The oxidative pentose phosphate(OPP)pathway provides metabolic intermediates for the shikimate pathway and directs carbon flow to the biosynthesis of aromatic amino acids(AAAs),which serve as basic protein building bl...The oxidative pentose phosphate(OPP)pathway provides metabolic intermediates for the shikimate pathway and directs carbon flow to the biosynthesis of aromatic amino acids(AAAs),which serve as basic protein building blocks and precursors of numerous metabolites essential for plant growth.However,genetic evidence linking the two pathways is largely unclear.In this study,we identified 6-phosphogluconate dehydrogenase 2(PGD2),the rate-limiting enzyme of the cytosolic OPP pathway,through suppressor screening of arogenate dehydrogenase 2(adh2)in Arabidopsis.Our data indicated that a single amino acid substitution at position 63(glutamic acid to lysine)of PGD2 enhanced its enzyme activity by facilitating the dissociation of products from the active site of PGD2,thus increasing the accumulation of AAAs and partially restoring the defective phenotype of adh2.Phylogenetic analysis indicated that the point mutation occurred in a well-conserved amino acid residue.Plants with different amino acids at this conserved site of PGDs confer diverse catalytic activities,thus exhibiting distinct AAAs producing capability.These findings uncover the genetic link between the OPP pathway and AAAs biosynthesis through PGD2.The gain-of-function point mutation of PGD2 identified here could be considered as a potential engineering target to alter the metabolic flux for the production of AAAs and downstream compounds.展开更多
Glucose-6-phosphate dehydrogenase （G6PDH, E.C. 1.1.1.49） and 6-phosphogluconate dehydrogenase （6PGDH, EC 1.1.1.44） are both key enzymes of the pentose phosphate pathway （PPP）. The OsG6PDH1 and Os6PGDH1 genes enc...Glucose-6-phosphate dehydrogenase （G6PDH, E.C. 1.1.1.49） and 6-phosphogluconate dehydrogenase （6PGDH, EC 1.1.1.44） are both key enzymes of the pentose phosphate pathway （PPP）. The OsG6PDH1 and Os6PGDH1 genes encoding cytosolic G6PDH and cytosolic 6PGDH were Isolated from rice （Oryza sativa L.）. We have shown that Os6PGDH1 gene was up-regulated by salt stress. Here we reported the isolation and characterization of Os6PGDH2 from rice, which encode the plastidic counterpart of 6PGDH. Genomic organization analysis indicated that OsG6PDH1 and OsG6PDH2 genes contain multiple introns, whereas two Os6PGDH1 and Os6PGDH2 genes have no introns in their translated regions. In a step towards understanding the functions of the pentose phosphate pathway in plants in response to various abiotic stresses, the expressions of four genes in the rice seedlings treated by drought, cold, high salinity and abscisic acid （ABA） were investigated. The results show that OsG6PDH1 and OsG6PDH2 are not markedly regulated by the abiotic stresses detected. However, the transcript levels of both Os6PGDH1 and Os6PGDH2 are up-regulated in rice seedlings under drought, cold, high salinity and ABA treatments. Meanwhile, the enzyme activities of G6PDH and 6PGDH in the rice seedlings treated by various abiotic stresses were Investigated. Like the mRNA expression patterns, G6PDH activity remains constant but the 6PGDH Increases steadily during the treatments. Taken together, we suggest that the pentose phosphate pathway may play an important role in rice responses to abiotic stresses and the second key enzyme of PPP, 6PGDH, may function as a regulator controlling the efficiency of the pathway under abiotic stresses.展开更多
基金supported by the National Key Research and Development Program of China(2019YFA0903900)the National Natural Science Foundation of China(32300233)+1 种基金Guangdong Provincial Key Laboratory of Synthetic Genomics(2023B1212060054)Shenzhen Key Laboratory of Synthetic Genomics(ZDSYS201802061806209).
文摘The oxidative pentose phosphate(OPP)pathway provides metabolic intermediates for the shikimate pathway and directs carbon flow to the biosynthesis of aromatic amino acids(AAAs),which serve as basic protein building blocks and precursors of numerous metabolites essential for plant growth.However,genetic evidence linking the two pathways is largely unclear.In this study,we identified 6-phosphogluconate dehydrogenase 2(PGD2),the rate-limiting enzyme of the cytosolic OPP pathway,through suppressor screening of arogenate dehydrogenase 2(adh2)in Arabidopsis.Our data indicated that a single amino acid substitution at position 63(glutamic acid to lysine)of PGD2 enhanced its enzyme activity by facilitating the dissociation of products from the active site of PGD2,thus increasing the accumulation of AAAs and partially restoring the defective phenotype of adh2.Phylogenetic analysis indicated that the point mutation occurred in a well-conserved amino acid residue.Plants with different amino acids at this conserved site of PGDs confer diverse catalytic activities,thus exhibiting distinct AAAs producing capability.These findings uncover the genetic link between the OPP pathway and AAAs biosynthesis through PGD2.The gain-of-function point mutation of PGD2 identified here could be considered as a potential engineering target to alter the metabolic flux for the production of AAAs and downstream compounds.
基金Suported by the Ph.D Foundation Program from the Ministry of Education （20050307013）, the National Natural Science Foundation of China （30470921）, the Jiangsu Provincial Natural Science Foundation of China （BK2005090） and the Changjiang Scholars and Innovative Research Team in University （PCSIRT）.The nucleotide sequences reported in this paper have been deposited in GenBank under the accession numbers AY078072, AY339367, AF486280 and AY278362, respectively,
文摘Glucose-6-phosphate dehydrogenase （G6PDH, E.C. 1.1.1.49） and 6-phosphogluconate dehydrogenase （6PGDH, EC 1.1.1.44） are both key enzymes of the pentose phosphate pathway （PPP）. The OsG6PDH1 and Os6PGDH1 genes encoding cytosolic G6PDH and cytosolic 6PGDH were Isolated from rice （Oryza sativa L.）. We have shown that Os6PGDH1 gene was up-regulated by salt stress. Here we reported the isolation and characterization of Os6PGDH2 from rice, which encode the plastidic counterpart of 6PGDH. Genomic organization analysis indicated that OsG6PDH1 and OsG6PDH2 genes contain multiple introns, whereas two Os6PGDH1 and Os6PGDH2 genes have no introns in their translated regions. In a step towards understanding the functions of the pentose phosphate pathway in plants in response to various abiotic stresses, the expressions of four genes in the rice seedlings treated by drought, cold, high salinity and abscisic acid （ABA） were investigated. The results show that OsG6PDH1 and OsG6PDH2 are not markedly regulated by the abiotic stresses detected. However, the transcript levels of both Os6PGDH1 and Os6PGDH2 are up-regulated in rice seedlings under drought, cold, high salinity and ABA treatments. Meanwhile, the enzyme activities of G6PDH and 6PGDH in the rice seedlings treated by various abiotic stresses were Investigated. Like the mRNA expression patterns, G6PDH activity remains constant but the 6PGDH Increases steadily during the treatments. Taken together, we suggest that the pentose phosphate pathway may play an important role in rice responses to abiotic stresses and the second key enzyme of PPP, 6PGDH, may function as a regulator controlling the efficiency of the pathway under abiotic stresses.
