Biology subjects

Ratcliffe, R. G.

Publications and source records attributed to Ratcliffe, R. G..

4 recordsLinked to original sources

bioRxiv2022

Leaf specific overexpression of a mitochondrially-targeted glutamine synthetase in tomato increased assimilate export resulting in earlier fruiting and elevated yield

To identify potential strategies for increasing the efficiency of tomato leaf metabolism, with a focus on the links between nitrogen/carbon metabolism, we explored a diel Flux Balance Analysis (FBA) model of a source leaf in which the metabolic output was varied up to the theoretically-achievable maximum. We noticed a potentially interesting switch in the use of glutamine synthetase (GS) isoforms -from the chloroplast isoform to the mitochondrial one- for nitrogen assimilation. To further explore this prediction, we characterized transgenic tomato plants over-expressing two tomato GS genes, GS1 and GS2, targeted to mitochondria. Both sets of transgenic plants were characterized as displaying faster growth rate, early flowering and increased fruit yield. In leaves, metabolomic profiling and enzyme activity analysis pointed that GS activity in mitochondrial plays a role in increasing the intracellular synthesis and subsequent export of sugar. Consistent with these changes, higher sucrose concentration in leaf exudates and reduced activities of enzymes involved in leaf starch synthesis were observed. Moreover, mitochondrial GS activity affected chloroplast redox status in a manner that modulated photorespiration and nitrogen metabolism. The combined data reveal the influence of mitochondrial GS activity on both foliar carbon/nitrogen balance and regulation of source-sink metabolism in tomato plants.

plant biology↗

bioRxiv2021

Metabolic constraints on nitrogen fixation by rhizobia in legume nodules

Rhizobia induce nodule formation on legume roots and differentiate into bacteroids, which use plant-derived dicarboxylates as energy and electron sources for reduction of atmospheric N2 into ammonia for secretion to plants. Using heterogeneous genome-scale datasets, we reconstructed a model of bacteroid metabolism to investigate the effects of varying dicarboxylate and oxygen supply on carbon and nitrogen allocation. Modelling and 13C metabolic flux analysis in bacteroids indicate that microaerobiosis restricts the decarboxylating arm of the TCA cycle and limits ammonia assimilation into glutamate. Catabolism of dicarboxylates induces a higher oxygen demand but also a higher NADH/NAD+ ratio compared to sugars. Carbon polymer synthesis and alanine secretion by bacteroids facilitate redox balance in microaerobic nodules with alanine secretion increasing as oxygen tension decreases. Our results provide a framework for understanding fundamental constraints on rhizobial metabolism during symbiotic nitrogen fixation.

microbiology↗

bioRxiv2020

CAM emerges in a leaf metabolic model under water-saving constraints in different environments

Crassulacean Acid Metabolism (CAM) evolved in arid environments as a water-saving alternative to C3 photosynthesis. There is great interest in engineering more drought-resistant crop species by introducing CAM into C3 plants. However, one of the open questions is whether full CAM or alternative water-saving flux modes would be more productive in the environments typically experienced by C3 crops. To study the effect of temperature and relative humidity on plant metabolism we coupled a time-resolved diel model of leaf metabolism to an environment-dependent gas-exchange model. This model allowed us to study the emergence of CAM or CAM-like behaviour as a result of a trade-off between leaf productivity and water-saving. We show that vacuolar storage capacity in the leaf is a major determinant of the extent of CAM and shapes the occurrence of phase II and IV of the CAM cycle. Moreover, the model allows us to study alternative flux routes and we identify mitochondrial isocitrate dehydrogenase (ICDH) and an isocitrate-citrate-proline-2OG cycle as a potential contributor to initial carbon fixation at night. Simulations across a wide range of environmental parameters show that the water-saving potential of CAM strongly depends on the environment and that the additional water-saving effect of carbon fixation by ICDH can reach up to 4% for the conditions tested.

plant biology↗

bioRxiv2019

Predicting metabolism during growth by osmotic cell expansion

Cell expansion is a significant contributor to organ growth and is driven by the accumulation of osmolytes to increase cell turgor pressure. Metabolic modelling has the potential to provide insights into the processes that underpin osmolyte synthesis and transport, but the main computational approach for predicting metabolic network fluxes, flux balance analysis (FBA), typically uses biomass composition as the main output constraint and ignores potential changes in cell volume. Here we present GrOE-FBA (Growth by Osmotic Expansion-Flux Balance Analysis), a framework that accounts for both the metabolic and ionic contributions to the osmotica that drive cell expansion, as well as the synthesis of protein, cell wall and cell membrane components required for cell enlargement. Using GrOE-FBA, the metabolic fluxes in dividing and expanding cell were analyzed, and the energetic costs for metabolite biosynthesis and accumulation in the two scenarios were found to be surprisingly similar. The expansion phase of tomato fruit growth was also modelled using a multi-phase single optimization GrOE-FBA model and this approach gave accurate predictions of the major metabolite levels throughout fruit development as well as revealing a role for transitory starch accumulation in ensuring optimal fruit development.

plant biology↗