The level of flavonoids and amines in de-etiolated and methyl jasmonate treated seedling of common buckwheat

The effects of atmospheric methyl jasmonate on the level of flavonoids and biogenic amines in de-etiolated seedlings of common buckwheat (Fagopyrum esculentum Moench) were investigated. In cotyledons and hypocotyls of etiolated seedlings, some traces of anthocyanins were found, with no flavones and flavonols identified. A measurable content of flavones and flavonols was, however, determined in roots. De-etiolation process stimulated the accumulation of all flavonoid types. Methyl jasmonate clearly decreased the content of anthocyanins in the hypocotyl, not affecting their level in cotyledons. In case of roots, the content of anthocyanins increased after a 4-day treatment. In general, reduction in the level of flavones and flavonols was recorded only in the hypocotyl, however it was not always significant. Cotyledons of the seedlings treated with methyl jasmonate responded by a slight increase in flavonoids level. Methyl jasmonate considerably induced the accumulation of 2-phenylethylamine in all the seedling organs, increasing the content of putrescine and tryptamine in cotyledons, and decreasing the level of tryptamine in roots.     

Effect of the hypocotyl hook on greening in etiolated cucumber cotyledons

Four-day-old etiolated cucumber cotyledons (Cucumis sativus, L.) were excised and allowed to green in white fluorescent light at 28 C. Cotyledons excised with a full hypocotyl hook exhibited a lag phase of 1 hour before entering the rapid greening phase, whereas cotyledons excised without any hypocotyl hook exhibited a lag phase of 6 hours. Cotyledons excised with varying lengths of hypocotyl hook accumulated chlorophyll roughly in proportion to the hook length. When cotyledons were excised with a full hook and were partially or totally shielded from light with aluminum foil, the samples with the hook covered accumulated more chlorophyll than the wholly exposed samples. The samples with the cotyledons covered showed no net accumulation of chlorophyll irrespective of hook's exposure to light. These data suggest the contribution of some factor or factors by the hypocotyl hook which reduce the lag phase during greening.     

The Role of Phytochrome in Anthocyanin Synthesis in Seedlings of Vigna radiata (L.) Wilczek

Previously, it has been demonstrated that the red light-inducedanthocyanin accumulation in mung bean seedlings is mediatedby phytochrome [Dumortier and Vendrig Z. Pflanzenphysiol. 87:313 (1978)]. In this paper the importance of phytochrome forthe accumulation of anthocyanins in seedlings of mung beanswas studied in non-irradiated seedlings and in seedlings irradiatedwith 5 min R. A short FR-irradiation given early after sowing reduced theamount of anthocyanins which were normally found in non-irradiatedseedlings. This indicates that P_FR may be important for at leastpart of the anthocyanin synthesis in the dark. As for the redlight-mediated anthocyanin accumulation, irradiation appearedto be most effective when given to seedlings at the age of 36–48hr. Although the seedlings were sensitive to red light irradiationbefore that time, they were not able to synthesize anthocyaninsuntil they had reached the age of 36 hr. Complete escape ofred/far-red reversibility occurred only when far-red was given12 hr after red, although partial escape could be observed witha shorter time-interval. Furthermore, the time-course of anthocyaninaccumulation after a two-fold R-irradiation was compared withthe effect of a single R-exposure. From the results could beconcluded that the pattern of anthocyanin accumulation is dependenton the time during which P_FR is present in the seedlings. Theseexperiments also indicate that P_FR not only plays a role inthe synthesis of anthocyanins but probably also in their degradation. The results of our study show that phytochrome is importantfor anthocyanin accumulation in non-irradiated mung bean seedlingsas well as in R-irradiated, and that it probably is also involvedin the degradation of the pigment. (Received January 18, 1982; Accepted April 30, 1982)     

Control by Light of Hypocotyl Elongation and Levels of Endogenous Gibberellins in Seedlings of Lactuca sativa L.

