Requirement of cotyledons for gibberellic acid-induced hypocotyl elongation in lettuce seedlings. Isolation of the cotyledon factor active in enhancing the effect of gibberellic acid

The role of cotyledons in hypocotyl elongation caused by gibberellicacid was studied using young seedlings of lettuce, Lactuca saliva,var. ‘Grand Rapids’. Removal of cotyledons fromintact seedlings resulted in a depression of hypocotyl elongationcaused by gibberellic acid. Gibberellic acid-induced hypocotylelongation in decotylized seedlings, was however, substantiallyenhanced by incubating the seedlings together with excised cotyledons.The exudate from excised cotyledons also enhanced the effectof gibberellic acid on hypocotyl elongation in decotylized seedlings.This active principle (named the cotyledon factor) in the cotyledonexudate was stable against heating at 100?C for 15 min, permeatedthe dialysis membrane, and was extractable with ethyl acetate.Biological activity of the cotyledon factor was not replacedby indole-3-acetic acid, kinetin, cyclic AMP, vitamins, sucroseor inorganic nutrients. The biological significance of the cotyledonfactor is discussed in relation to the action of gibberellicacid. (Received February 14, 1973; )     

Cloning and expression of sesquiterpene synthase genes from lettuce (Lactuca sativa L.)

Sesquiterpenoid lactones (SLs) from lettuce (Lactuca sativa L.) include constitutive components of latex such as lactucin and the induced phytoalexin, lettucenin A. A redundant primer strategy was used to recover two full length cDNA clones (LTC1 and LTC2) encoding sesquiterpene synthases from a cDNA library derived from seedlings with the red spot disorder, which accumulate phytoalexins. Recombinant enzymes produced from LTC1 and LTC2 in Escherichia coli catalysed the cyclisation of farnesyl diphosphate to germacrene A, potentially an early step in the biosynthesis of SLs. RT-PCR analysis showed LTC1 and LTC2 were expressed constitutively in roots, hypocotyls and true leaves but not in cotyledons. Expression in cotyledons was induced by challenge with the downy mildew pathogen Bremia lactucae in the disease resistant cultivar Diana. Southern hybridisation experiments showed that LTC1 and LTC2 were not part of a multigene family. The germacrene A synthases provide targets for modified expression to generate beneficial modifications to the SL profile in lettuce.     

Frugivory by insects on mangrove propagules: effects on the early life history of Avicennia marina

This study investigates how herbivory by larval insects on fruit of the mangrove Avicennia marina affects the subsequent establishment, growth, and survivorship of its seedlings. Research was done in a temperate mangrove forest in Australia where the larvae of two species of insects were the dominant frugivores: the mangrove fruit fly Euphranta marina and the mangrove plume moth Cenoloba obliteralis. Larvae consumed the cotyledons of fruit, but not their embryonic axes. Damage to the cotyledons of fruit while on the tree was obvious in abscised propagules and the cotyledons of seedlings, indicating that larvae within fruit continue to consume the cotyledons and develop while the propagule disperses and the seedling establishes. We found that 53% of fruit, 69% of abscised propagules, and 80% of the cotyledons of seedlings had been attacked by larval insects. The degree of damage to the cotyledons was positively related to the densities of larvae within the fruit and the number of emergence or exit holes in the surface of the cotyledons. Consequently, the number of exit holes could be used as a proxy for the amount of damage to the cotyledons. An experiment in which we placed propagules with different levels of damage (zero, one, two, or three exit holes) on the forest floor revealed that frugivory did not influence the establishment of seedlings. In another experiment, we monitored the performance of newly established seedlings with cotyledons that had zero exit holes, one or more exit holes, or where we had removed their cotyledons. In both experiments, the growth of seedlings as measured by their heights and number of leaves was negatively related to the degree of frugivory. Consumption of the cotyledons by larvae apparently reduces the energy reserves available for the initial growth of seedlings. The early mortality of seedlings in both experiments was minimal and appeared to be independent of differences in size due to frugivory. Experimentally removing the cotyledons of seedlings produced extremely short seedlings with increased mortality, confirming the importance of cotyledonary reserves to the initial growth and survivorship of seedlings of A. marina. This study highlights how the early life history of plants may be impacted by conditions experienced by the propagules before they disperse from the parental plant.     

