S stable isotope discrimination in rice: isotope vs molecular phenotypes

Sulfur has four stable isotopes: 32S, 33S, 34S, and 36S; their percentage abundances are 0.9499, 0.0075, 0.0425, and 0.0001 atom fraction, respectively. Mass differences among the S isotopes result in small but yet significant differences in their chemical and physical properties, which may produce considerable separation of the S isotopes during chemical reactions. The most abundant isotopes - 32S and 34S - are now commonly measured using elemental analyzers coupled with isotope ratio mass spectrometers, and S isotope abundance are generally express in term of 34S/32S abundance ratio, using the standard δ34S notation: δ34S = [(34S/32S)sample/(34S/32S)VCDT -1] x 1000 which express the part per thousand deviation of the isotope ratio 34S/32S of a sample relative to an international standard, the Vienna Canyon Diablo Troilite (VCDT). Unlike what has happened with carbon and nitrogen, natural abundance S stable isotope analysis techniques have scarcely been employed to study S allocation and metabolism in plants mainly because of the lack of knowledge about the 32S/34S isotope effects occurring during S metabolism and partitioning among the different organs. Most of the irreversible reactions involving S discriminate between 32S and 34S by favoring the light isotope (32S), thus enriching in 34S the residual substrate molecules left behind. That is to say, that irreversible reactions that do not consume all the substrate may likely produce a detectable fractionation of S stable isotopes at natural abundance, providing crucial insights in the understanding of S metabolic fluxes inside the plants, thus preventing costly or radioactive labeling. Preliminary analyses performed on rice plants grown in complete nutrient solutions revealed some discrepancies amongst δ34S values of the S source and total S measured in the whole biomass or organs. Total S in the whole plant was significantly depleted in 34S by -1.40 ± 0.08 ‰ with respect to the S source. Moreover, the analysis of organ-specific δ34S values measured for total S revealed that roots were depleted in 34S by -0.52 ± 0.05 ‰ while shoots were enriched by 0.62 ± 0.04 relative to the same S source. Moreover, S stable isotope mass balance studies revealed that different isotope phenotypes can be associated to the preferential expresion of specific sulfate transporters involved in sulfate uptake. Taken as a whole, data strongly indicate that 32S/34S isotope effects occur during sulfate uptake and S partitioning.