control coefficient and rationalizes why manipulation of other enzymes has little effect on ascorbate concentration.

Citovat

FENECH, M., V. AMORIM-SILVA, A.E. DEL VALLE, Dominique ARNAUD, N. RUIZ-LOPEZ, A.G. CASTILLO, N. SMIRNOFF a M.A. BOTELLA. The role of GDP-L-galactose phosphorylase in the control of ascorbate biosynthesis. Plant Physiology. ROCKVILLE: American Society of Plant Physiologists, 2021, roč. 185, č. 4, s. 1574-1594. ISSN 0032-0889. Dostupné z: https://dx.doi.org/10.1093/plphys/kiab010.

@article{1834787,
   author = {Fenech, M. and AmorimandSilva, V. and del Valle, A.E. and Arnaud, Dominique and RuizandLopez, N. and Castillo, A.G. and Smirnoff, N. and Botella, M.A.},
   article_location = {ROCKVILLE},
   article_number = {4},
   doi = {http://dx.doi.org/10.1093/plphys/kiab010},
   keywords = {Arabidopsis; Ascorbic Acid; Galactose; Gene Expression Regulation; Plant; Genetic Variation; Genotype; Guanosine Diphosphate; Mutation; Phosphorylases; Genetically Modified; Tobacco},
   language = {eng},
   issn = {0032-0889},
   journal = {Plant Physiology},
   title = {The role of GDP-L-galactose phosphorylase in the control of ascorbate biosynthesis},
   url = {https://academic.oup.com/plphys/article/185/4/1574/6119655},
   volume = {185},
   year = {2021}
}

TY  - JOUR
ID  - 1834787
AU  - Fenech, M. - Amorim-Silva, V. - del Valle, A.E. - Arnaud, Dominique - Ruiz-Lopez, N. - Castillo, A.G. - Smirnoff, N. - Botella, M.A.
PY  - 2021
TI  - The role of GDP-L-galactose phosphorylase in the control of ascorbate biosynthesis
JF  - Plant Physiology
VL  - 185
IS  - 4
SP  - 1574-1594
EP  - 1574-1594
PB  - American Society of Plant Physiologists
SN  - 00320889
KW  - Arabidopsis
KW  - Ascorbic Acid
KW  - Galactose
KW  - Gene Expression Regulation
KW  - Plant
KW  - Genetic Variation
KW  - Genotype
KW  - Guanosine Diphosphate
KW  - Mutation
KW  - Phosphorylases
KW  - Genetically Modified
KW  - Tobacco
UR  - https://academic.oup.com/plphys/article/185/4/1574/6119655
N2  - The enzymes involved in l-ascorbate biosynthesis in photosynthetic organisms (the Smirnoff-Wheeler [SW] pathway) are well established. Here, we analyzed their subcellular localizations and potential physical interactions and assessed their role in the control of ascorbate synthesis. Transient expression of C terminal-tagged fusions of SW genes in Nicotiana benthamiana and Arabidopsis thaliana mutants complemented with genomic constructs showed that while GDP-D-mannose epimerase is cytosolic, all the enzymes from GDP-D-mannose pyrophosphorylase (GMP) to L-galactose dehydrogenase (L-GalDH) show a dual cytosolic/nuclear localization. All transgenic lines expressing functional SW protein green fluorescent protein fusions driven by their endogenous promoters showed a high accumulation of the fusion proteins, with the exception of those lines expressing GDP-L-galactose phosphorylase (GGP) protein, which had very low abundance. Transient expression of individual or combinations of SW pathway enzymes in N. benthamiana only increased ascorbate concentration if GGP was included. Although we did not detect direct interaction between the different enzymes of the pathway using yeast-two hybrid analysis, consecutive SW enzymes, as well as the first and last enzymes (GMP and L-GalDH) associated in coimmunoprecipitation studies. This association was supported by gel filtration chromatography, showing the presence of SW proteins in high-molecular weight fractions. Finally, metabolic control analysis incorporating known kinetic characteristics showed that previously reported feedback repression at the GGP step, combined with its relatively low abundance, confers a high-flux control coefficient and rationalizes why manipulation of other enzymes has little effect on ascorbate concentration.
ER  -

FENECH, M., V. AMORIM-SILVA, A.E. DEL VALLE, Dominique ARNAUD, N. RUIZ-LOPEZ, A.G. CASTILLO, N. SMIRNOFF a M.A. BOTELLA. The role of GDP-L-galactose phosphorylase in the control of ascorbate biosynthesis. \textit{Plant Physiology}. ROCKVILLE: American Society of Plant Physiologists, 2021, roč.~185, č.~4, s.~1574-1594. ISSN~0032-0889. Dostupné z: https://dx.doi.org/10.1093/plphys/kiab010.

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