Four 13-hydroxygibberellins, gibberellin A₁ (GA₁), 3-epi-GA₁,GA₁₉ and GA₂₀ were identified by full-scan GC/MS in extractsof lettuce seedlings (Lactuca sativa L. cv. Grand Rapids). Theresults suggest that the early-13-hydroxylation biosyntheticpathway to GA₁ functions in the lettuce seedlings. It was alsofound that GA₁ is active per se in the control of hypocotylelongation in lettuce seedlings. To investigate the relationshipbetween control by light of hypocotyl elongation and levelsof endogenous GAs in lettuce, endogenous levels of GAs werequantified by radioimmunoassay in seedlings that had been grownfor 5 days in the dark (5D) and in those that had been grownfor 4 days in the dark and then under white light for 1 day(4D1L). The endogenous level of GA₁ in the upper and lower partsof hypocotyls in 5D seedlings was about four times higher thanthat in 4D1L seedlings. The response of explants (hypocotylsegments with cotyledons) from dark-grown seedlings to GA₁ isknown to be similar in the dark and under white light when theexplants are treated with inhibitors of the biosynthesis ofGA. Therefore, the above information suggests that the highlevel of GA₁ in hypocotyls of dark-grown seedlings is responsiblefor the rapid elongation of hypocotyl, while irradiation bywhite light decreases the endogenous level of GA₁ in the hypocotylswith a resultant decrease in the rate of hypocotyl elongation. (Received March 13, 1992; Accepted May 21, 1992)     

Organ- and Tissue-specific Biosynthesis of Flavonoids in Seedlings of Oenothera odorata (Onagraceae)

In developing Oenothera odorata seedlings, phytochrome-mediated accumulation of various flavonoids (mainly glycosides of cyanidin and quercetin) is detectable, subsequent to a transient induction of the key enzymes of the general phenylpropanoid metabolism, L-phenylalanine ammonia lyase (PAL) and of flavonoid biosynthesis, chalcone synthase (CHS). Organ- and tissue-specific distribution of these enzymes and of the flavonoid end products was investigated in seedlings, irradiated with continuous far-red light. Anthocyanins and quercetin glycosides are mainly localized in both the upper and lower epidermis of the cotyledons and to a lesser extent also in the epidermal cell layer of the hypocotyl. An obvious organ-specific distribution was observed for the anthocyanins: cyanidin-3,5-O-diglucoside accumulates in the epidermal cells of the cotyledons, whereas cyanidin-3-O-glucoside is restricted to the epidermis of the hypocotyl. By contrast the pattern of quercetin glycosides is the same in the cotyledons and in the hypocotyl. The methylated flavonol aglycone 3-0-methylquercetin was found to be localized in the seed coat. According to this organ- and tissue-specific pattern of flavonoids, immunochemical and immunohistochemical detection of PAL and CHS revealed a predominant localization of theenzymes in the epidermal layers of the cotyledons and the hypocotyl but also in the cells surrounding the vascular bundles. The role of compartmentation in regulation of flavonoid biosynthesis and putative functions of flavonoid compounds are discussed.     

A double enhancement of greening in the cotyledons of cucumber seedlings by excision and red light

Cotyledons excised without the hypocotyl hook from 6-day-old etiolated cucumber ( Cucumis sativus L. var. Elem) seedlings accumulated a significantly higher amount of chlorophyll than cotyledons excised with hooks or intact cotyledons. It was found that maximum ehancement of greening was achieved after 2 h of dark incubation following excision. Pretreatments with red light effected an additive rise in chlorophyll level in subsequent white light after a dark incubation, suggesting that the effects of excision and phytochrome on greening act independently. Etiolated seedlings were variously dissected before greening and it was found that enhancement occurred only when cotyledons were excised at the level of the hypocotyl hook or above it. Similar results were obtained when the dissected plants were pre-treated with red light.     

Opening of the hypocotyl hook in seedlings as influenced by light and adjacent tissues

The influence of the cotyledons and apical bud and the root system on the light-induced opening of the hypocotyl hook of etiolated seedlings of Gossypium hirsutum L., Phaseolus vulgaris L., Helianthus annuus L., Ipomoea alla L., Ipomoea sp., Cucumis sativus L., Linum usitatissimum L., Hibiscus esculentus L., and Raphanus sativus L. was studied. Light stimulated the opening of hypocotyl hook in all cases, but the cotyledons and roots had different effects in different plants. Generally, the presence of cotyledons and the remainder of the shoot apical to the hook inhibited light-mediated opening, but in Gossypium the organs stimulated light-mediated opening. Presence of roots either promoted opening, had no effect, or had an effect only when the cotyledons were present. In the dark the adjacent organs had a reduced effect over that shown in the light, but one cultivar of cotton, Acala SJ1, opened the hook in the dark without cotyledons as much as under any condition in the light. The variation between species in hook opening may related to the need of that process for a proper hormonal balance, as affected by light, which must be obtained from adjacent tissues.     

