Unraveling delta1-pyrroline-5-carboxylate-proline cycle in plants by uncoupled expression of proline oxidation enzymes.
The two-step oxidation of proline in all eukaryotes is performed at the inner mitochondrial membrane by the consecutive action of proline dehydrogenase (ProDH) that produces Delta(1)-pyrroline-5-carboxylate (P5C) and P5C dehydrogenase (P5CDH) that oxidizes P5C to glutamate. This catabolic route is down-regulated in plants during osmotic stress, allowing free Pro accumulation. We show here that overexpression of MsProDH in tobacco and Arabidopsis or impairment of P5C oxidation in the Arabidopsis p5cdh mutant did not change the cellular Pro to P5C ratio under ambient and osmotic stress conditions, indicating that P5C excess was reduced to Pro in a mitochondrial-cytosolic cycle. This cycle, involving ProDH and P5C reductase, exists in animal cells and now demonstrated in plants. As a part of the cycle, Pro oxidation by the ProDH-FAD complex delivers electrons to the electron transport chain. Hyperactivity of the cycle, e.g. when an excess of exogenous l-Pro is provided, generates mitochondrial reactive oxygen species (ROS) by delivering electrons to O(2), as demonstrated by the mitochondria-specific MitoSox staining of superoxide ions. Lack of P5CDH activity led to higher ROS production under dark and light conditions in the presence of Pro excess, as well as rendered plants hypersensitive to heat stress. Balancing mitochondrial ROS production during increased Pro oxidation is therefore critical for avoiding Pro-related toxic effects. Hence, normal oxidation of P5C to Glu by P5CDH is key to prevent P5C-Pro intensive cycling and avoid ROS production from electron run-off.
Miller
Gad
G
Department of Plant Science, Tel Aviv University, Tel-Aviv 69978, Israel.
Honig
Arik
A
Stein
Hanan
H
Suzuki
Nobuhiro
N
Mittler
Ron
R
Zilberstein
Aviah
A
D050842:1-Pyrroline-5-Carboxylate Dehydrogenase / Q000235:genetics / Q000378:metabolism*; D017360:Arabidopsis / Q000201:enzymology / Q000235:genetics / Q000378:metabolism; D029681:Arabidopsis Proteins / Q000235:genetics / Q000378:metabolism; D055864:Droughts; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D018506:Gene Expression Regulation, Plant / Q000187:drug effects; D018698:Glutamic Acid / Q000378:metabolism; D006358:Hot Temperature; D000455:Medicago sativa / Q000201:enzymology / Q000235:genetics; D008928:Mitochondria / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism; D009154:Mutation; D010084:Oxidation-Reduction; D010944:Plants / Q000201:enzymology / Q000235:genetics / Q000378:metabolism*; D030821:Plants, Genetically Modified; D011392:Proline / Q000378:metabolism / Q000494:pharmacology; D011394:Proline Oxidase / Q000235:genetics / Q000378:metabolism*; D011758:Pyrroles / Q000378:metabolism*; D017382:Reactive Oxygen Species / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012965:Sodium Chloride / Q000494:pharmacology; D014026:Nicotiana / Q000201:enzymology / Q000235:genetics / Q000378:metabolism
D029681:Arabidopsis Proteins; D011758:Pyrroles; D017382:Reactive Oxygen Species; C015485:delta-1-pyrroline-5-carboxylate; D018698:Glutamic Acid; D012965:Sodium Chloride; D011392:Proline; D050842:1-Pyrroline-5-Carboxylate Dehydrogenase; C508255:ALDH12A1 protein, Arabidopsis; D011394:Proline Oxidase