Transgenic Introduction of a Glycolate Oxidative Cycle into A. thaliana Chloroplasts Leads to Growth Improvement

Abstract

The photorespiratory pathway helps illuminated C3-plants under conditions of limited CO2 availability by effectively exporting reducing equivalents in form of glycolate out of the chloroplast and regenerating glycerate-3-P as substrate for RubisCO. On the other hand, this pathway is considered as probably futile because previously assimilated CO2 is released in mitochondria. Consequently, a lot of effort has been made to reduce this CO2 loss either by reducing fluxes via engineering RubisCO or circumventing mitochondrial CO2 release by the introduction of new enzyme activities. Here we present an approach following the latter route, introducing a complete glycolate catabolic cycle in chloroplasts of Arabidopsis thaliana comprising glycolate oxidase (GO), malate synthase (MS), and catalase (CAT). Results from plants bearing both GO and MS activities have already been reported (Fahnenstich et al., 2008). This previous work showed that the H2O2 produced by GO had strongly negative effects. These effects can be prevented by introducing a plastidial catalase activity, as reported here. Transgenic lines bearing all three transgenic enzyme activities were identified and some with higher CAT activity showed higher dry weight, higher photosynthetic rates, and changes in glycine/serine ratio compared to the wild type. This indicates that the fine-tuning of transgenic enzyme activities in the chloroplasts seems crucial and strongly suggests that the approach is valid and that it is possible to improve the growth of A. thaliana by introducing a synthetic glycolate oxidative cycle into chloroplasts.

From Endoplasmic Reticulum to Mitochondria: Absence of the Arabidopsis ATP Antiporter Endoplasmic Reticulum Adenylate Transporter1 Perturbs Photorespiration

“…This metabolic process serves to recycle 2-phosphoglycolate, which is unavoidably produced by ribulose-1,5-bis-phosphate carboxylase/oxygenase, into 3-phosphoglycerate but also results in considerable rates of CO 2 release. Because photorespiration was long considered a wasteful process leading to reduced plant growth and crop yield, it is not surprising that several molecular-genetic strategies were pursued to possibly reduce photorespiratory CO 2 losses (Kebeish et al, 2007;Maier et al, 2012). After its discovery in the 1950s, it turned out photorespiration is based on a complex metabolic pathway spread over three cellular compartments: chloroplasts, peroxisomes, and mitochondria (Tolbert, 1997;Foyer et al, 2009).…”

The Benefits of Photorespiratory Bypasses: How Can They Work?

“…Previous studies suggested that this pathway theoretically requires less energy and shifts CO 2 release from mitochondria to chloroplasts (Peterhansel and Maurino, 2011;Peterhansel et al, 2013); experimental results indicated that the bypass allowed for increased net photosynthesis and biomass production in Arabidopsis (Kebeish et al, 2007). There are reports of two other photorespiratory bypass pathways in the literature (Carvalho, 2005;Carvalho et al, 2011;Maier et al, 2012). In the Carvalho bypass (Carvalho, 2005;Carvalho et al, 2011), glyoxylate is converted to hydroxypyruvate in the peroxisome.…”

“…Schematic representation of the C3 photosynthesis kinetic model with three different photorespiratory bypass pathways. The bypass described by Kebeish et al (2007) is indicated in blue, the bypass described by Maier et al (2012) in pink, and the bypass described by Carvalho et al (2011) in green. The original photorespiratory pathway is marked in orange, and CO 2 released from photorespiration (including the original pathway and bypass pathways) is indicated in red.…”

The role of membrane transport in metabolic engineering of plant primary metabolism

“…Possibly, the formation of glycine from glycolate during photorespiration was of selective advantage, which favored the loss of the tatronate semialdehyde route that does not produce glycine as a pathway intermediate. While shortcutting photorespiration has already been demonstrated to lead to increased biomass production at least under short day conditions in engineered Arabidopsis plants in two independent studies [18,19 ], a proof of concept for transforming a C 3 to a C 4 plant is still lacking. As outlined below, the implementation of both, shortcutting photorespiration and transforming C 3 to C 4 , will benefit from a better understanding of intracellular solute transport.…”