Ethylene-mediated enhancement of apical hook formation in etiolated Arabidopsis thaliana seedlings is gibberellin dependent

Dark-grown Arabidopsis seedlings develop an apical hook by differential elongation and division of hypocotyl cells. This allows the curved hypocotyl to gently drag the apex, which is protected by the cotyledons, upwards through the soil. Several plant hormones are known to be involved in hook development, including ethylene, which causes exaggeration of the hook. We show that gibberellins (GAs) are also involved in this process. Inhibition of GA biosynthesis with paclobutrazol (PAC) prevented hook formation in wild-type (WT) seedlings and in constitutive ethylene response (ctr)1-1, a mutant that exhibits a constitutive ethylene response. In addition, a GA-deficient mutant (ga1-3) did not form an apical hook in the presence of the ethylene precursor 1-aminocyclopropane-1-carboxylate (ACC). Analysis of transgenic Arabidopsis seedlings expressing a green fluorescent protein (GFP)-repressor of ga1-3 (RGA) fusion protein suggested that ACC inhibits cell elongation in the apical hook by inhibition of GA signaling. A decreased feedback of GA possibly causes an induction of GA biosynthesis based upon the expression of genes encoding copalyl diphosphate synthase (CPS; GA1) and GA 2-oxidase (AtGA2ox1). Furthermore, expression of GASA1, a GA-response gene, suggests that differential cell elongation in the apical hook might be a result of differential GA-sensitivity.     

The role of the apex in normal and tropic growth of sunflower hypocotyls

Regional growth in vertical and horizontal etiolated sunflower hypocotyls from which the apical hook tissue had been either partly or wholly excised, was measured 24 h later, the regions having been demarcated with resin beads. Removal of the cotyledons (an excision which included the distal end of the shoot apex) had little effect on growth during this period but excision of the apical hook significantly reduced growth. In vertically orientated seedlings, removal of half of the hook severely reduced growth in all other growing regions and removal of the entire hook totally inhibited growth. This inhibition of growth was not a consequence of the removal of the region of growth but a consequence of the removal of a region on which growth was dependent. In horizontal seedlings, the situation was more complex inasmuch as a horizontal orientation itself induced growth in previously non-growing regions. This new growth was localised in its extent and was not as severely affected by progressive excision of the hook as was growth in vertical seedlings. The results are discussed in terms of overall growth co-ordination in the hypocotyl.     

Characteristics of hook formation by bean seedlings

Explants were isolated from 6-day-old etiolated bean seedlings (Phaseolus vulgaris L. cv. Black Valentine) containing the cotyledons with 4 mm of hypocotyl just below the node and/or the epicotyl. During incubation on distilled water, uneven growth of the hypocotyl or epicotyl occurred resulting in the formation of a hook. The more rapid growth of the side which became convex was not dependent upon the presence of the slower growing concave side. It was concluded that the main axis has an intrinsic capacity for asymmetric growth. The growth leading to hook formation was inhibited by α-naphthaleneacetic acid at concentrations above 0.2 milligram per liter.     

The positive hook: the role of gravity in the formation and opening of the apical hook

Photographic observations on germinating seedlings of Lepidium sativum L., Cucumis sativus L., and Helianthus annuus L. showed that the hypocotyl hook is not present in the seed but forms during the early stages of growth. Evidence that gravity plays a major role in inducing curvature of the hypocotyl, and in maintaining the hook once it has been formed, was obtained from clinostat experiments, from the use of morphactin to remove geotropic sensitivity and from inversion of seedlings to change the direction of the geostimulus. In L. sativum and H. annuus gravity perception seemed to be the only mechanism responsible for hook formation. In C. sativus hook formation was additionally aided by the mode of emergence of the cotyledons from the seed coat but gravity played an indirect role in regulating such emergence. Further evidence that hook formation is linked to a georesponse was derived from a comparison of hypocotyl development in wild-type Arabidopsis thaliana seedlings with that of an ageotropic mutant, hook formation being found to occur only in the wild type. Hook formation and maintenance is discussed in terms of contrasting geosensitivity between the apical and basal ends of the hypocotyl and it is suggested that light-induced hook opening is a reversal to a condition of uniformly negative georesponse throughout the hypocotyl.     