Involvement of gibberellins in expression of a cysteine proteinase (SH-EP) in cotyledons of Vigna mungo seedlings.

The expression of a papain-type proteinase, designated SH-EP, in cotyledons of Vigna mungo seedlings has been shown to require some factors in the embryonic axes. Gibberellin A1 (GA(1)) and GA(20) were identified by GC-MS in embryonic axes of V. mungo seedlings. The level of accumulation of SH-EP in cotyledons of V. mungo seedlings was greatly reduced by treatment of the seeds with uniconazole-P, an inhibitor for GA biosynthesis. The reduced level of accumulation of SH-EP in cotyledons by uniconazole-P was recovered by exogenous application of GA(1) and GA(20) to the seedlings.     

Light and molybdenum requirements for substrate induction of nitrate reductase in cotyledons of lettuce,Lactuca sativa L.

Light was required for induction of nitrate reductase (NR, E.C. 1.6.6.1) in intact cotyledons of 2-day old seedlings ofLactuca sativa L. Molybdate strongly enhanced efficiency of induction. Benzyladenine (BA), gibberellin, and succinic acid-2,2-dimethylhydrazide reduced the enzyme activity. BA thrice enhanced incorporation of labelled leucine to the protein fraction. (2-chloroethyl)trimethylammonium chloride did not affect NR activity and markedly inhibited greening and protein synthesis. KNO₃ stimulated protein synthesis as well as growth of the cotyledons.     

Phytochrome-mediated control of respiratory activities of mitochondria in cotyledons of dark-grown cucumber seedlings

Mitochondria isolated from cotyledons of dark-grown cucumber ( Cucumber sativus L., cv. Shimotsuki-Aonaga) seedlings after illumination with continuous far-red light showed an increased capacity for oxidation of malate or α-ketoglutarate, as compared with those from cotyledons of non-illuminated seedlings. This increase is supposed to be caused by phytochrome action (high irradiance response). Exogenous NAD⁺ had no effect on the rate of the oxidation of α-ketoglutarate or malate by mitochondria isolated from far-red light-treated cotyledons, but it enhanced the oxidation rate of mitochondria from control cotyledons to the level of mitochondria from light-treated ones. The NAD (NAD⁺+ NADH) content was higher in mitochondria isolated from continuously far-red light-treated cotyledons than in mitochondria from controls. The NAD content was also increased by the treatment with a red light pulse and this response was reversed by a subsequent far-red light pulse. It is proposed that phytochrome controls respiratory activities of cucumber mitochondria by changing the size of the NAD pool in the mitochondria.     

Stimulation of cytokinins and chlorophyll synthesis in cucumber cotyledons by triadimefon

Triadimefon [1-(4-chlorophenoxy)-3,3-dimethyl-1-(1,2,4-triazol-1-yl)-2-butanone] changes the morphology and partitioning of dry matter in cucumber ( Cucumis sativus L. cv. National Pickling) seedlings. The dry weights, potassium and cytokinin levels in the cotyledons and roots of the treated seedlings were higher, whereas the hypocotyl weights were lower than the controls. When etiolated intact seedlings or cotyledons excised from triadimefon-pretreated dark-grown seedlings were exposed to light, chlorophyll synthesis in the pretreated cotyledons was stimulated. Triadimefon does not have cytokinin-like activity in the cucumber cotyledon greening bioassay, but appears to induce the plants to produce more cytokinims, probably by stimulating root growth. Hence it is proposed that the stimulation of chlorophyll production by triadimefon in cucumber cotyledons is mediated by maintaining high levels of potassium and cytokinins in the cotyledons.     

Nicotinate riboside salvage in plants: presence of nicotinate riboside kinase in mungbean seedlings.

Salvage of nicotinate riboside for NAD synthesis was investigated in mungbean seedlings. Nicotinate riboside kinase activity was detected in extracts from cotyledons. Exogenously supplied [carboxyl-(14)C]nicotinate riboside was readily converted into pyridine nucleotides in cotyledons of mungbean seedlings. This conversion was also found in embryonic axes, but the rate was lower than in cotyledons. These results suggest that, in addition to the seven-component pyridine nucleotide cycle (PNC VII), an eight-component cycle (PNC VIII) involving nicotinate riboside kinase operates in plants.     

