The induction of the sucrose synthase (SuSy) gene (SuSy) by low O 2 , low temperature, and limiting carbohydrate supply suggested a role in carbohydrate metabolism under stress conditions. The isolation of a maize (Zea mays L.) line mutant for the two known SuSy genes but functionally normal showed that SuSy activity might not be required for aerobic growth and allowed the possibility of investigating its importance during anaerobic stress. As assessed by root elongation after return to air, hypoxic pretreatment improved anoxic tolerance, in correlation with the number of SuSy genes and the level of SuSy expression. Furthermore, root death in doublemutant seedlings during anoxic incubation could be attributed to the impaired utilization of sucrose (Suc). Collectively, these data provide unequivocal evidence that Suc is the principal C source and that SuSy is the main enzyme active in Suc breakdown in roots of maize seedlings deprived of O 2 . In this situation, SuSy plays a critical role in anoxic tolerance.

Sucrose synthase activity increased in 2-day-old rice (Oryza sativa) seedlings submitted to anaerobic stress. Likewise The unusual behavior of maize sucrose synthase isozymes under anaerobiosis led us to study the anaerobic induction of the enzyme in rice. Rice shares the metabolic strategy of maize roots in response to anaerobic stress. However, in contrast to maize roots, rice is extremely resistant to anoxia. Rice seedlings that have germinated for 48 h aerobically survive more than 21 d and are capable of coleoptile elongation under complete oxygen deprivation. Their metabolism is high enough to permit DNA, RNA, as well as protein synthesis (1,12,13). We have already shown that many of the glycolytic and fermentative enzymes increase in activity (19). ADH (16), cytosolic glyceraldehyde 3-phosphate dehydrogenase (18), and PDC (our unpublished data) are all induced at both the mRNA and protein levels. The rice seedlings referred to above are still attached to the seed that contains starchy reserves. It seemed likely that the existence of these reserves played a role in the active metabolism seen in such seedlings under anoxia. Because starch mobilization is mediated by sucrose synthase, we hypothesized that in rice, sucrose synthase activity and protein levels should be induced by anaerobiosis coordinately with the other enzymes of sugar phosphate metabolism. In maize roots, however, lack ofstarch reserves means that sucrose synthase accumulation would be unnecessary.Our results were obtained using several independent techniques, including enzyme activity measurements, immunodetection on Western blots, SDS, and two-dimensional PAGE of in vivo-and in vitro-labeled total proteins and immunoprecipitates, as well as hybridization analyses of Northern blots and run-on transcripts. Sucrose synthase activity and protein both increased upon anaerobic stress. This was accompanied by an increase in the steady-state level of sucrose synthase mRNA and in the rate of transcription ofthe sucrose synthase gene. We conclude that sucrose synthase is a typical anaerobic polypeptide in rice and is induced at both the protein and mRNA levels. MATERIALS AND METHODS Plant MaterialInbred rice (Oryza sativa L., var Cigalon) seeds were surface-sterilized with a commercial preparation of bleach, then germinated for 48 h in the dark at 25°C under distilled water with vigorous agitation ( 16). Under such conditions, seedlings are under normoxia and have coleoptiles and primary roots about 0.5 cm in length. Anaerobic treatment consisted of www.plantphysiol.org on April 3, 2019 -Published by Downloaded from

Hypoxic pretreatment of tomato (Lycopersiron esculentum M.)roots induced an acclimation to anoxia. Survival in the absence of oxygen was improved from 10 h to more than 36 h if externa1 sucrose was present. The energy charge value of anoxic tissues increased during the course of hypoxic acclimation, indicating an improvement of energy metabolism. In acclimated roots ethanol was produced immediately after transfer to anoxia and little lactic acid accumulated in the tissues. In nonacclimated roots significant ethanol synthesis occurred after a 1-h lag period, during which time large amounts of lactic acid accumulated in the tissues. Severa1 enzyme activities, including that of alcohol dehydrogenase, lactate dehydrogenase, pyruvate decarboxylase, and sucrose synthase, increased during the hypoxic pretreatment. In contrast to maize, hexokinase activities did not increase and phosphorylation of hexoses was strongly inhibited during anoxia in both kinds of tomato roots. Sucrose, but not glucose or fructose, was able to sustain glycolytic flux via the sucrose synthase pathway and allowed anoxic tolerance of acclimated roots. These results are discussed in relation to cytosolic acidosis and the ability of tomato roots to survive anoxia.

Alcohol dehydrogenase (ADH) activity was present in roots and shoots of 48-h rice embryos and rose in response to anoxia. The increase was accompanied by changes in the ADH isozyme pattern. Translatable levels of mRNA for two ADH peptides increases as early as 1 h after the beginning of anoxic treatment. Adh mRNA was detected in aerobically grown rice embryos by hybridization to maize Adh1 cDNA: its level increased significantly after 3 h of anoxia.

The objective of this study was to evaluate the role of enzyme induction and aerenchyma formation in prolonged tolerance to soil flooding in a variety of underground clover (Trifolium subterraneum 'Park') previously selected for resistance. Seedlings were grown in hydroponic tanks, initially with aeration for 3 weeks and subsequently in the absence of aeration for up to 3 weeks. After 1 h in the absence of aeration, the oxygen concentration in the hydroponic medium had decreased to 1.5 %. During the 3 weeks of extreme oxygen deficiency, primary roots died and were replaced by considerable numbers of adventitious roots. Activities of many glycolytic and fermentative enzymes increased in adventitious roots. Excised adventitious roots were capable of immediate induction of ethanol in the absence of lactate production, in association with energy charge higher than that in excised roots of aerobically maintained controls. Energy charge was even higher when measured in adventitious roots in planta. Interestingly, haemoglobin protein could be correlated with energy charge. Aerenchyma was readily visualized in adventitious roots by optical microscopy of longitudinal and transverse sections. We conclude that avoidance of root anoxia via aerenchyma is the major mechanism for prolonged root tolerance in Trifolium subterraneum 'Park'.