David K. Liscombe, Yusuke Kamiyoshihara, Jérémie Ghironzi, Christine J. Kempthorne, Kevin Hooton, Blandine Bulot, Vassili Kanellis, James McNulty, Nghi B. Lam, Louis Félix Nadeau, Michael Pautler, Denise M. Tieman, Harry J. Klee, and Charles Goulet

PNAS February 15, 2022 119 (7) e2118676119;

Significance

Aroma is an important factor in consumer perception and acceptance of fresh tomatoes and involves a cocktail of several dozen compounds. Tomato fruits produce uncommon nitrogen-containing volatiles derived mainly from the amino acids leucine and phenylalanine. These volatiles have strong positive correlations with consumer liking. We show that an enzyme active in ripening tomatoes is responsible for the production of all nitrogenous volatiles in tomato fruit, at the expense of substrates derived from cysteine and volatile aldehydes. This discovery defines a cysteine-dependent route to nitrogenous volatiles in plants, prompting a reconsideration of the impact of sulfur metabolism on tomato flavor and quality.

Abstract

Tomato (Solanum lycopersicum) produces a wide range of volatile chemicals during fruit ripening, generating a distinct aroma and contributing to the overall flavor. Among these volatiles are several aromatic and aliphatic nitrogen-containing compounds for which the biosynthetic pathways are not known. While nitrogenous volatiles are abundant in tomato fruit, their content in fruits of the closely related species of the tomato clade is highly variable. For example, the green-fruited species Solanum pennellii are nearly devoid, while the red-fruited species S. lycopersicum and Solanum pimpinellifolium accumulate high amounts. Using an introgression population derived from S. pennellii, we identified a locus essential for the production of all the detectable nitrogenous volatiles in tomato fruit. Silencing of the underlying gene (SlTNH1;Solyc12g013690) in transgenic plants abolished production of aliphatic and aromatic nitrogenous volatiles in ripe fruit, and metabolomic analysis of these fruit revealed the accumulation of 2-isobutyl-tetrahydrothiazolidine-4-carboxylic acid, a known conjugate of cysteine and 3-methylbutanal. Biosynthetic incorporation of stable isotope-labeled precursors into 2-isobutylthiazole and 2-phenylacetonitrile confirmed that cysteine provides the nitrogen atom for all nitrogenous volatiles in tomato fruit. Nicotiana benthamiana plants expressing SlTNH1 readily transformed synthetic 2-substituted tetrahydrothiazolidine-4-carboxylic acid substrates into a mixture of the corresponding 2-substituted oxime, nitro, and nitrile volatiles. Distinct from other known flavin-dependent monooxygenase enzymes in plants, this tetrahydrothiazolidine-4-carboxylic acid N-hydroxylase catalyzes sequential hydroxylations. Elucidation of this pathway is a major step forward in understanding and ultimately improving tomato flavor quality.

See: https://www.pnas.org/content/119/7/e2118676119

Fig. 1.

A gene on Chr12 (Solyc12g013690) is responsible for nitrogenous volatiles biosynthesis in tomato fruit. (A) Structures of main nitrogenous volatiles detected in tomato fruit. (B) Emissions of nitrogenous volatiles from cut ripe fruits of ILs and the tomato parent, cv. M82 (±SE, n = 6). The amount of each nitrogenous volatile is significantly different between M82 and the ILs (P < 0.01). n.d., not detectable. (C) Fine mapping of a nitrogenous volatile-related QTL on chromosome 12. A set of recombinants with low or high (wild type) content of nitrogenous volatiles was derived from a cross between M82 and IL12-2. The illustration shows a small segment of chromosome 12 where the QTL is located. White and dark-gray sections represent genomic segments from S. lycopersicum and S. pennellii, respectively. Arrows indicate the positions of markers used to define recombinants. The light-gray sections represent the regions where a recombination occurred, as delimited by molecular markers. The color inside the diamonds indicates whether the ILs contain the S. lycopersicum (white) or S. pennellii (dark-gray) version of Solyc12g013690. The QTL region as defined by volatile content is shown by dashed lines and enlarged below to show details (assembly SL2.50).