Identification in lettuce seedlings of a catecholamine active in synergistically enhancing the gibberellin effect on lettuce hypocotyl elongation

The occurrence of catecholamines in lettuce seedlings was examined by bioassay and gas chromatography-mass spectrometry (GC-MS), since synthetic catecholamines can synergistically enhance the stimulating effect of gibberellic acid (GA₃) on hypocotyl elongation of decotylized lettuce seedlings. The catecholamine fraction on TLC obtained from lettuce seedlings synergistically enhanced the GA₃ effect on hypocotyl elongation. The analysis of the catecholamine fraction from lettuce seedlings by GC-MS demonstrated the occurrence of dopamine.     

Development of the primary root and mobilisation of reserves in etiolated seedlings of Brassica napus grown on a slowly rotating clinostat

The effect of the slow rotating clinostat (1 rpm) on the growth of the primary root was studied on Brassica napus seedlings. After 5 d in darkness, the primary root was longer and thinner in seedlings grown on the clinostat than in seedlings grown in the vertical position. However, the breakdown of lipid reserves, sucrose level and transport of 14C-labeled sucrose from the cotyledons to the primary root, were not altered by growth on the clinostat. Moreover, the activity of isocitrate lyase, one of the two enzymes necessary for the conversion of lipids into glucids also was also not modified in the cotyledons of clinorotated seedlings. Thus, there was clear evidence that clinorotation had a direct effect on the growth of the primary root that was independent of the mobilisation of lipid reserves in the cotyledons. As a sink, the primary root had the same strength on the clinostat as in the vertical position, but the reserves were used in a different way. The increase in root elongation on the clinostat could be due to the slight, but continuous, omnilateral gravitropic stimulation due to the rotation of the seedlings about a horizontal axis.     

Induction of lincomycin and streptomycin resistance by nitrosomethylurea and ethyl methanesulphonate in Capsicum annuum L.

A high frequency of plastid-encoded antibiotic-resistant variants of Capsicum annuum were isolated on selective media following treatment by ethyl methanesulphonate (EMS) and nitrosomethylurea (NMU). Seeds and explants were mutagenized with 0.1% EMS and 5 mm NMU separately. Non-mutagenized cotyledons (controls), mutagenized cotyledons from EMS-treated seedlings and NMU-treated cotyledons were placed on regeneration medium supplemented with the antibiotics streptomycin or lincomycin. Resistant shoots appeared at a high frequency in mutagenized cotyledons, whereas in controls morphogenesis was suppressed, accompanied by bleaching. The stability of streptomycin and lincomycin resistance was confirmed by leaf assay. NMU-treated cotyledons gave a higher frequency of variants than cotyledons from EMS-treated seedlings. The mutagenic effect of EMS was more pronounced using whole seeds rather than cotyledons; in contrast, NMU was more effective in inducing variations in cotyledons than in seeds. Received: 1 October 1996 / Revision received: 12 May 1997 / Accepted: 3 June 1997     

Influence of UV-B Radiation on Early Seedling Growth and Translocation of 65Zn from Cotyledons in Cotton

The influence of UV-B radiation from filtered or unfiltered fluorescent sunlamps on early seedling growth and translocation of ⁶⁵Zn from cotyledons to the shoot was examined in two cultivars of cotton, Acala and Gregg. Ten-day-old seedlings which had been irradiated in the greenhouse for 6 h continuously each day for 14 days with 0.81 or 1.61 W × m^-2 UV-B radiation under two unfiltered FS-40 sunlamps, showed pronounced phytotoxic damage. This was characterized at first by bronzing and glazing of the cotyledons and later by upward curling of the leaves and abscission. Leaf expansion, dry matter accumulation, and mobilization of ⁶⁵Zn from the cotyledons was severely impaired in the young developing shoot under unfiltered UV-B radiation. A significant stress response also was observed in seedlings exposed to 0.61 W × m^-2 UV-B radiation through a polystyrene filter and 0.73 W × m^-2 UV-B radiation through a cellulose-acetate filter. This stress response was characterized by the formation of a red pigment in the petioles of the cotyledons, reduced leaf expansion, and reduced transport of ⁶⁵Zn. Control seedlings exposed to 0.03 W × m^-2 UV-B radiation through a mylar filter were green, had maximum leaf size and dry-matter accumulation, and had the greatest percentage of ⁶⁵Zn translocated from the cotyledons.     