Osmotic potential of the epidermal cells of cucumber hypocotyls as affected by gibberellin and cotyledons

By using light-grown cucumber (cv. Aonagajibae) seedlings withfully expanded cotyledons, the osmotic potential of the epidermalcells of the hypocotyl, as determined by the plasmolysis methodwas examined in relation to the effects of gibberellin (GA₇)and cotyledons. GA₇ at a dosage level of 0.05 µg/plantapplied to the apical bud lowered the osmotic potential; thedecrease was significant 5 hr after the GA₇ treatment. Removalof the cotyledons, covering them with aluminum foil, or applyinglinuron, a DCMU analogue, to them caused an increase in thepotential. In light-grown cucumber seedlings, the maintenanceof a lower osmotic potential in the epidermal cells of the hypocotylwas concluded to be regulated by the photosynthetic activityof the cotyledon and also by gibberellin. ¹Noriko Katsu, Kitano-Haitsu 203, Saginuma 1-9-10, Takatsu-ku,Kawasaki 218, Japan. (Received May 30, 1980; )     

Effect of Sink-Source Manipulations on the Photosynthetic Rate and Carbohydrate Content of Cucumber Cotyledons

Mayoral, M. L., Plaut, Z. and Reinhold, L. 1985. Effect of sink-sourcemanipulations on the photosynthetic rate and carbohydrate contentof cucumber cotyledons.-J. exp. Bot. 36 1551–1558. The photosynthetic rate of cucumber cotyledons (Cucumis sativuscv. Dahla) reached a maximum value of 12 mg dm^–2 h^–1,10 d after emergence. In 12-d-old seedlings removal of one cotyledondoubled the CO₂ fixation rate of the other, as observed 3 dafter treatment. When the primary leaf was removed, the photosyntheticrate of the cotyledons was decreased by 33%. At this stage ofgrowth elimination of the roots as a sink for assimilates bygirdling the hypocotyl affected neither the photosynthetic ratenor the carbohydrate content of the cotyledons. By contrast,in 18-d-old seedlings removal of the first leaf brought abouta 42% increase in the photosynthetic rate of the cotyledons.The simultaneous removal of the first leaf and one cotyledondoubled the rate of CO₂ fixation of the remaining cotyledon.Girdling the hypocotyl lowered the photosynthetic rate of thecotyledons by 73%. In both 12- and 18-d-old seedlings a decreaseor increase in the sink-source ratio was correlated with anincrease or a decrease respectively in the carbohydrate contentof the cotyledons. The stomatal resistance of the cotyledonswas not affected by any of the treatments. The effect of sink-sourcemanipulations on photosynthesis and on the level of carbohydratespresent in the cotyledons was more evident in those seedlingsgrowing under high light intensity (580 µE m^–2 s^–1),than in those exposed to 300 µE m^–2 s^–1 Key words: Sink-source relationship, cotyledons, photosynthesis     

Effect of darkening the cotyledons on the growth and curvature of the sunflower hypocotyl: evidence of hydraulic signalling

When both cotyledons of light-grown sunflower seedlings (Helianthusannuus L.) were darkened by covering them with aluminium foil,the resulting increase in the rate of elongation of the hypocotylwas closely proportional to the associated reduction in seedlingtranspiration and to the increase in xylem water potential (_PX.Covering only one cotyledon induced curvature away from thatside. This response was associated with a higher _PX in the vascularbundles connected directly with the covered cotyledon than inthose connected with the illuminated cotyledon. The water contentof peripheral tissues of the hypocotyl below the covered cotyledonwas also higher than that of similar samples from below theilluminated cotyledon. These results are consistent with thehypothesis that the effect of darkening the cotyledons on thegrowth and curvature of the hypocotyl is mediated by hydraulicsignalling, characterized by transmission to the hypocotyl ofan increase in _PX resulting from the reduction in cotyledontranspiration. Key words: Cotyledons, Helianthus annuus, hydraulic signalling, hypocotyl, transpiration, water potential     

Promotion of Hypocotyl Elongation in Watermelc Seedlings by 6-Benzyladenine

Hypocotyl elongation under white fluorescent light was aboutdoubled in dwarf watermelon (Citrullus lanatus0 (Thunb.) Matsu.and Nakai) seedlings treated with 0.1 to 0.3 µg apicaland 3 x 10–6 to 10.3 M root applications of 6-benzyladenine(BA). BA-enhancement of growth occurred primarily during thefirst 48 h after treatment. Increased hypocotyl length in BA-treatedseedlings was attributed more to longer cells than to an increasein cell number. Early hypocotyl growth of normal seedlings wasalso significantly enhanced by BA although final hypocotyl lengthwas not substantially affected. Benzyladenine caused expansion of cotyledons and, at higherdoses, lateral expansion of hypocotyls. BA-induced increasesin fresh weight of cotyledons and hypocotyls were accompaniedby an increase in dry weight of hypocotyls at the expense ofroots which had less dry matter than untreated seedlings.     

