A large decrease of cytosolic triosephosphate isomerase in transgenic potato roots affects the distribution of carbon in primary metabolism.
Triosephosphate isomerase (TPI, EC 5.3.1.1) catalyzes the interconversion of dihydroxyacetone-P and glyceraldehyde 3-P in the glycolytic pathway. A constitutively expressed antisense construct for cytosolic TPI was introduced into potato (Solanum tuberosum) using Agrobacterium rhizogenes to examine the metabolic effects of a reduction in cytosolic TPI in roots. We obtained a population of transgenic root clones displaying ~36 to 100 % of the TPI activity found in control clones carrying an empty binary vector. Ion exchange chromatography and immunoblot analysis showed that the antisense strategy significantly decreased the cytosolic TPI isoform, while levels of plastidial TPI activity remained apparently unaffected. Transgenic roots were characterized with respect to the activity of glycolytic enzymes, their metabolite contents and carbon fluxes. Metabolite profiling of sugars, organic acids, amino acids and lipids showed elevated levels of sucrose, glucose, fructose, fumarate, isocitrate, 4-aminobutyrate, alanine, glycine, aromatic amino acids and saturated long chain fatty acids in roots containing the lowest TPI activity. Labelings with (14)C-glucose, (14)C-sucrose and (14)C-acetate indicated that a reduction of cytosolic TPI activity in roots increased carbon metabolism through the pentose phosphate pathway, O(2) uptake and catabolism of sucrose to CO(2), and capacity for lipid synthesis. These results demonstrate that a large reduction of cytosolic TPI alters the distribution of carbon in plant primary metabolism.
IRBV, Université de Montréal, 4101 rue Sherbrooke est, Montréal, QC H1X 2B2, Canada.
Sonia
S
Dorion
Audrey
A
Clendenning
Julie
J
Jeukens
Joaquín J
JJ
Salas
Nanhi
N
Parveen
Andrea A
AA
Haner
R David
RD
Law
Enrique Martínez
EM
Force
Jean
J
Rivoal
D000596:Amino Acids / Q000032:analysis / Q000378:metabolism; D001692:Biological Transport; D050260:Carbohydrate Metabolism; D002241:Carbohydrates / Q000032:analysis; D002244:Carbon / Q000378:metabolism*; D002250:Carbon Radioisotopes / Q000032:analysis; D002264:Carboxylic Acids / Q000032:analysis / Q000378:metabolism; D003600:Cytosol / Q000201:enzymology; D006019:Glycolysis; D007527:Isoenzymes; D050356:Lipid Metabolism; D008055:Lipids / Q000032:analysis; D009711:Nucleotides / Q000032:analysis / Q000378:metabolism; D010100:Oxygen / Q000378:metabolism*; D018517:Plant Roots / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D030821:Plants, Genetically Modified / Q000378:metabolism; D016372:RNA, Antisense / Q000235:genetics; D018749:RNA, Plant / Q000235:genetics; D011198:Solanum tuberosum / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D014305:Triose-Phosphate Isomerase / Q000235:genetics / Q000378:metabolism*
D000596:Amino Acids; D002241:Carbohydrates; D002250:Carbon Radioisotopes; D002264:Carboxylic Acids; D007527:Isoenzymes; D008055:Lipids; D009711:Nucleotides; D016372:RNA, Antisense; D018749:RNA, Plant; D002244:Carbon; D014305:Triose-Phosphate Isomerase; D010100:Oxygen