Effect of horizontal clinorotation on the root system development and on lipid breakdown in rapeseed (Brassica napus) seedlings

Seedlings of Brassica napus were cultivated on a slowly rotating clinostat (1 rpm) or in the vertical control for 5 d. The root growth, the cotyledonary reserves and the transport of 14C-labeled sucrose from cotyledons to root system were studied in both cultural conditions. The biomass (fresh weight) of the root system was 35% higher in the horizontally clinorotated seedlings than in the controls. This increase was correlated with a greater degradation of reserve lipids and faster accumulation of sucrose in the cotyledons. The activity of isocitrate lyase, one of the two enzymes necessary to conversion of lipids into glucids, was also greater in the cotyledons of clinorotated seedlings. The labeling distribution of 14C in the cotyledons, the hypocotyl and the root system after 30, 60 and 120 min of application of 14C-labeled sucrose on the cotyledons showed higher translocation of the cotyledonary sucrose to the root system of clinorotated seedlings. In addition, we studied the effects of clinorotation on the biomass of the excised root system (of 10 d old seedlings) cultivated in a medium containing 1% sucrose. The horizontally clinorotated root system grew more than that of the controls. These results showed that the horizontal clinorotation acted on the root system growth and provoked a higher sucrose translocation from source to sink, i.e. from cotyledons to root system.     

Water stress in cucumber cotyledons infected withXanthomonas campestris pv.cucurbitae

Xanthomonas compestris pv.cucurbitae strain 62 was isolated from diseased cucumber seedlings grown in an open field in Egypt and identified in this study. Inoculation of cucumber seedlings with this strain resulted in reduction of relative water content (RWC) and increase in free proline content of infected cotyledons. RWC was found to be decreased by the increase of disease severity. The reverse was true for free proline content which was found to be increased by the increase of disease severity. These results indicate that water stress increased in direct proportion to the disease severity. Highly significant increase in free proline content was found to occur in organs of cucumber seedlings inoculated with strain 62 and showing disease symptoms on cotyledons only. This implied that water stress was not limited to symptomatic cotyledons but extended to the whole seedling.     

A development-dependent hemagglutinin from cucumber surfaces: I. Hemagglutinin activity of cotyledon surfaces

During studies on the chemistry of plant surfaces, we observed that concomitant with the development of cucumber (Cucumis sativus L. cv `ELEM') cotyledons an agglutinin that agglutinates human erythrocytes appeared on epicuticular surfaces. The agglutinin was released from cotyledon surfaces into distilled water by a 1-minute immersion (or even less). Homogenization of the washed cotyledons released residual agglutinating activity. The surface-located hemagglutinating activity was age-dependent and occurred in dark- and light-grown cotyledons. Agglutinating activities were present in light-grown cotyledons of 2- to 14-day-old seedlings and in dark-grown cotyledons of 3- to 17-day-old seedlings. Agglutinating activities were maximal in cotyledons of 3- to 4-day-old seedlings. No activity could be detected in dry seeds or in seedlings up to 2 days and after 17 days of germination. The hemagglutinating activity was specifically inhibited by N,N′,N″-triacetylchitotriose, chitin, and chitosan.     

Anthocyanin Synthesis in Seedlings of Vigna radiata (L.) Wilczek: Interaction between Hypocotyl and Cotyledons

During the development of mung bean seedlings, cotyledons graduallyacquired the capacity to synthesize anthocyanins after red lightirradiation. When isolated from 36 hr old seedlings, cotyledonsaccumulated anthocyanins with a time-course similar to the oneobserved with cotyledons of intact seedlings (Dumortier andVendrig 1982). In both cases, there were two periods of rapidpigment accumulation separated by a stagnation phase. Experimentsin which the hypocotyl hook or the cotyledons had been shieldedduring irradiation led to the conclusion that cotyledons containphotoreceptors for anthocyanin accumulation in the hypocotylhook (which also had to be irradiated). The reverse was alsotrue: irradiation of the hypocotyl hook influenced pigment accumulationin the cotyledons in a positive way. A hypothesis is presentedconcerning the level at which P_FR exerts its influence. (Received January 18, 1982; Accepted April 30, 1982)     

