Effet de la salinité pendant la croissance de plants de riz sur la biosynthèse de la 2-acétyl-1-pyrroline (2AP), composé aromatique clé des variétés de riz aromatique,le rôle de la 1-pyrroline dans la biosynthèse de la 2AP ainsi que l'origine du groupe acétyle ont été étudiés. La variété aromatique de Camargue (France), Aychade, a été cultivée soit dans un milieu liquide soit dans des pots en serre. Dans le milieu de culture liquide, une augmentation significative de la teneur en 2AP dans les feuilles a été observée quelle que soit la concentration en NaCl (de 1,7 à 170 mM). Une corrélation positive a été mise en évidence entre la teneur en sel et en 2AP. A de plus fortes concentrations en NaCl, il a également pu être montré qu'il y avait une relation entre la concentration en proline, un précurseur aminé majeur de la 2AP, et la teneur en 2AP. Inversement, les changements en acide -aminobutyrique (GABA), un des métabolites compétitif de la biosynthèse de la 2AP, n'ont pas été corrélés avec ceux de la 2AP. Dans les expérimentations menées en serre, la variété Aychade a été cultivée dans une solution saline à 30 mM, correspondant à une conductivité électrique de 3800 ± 400 S.cm-1, valeur considérée comme suffisante pour induire un stress salin sur le plant de riz. Cette solution saline a été appliquée à des dates et pour des durées différentes en fonction du cycle de développement de la plante. A la phase végétative, une relation positive a été observée dans les feuilles entre le traitement salin et la teneur en 2AP ainsi qu'en proline. Il est intéressant de noter que tous les grains provenant des plants traités avec la solution saline contenaient des teneurs nettement plus élevées en 2AP par rapport au contrôle. La synthèse de la 2AP la plus importante s'est produite lorsque les plants ont été soumis à un traitement salin pendant l'ensemble des phases végétative et reproductive. Cependant, dans le dernier cas, le rendement en grain était beaucoup plus faible. En supplémentant des cals de riz avec de la 1-pyrroline, il a été démontré que cette imine cyclique était un facteur limitant pour la biosynthèse de la 2AP non seulement dans la variété aromatique, mais également dans la non aromatique. Les essais conduits avec le glucose et les acides gras marqués 13C ont montré que ces molécules pourraient être précurseurs du groupe acétyle de la molécule de 2AP. / The effect of salinity during rice plant growth on the biosynthesis of 2-acetyl-1-pyrroline (2AP), characteristic flavour compound of fragrant rice cultivars was studied. Additionally role of 1-pyrroline in 2AP biosynthesis together with source of acetyl group were investigated. Aychade, a fragrant rice was cultivated either in a liquid medium adapted for seedlings or in pots in a greenhouse. A significant increase in 2AP levels in the leaves was observed whatever the concentration of NaCl (1.7 to 170 mM NaCl) in the culture medium. A positive correlation was found between the level of salt and that of 2AP. There was a relationship between the concentration of proline, a major amino precursor of 2AP and 2AP level at highest concentrations of NaCl. Conversely the changes in -aminobutyric acid (GABA), a competitive metabolite in 2AP biosynthesis were not correlated with those of 2AP. In greenhouse experiments, Aychade cultivar was grown under one level of salt solution, 30 mM that corresponds to EC of 3800 ±400 S.cm-1, value considered sufficient to induce salt stress in rice. Timing and duration of application of salt solution varied according to the growth stages. At vegetative phase a positive relation was observed between salt treatment and 2AP as well as proline levels in the leaves. Interestingly grains from all salt treated plants contained significantly higher levels of 2AP than those from control. The highest synthesis occurred when plants were subjected to salt treatment during whole vegetative and reproductive phases. However in the latter case crop yield decreased significantly. Through supplementation of rice callus with 1-pyrroline it was demonstrated that this cyclic imine was limiting factor for the 2AP biosynthesis not only in a fragrant variety but also in a non fragrant variety. Experiments with 13C labelled glucose and fatty acids showed that glucose and fatty acids could be precursors of acetyl group of 2AP.

The flavour or fragrance of Basmati rice is associated with the presence of 2-acetyl-1- pyrroline. This work shows that a gene with homology to betaine aldehyde dehydrogenase (BAD) has significant polymorphisms in the coding region of fragrant genotypes relative to non fragrant genotypes. Accumulation of 2-acetyl-1-pyrroline in fragrant rice genotypes may be explained by the presence of mutations resulting in loss of function of the fgr gene product. The fgr gene corresponds to the gene encoding BAD2 in rice while BAD1 is encoded by a gene on chromosome 4. Development of an allele specific amplification (ASA) based around the deletion in the gene encoding BAD2 allows, perfect, simple and low cost discrimination between fragrant and non-fragrant rice varieties and identifies homozygous fragrant, homozygous non-fragrant and heterozygous non-fragrant individuals in a population segregating for fragrance. The cDNAs transcribed from rice chromosomes 4 and 8, each encoding an enzyme with homology to betaine aldehyde dehydrogenase were cloned and expressed in E. coli. The enzyme responsible for fragrance, encoded from chromosome 8, had optimum activity at pH 10, showed low affinity towards betaine aldehyde (bet-ald) with Km value of approximately 63ìM but a higher affinity towards -aminobutyraldehyde (GABald) with a Km value of approximately 9ìM. The enzyme encoded from chromosome 4 had optimum activity at pH 9.5 and showed generally lower affinity towards most substrates compared to the enzyme encoded from chromosome 8, substrate specificities suggest that the enzymes have higher specificity to aminoaldehydes and as such both should be renamed as an aminoaldehyde dehydrogenase (AAD). The inactivation of AAD2 (BAD2) in fragrant rice varieties likely leads to accumulation of its main substrate GABald which then cyclises to 1-pyrroline the immediate precursor of 2AP.

The flavour or fragrance of Basmati rice is associated with the presence of 2-acetyl-1- pyrroline. This work shows that a gene with homology to betaine aldehyde dehydrogenase (BAD) has significant polymorphisms in the coding region of fragrant genotypes relative to non fragrant genotypes. Accumulation of 2-acetyl-1-pyrroline in fragrant rice genotypes may be explained by the presence of mutations resulting in loss of function of the fgr gene product. The fgr gene corresponds to the gene encoding BAD2 in rice while BAD1 is encoded by a gene on chromosome 4. Development of an allele specific amplification (ASA) based around the deletion in the gene encoding BAD2 allows, perfect, simple and low cost discrimination between fragrant and non-fragrant rice varieties and identifies homozygous fragrant, homozygous non-fragrant and heterozygous non-fragrant individuals in a population segregating for fragrance. The cDNAs transcribed from rice chromosomes 4 and 8, each encoding an enzyme with homology to betaine aldehyde dehydrogenase were cloned and expressed in E. coli. The enzyme responsible for fragrance, encoded from chromosome 8, had optimum activity at pH 10, showed low affinity towards betaine aldehyde (bet-ald) with Km value of approximately 63ìM but a higher affinity towards -aminobutyraldehyde (GABald) with a Km value of approximately 9ìM. The enzyme encoded from chromosome 4 had optimum activity at pH 9.5 and showed generally lower affinity towards most substrates compared to the enzyme encoded from chromosome 8, substrate specificities suggest that the enzymes have higher specificity to aminoaldehydes and as such both should be renamed as an aminoaldehyde dehydrogenase (AAD). The inactivation of AAD2 (BAD2) in fragrant rice varieties likely leads to accumulation of its main substrate GABald which then cyclises to 1-pyrroline the immediate precursor of 2AP.