Intramolecular carbon isotope signals reflect metabolite allocation in plants

Abstract

- Stable isotopes at natural abundance are key tools to study physiological processes occurring outside the temporal scope of manipulation and monitoring experiments. Whole-molecule carbon isotope ratios (13C/12C) enable assessments of plant carbon uptake yet conceal information about carbon allocation. Here, we identify an intramolecular 13C/12C signal at tree-ring glucose C-5 and C-6, develop experimentally testable theories on its origin, and test these theories. - First, we assess the potential of processes within C3 metabolism for signal introduction based (inter alia) on constraints on signal propagation posed by metabolic networks. Second, we support our theoretical framework by re-analysing published 13C/12C data and modelling signal-environment relationships. - We propose the intramolecular signal reports carbon allocation into major metabolic pathways in actively photosynthesising leaf cells including the anaplerotic, shikimate, and non-mevalonate pathway. This is supported by previously reported whole-molecule 13C/12C increases in cellulose of ozone-treated Betula pendula and a highly significant relationship between the intramolecular signal and tropospheric ozone concentration. Additionally, we postulate a pronounced preference of leaf-cytosolic triose-phosphate isomerase to catalyse the forward reaction in vivo (dihydroxyacetone phosphate to glyceraldehyde 3-phosphate). - In conclusion, intramolecular 13C/12C analysis resolves information about carbon uptake and allocation enabling more comprehensive assessments of carbon metabolism than whole-molecule 13C/12C analysis.

Metabolism is a major driver of hydrogen isotope fractionation recorded in tree-ring glucose of Pinus nigra

“…The present paper and recent studies on intramolecular 13 C/ 12 C signals (Wieloch et al, 2018(Wieloch et al, , 2021a show that there is much room for improvement. Thus, we recommend pushing the development of intramolecular and dual-isotope methodology.…”

“…In principle, whole-molecule stable carbon isotope ratios of plant organic matter, δ 13 C, enable Ci estimations because 13 C fractionation during carbon uptake is related to Ci/Ca (Farquhar et al, 1982;Evans et al, 1986). However, processes that are independent of carbon uptake fractionation exert strong control over δ 13 C values of the samples studied here (Wieloch et al, 2018(Wieloch et al, , 2021a. Therefore, we decided against using δ 13 Cderived Ci estimates to test this hypothesis.…”

Carbon flux around leaf-cytosolic glyceraldehyde-3-phosphate dehydrogenase introduces a 13C signal in plant glucose

“…These signals cover the period 1961–1995 and are expressed in terms of intramolecular 13 C discrimination, Δ i ′ , where i denotes glucose C–H positions ( Wieloch et al , 2018 ). In this notation, positive values denote discrimination against 13 C. Triose phosphate cycling in tree-ring cells causes fractional equilibration of 13 C between glucose C-1 and C-6, C-2 and C-5, and C-3 and C-4 (see SI 2 in Wieloch et al , 2021, Preprint ). The prime marks data corrected for heterotrophic triose phosphate cycling ( Wieloch et al , 2018 ).…”

“…Due to the substitution of the unknown R GAP by R VPDB (see above), Δ GAP′ develops from zero and Δ 3PGA develops towards zero; that is, the model exclusively returns the 13 C discrimination of the system of interest. Leaf-cytosolic 3PGA is partly reimported into chloroplasts, and chloroplast metabolism can be expected to transfer part of the GAPDH fractionation signal to other glucose carbon positions (see SI 1 in Wieloch et al , 2021, Preprint ). However, these effects are small and were neglected here for reasons of simplicity.…”

“…Both the commitment of GAP to 3PGA versus FBP metabolism and the contribution of np-GAPDH versus p-GAPDH to catalysing the GAP to 3PGA forward conversion may alter the 13 C/ 12 C ratio at tree-ring glucose C-4 and respond to environmental cues. In vivo , the reaction catalysed by cytosolic TPI is strongly displaced from equilibrium on the side of GAP ( Wieloch et al , 2021, Preprint ). That is, GAP and DHAP are not in isotopic equilibrium because cytosolic flux from GAP to DHAP is negligible.…”