Role of ethylene in chlorophyll degradation in radish cotyledons

The effect of age of radish seedlings on changes in chlorophyll concentration caused by ethylene was examined. Ethylene was produced at 2–4 nl g^?1 h^?1 following excision of cotyledons from 5-to 20-day-old seedlings. The youngest cotyledons maintained this rate, whereas ethylene synthesis declined by as much as 80% during a 24-h period in older cotyledons. The youngest cotyledons continued to accumulate chlorophyll in the dark, but after 7 days cotyledons lost chlorophyll and the proportion of chlorophyll lost increased with age. Ethylene promoted, and norbornadiene inhibited, this loss of chlorophyll; in combined treatments the effects of ethylene and norbornadiene were competitive. The maximal rate of chlorophyll loss occurred in 1μl L^?1 ethylene; extrapolation of the response to concentration indicated that half-maximum loss would occur at 0.005–0.01 μl L^?1 ethylene. In cotyledons from 20-day-old seedlings, chlorophyll degradation occurred mainly after 24 h from excision and transfer to the dark. Chlorophyll degradation during 48 h in the dark was affected by norbornadiene or ethylene applied from 0–24 h or from 24–48 h.     

一种鉴定金花茶人工杂交F_1代杂种苗的简易方法

本试验选用了两个以金花茶作父本的人工杂交组合F_1代实生苗,经根尖体细胞染色体观察鉴定四倍体杂种。一、云南野山茶(Camellia pitardii var.yunnanica六倍体)×金花茶(C.Chrysantha二倍体)。结果为:所获56株子叶和下胚轴为红色的F_1代杂种苗中,有55株(98.2％)是真正的四倍体杂种(X~2=0.0128,D.f.=1,0.95>p>0.90);所获50株子叶和下胚轴为黄白色的杂种苗中,有13株(26％)为真正四倍体杂种(X~2=27.38,D.f.=13,0.02>p>0.01)。二、云南山茶花(C.reticulata六倍体)×金花茶。结果为:在78株子叶和下胚轴为红色的杂交苗中,有77株(98.7％)为真正四倍体杂种(X~2=0.0128,D.f.=1,0.95>p>0.90);而在78株子叶及下胚轴为黄白色的杂种苗中,只有21株(26.9％)是真正四倍体杂种(X~2=41.65,D.f.=24,0.02>p>0.01) 在多数杂种实生苗中的这种红色素,是因种子直感现象而发生的父本金花茶的一种遗传性状。金花茶的另一特征——多子叶现象(3枚以上),则在一些F_1代杂种苗或杂种种子中表现得不明显。因此,利用F_1代杂种苗子叶和下胚轴所具有的红色特征,在杂种种子萌发期用来鉴别以金花茶为亲本的F_1代杂种的真伪,是一种简便、快速的、有发展前途的方法。     

The role of roots and cotyledons as storage organs in early stages of establishment in Quercus crispula: a quantitative analysis of the nonstructural carbohydrate in cotyledons and roots

Quercus seedlings have hypogeal cotyledons and tap roots, both of which act as storage organs. The importance of the storage function in the two organs may change as the seedling develops. Therefore, changes in carbohydrate reserves in cotyledons and roots of Q. crispula grown under 40 % and 3 % of full light from shoot emergence to the completion of the first leaf flush were monitored. In addition, a shoot-clipping treatment was performed to examine the relative contribution of the cotyledons and tap roots to resprouting. Cotyledons maintained large amounts of nonstructural carbohydrates during shoot development, and carbohydrates were still present in the cotyledons during the final phase of leaf flush. In addition, a notable increase in the amount of carbohydrates was observed in tap roots before leaf flush at both light levels. Since root development occurred before leaf flush, even in plants grown under 3 % light, the carbohydrate found in them presumably originated from seed reserves and was translocated to roots as storage reserves. When shoots were clipped at the leaf flushing stage, the amount of carbohydrate decreased only in the cotyledons after resprouting, suggesting that cotyledons act as the main storage organs during shoot development stages. However, it could be advantageous as a 'risk avoidance strategy' for the seedlings to store reserves in both cotyledons and roots, since cotyledons may be removed by predators during shoot development.