Effect of the Hypocotyl Hook on Chlorophyll Accumulation in Excised Cotyledons of Cucumis sativus L

Hypocotyl hooks have been shown to influence greening in excised cucumber (Cucumis sativus) cotyledons. The properties of the lag phase are greatly affected by the presence or absence of the hook tissue. A 45-second light pretreatment followed by 4 hours of darkness is sufficient to remove the lag phase from cotyledons with hooks, while hookless cotyledons require 2 hours of continuous illumination followed by 1 hour of dark incubation to break the lag phase. The effect of hooks on cotyledon greening is enhanced if the hooks are shielded from light. Cutting off the hooks after lag phase removal caused a marked decrease in chlorophyll accumulation in the cotyledons. These observations may indicate that the hypocotyl hooks produce a substance or substances needed in the greening process, which are translocated to the cotyledons. Indoleacetic acid, abscisic acid, gibberellin A₃, 6-benzylamino purine and δ-aminolevulinic acid do not show any activity; on the other hand, ethylene appears to replace partially the hypocotyl hooks.     

Factors involved in the opening of the hypocotyl hook of cotton and beans

Conditions influencing the opening of the bean (Phaseolus vulgaris L.) and cotton (Gossypium hirsutum L.) hypocotyl hook were defined. Such hooks were shown to undergo geotropic curvature; orientation of the hook with respect to gravity greatly affected the observed opening. Cotton and bean hooks behaved exactly opposite in regard to the presence of the cotyledons and apical bud. The cotton hook required the cotyledons for opening, but the corresponding tissue slowed or inhibited opening of the bean hook. With cotton, lower hypocotyl and root tissues stimulated hook opening, but with bean, the tissues below the hook section had little effect. Kinetin and gibberellic acid both modified hook opening in light and dark; the former was inhibitory and the latter was stimulatory. Indoleacetic acid, at concentrations above 10⁻⁵ M, caused pronounced hook closing in red light but not in the dark. These effects were generally the same with both plants. In opening of the cotton hook, the cotyledons were not necessary as a light receptor tissue. None of the growth substances tested were able to substitute completely for the cotton cotyledon. Coumarin was a pronounced inhibitor of opening of the cotton hook, and this response was expressed to such a degree as to cause hook closure with bean tissue. Reduced oxygen levels inhibited hook opening in bean. Oxygen was required in processes subsequent to the light reaction, but not for the photochemical process.     

Control of Hypocotyl Hook Angle in Phaseolus mungo L. Role of Growth Substances

Effects of growth hormones on the hook angle and light responseof Phaseolus mungo L. hypocotyl hooks are described and theresults are discussed with reference to the functions of otherparts of the seedling in controlling the growth and shape ofthe hook. Apically applied IAA (indolyl acetic acid) prevented hook openingin decapitated seedlings in the dark and in all the red-irradiatedseedlings. [¹⁴C]IAA experiments showed that only a small quantityof IAA (2–6 ?g per hook) was required to produce theseeffects, and that transport of IAA through the hook was negligibleand unaffected by red irradiation. ABA (abscisic acid) had little effect on the hooks or theirlight response. Applied ethylene and IAA-induced ethylene slightly closed thehooks, but only slightly reduced light-induced opening. IAAreduced the effect of ethylene in the dark, but after irradiationthe hooks appeared more sensitive to the ethylene in the presenceof IAA, resulting in light-induced hook closure. Basally applied kinetin (6-furfurylaminopurine) prevented decapitatedhooks from opening in the dark, especially when GA₃ (gibberellicacid) was also present. Some combinations of kinetin and GA₃(with high kinetin concentrations) also prevented light-inducedopening, but combinations with lower kinetin concentrationsallowed almost as much opening as was found in intact hooks. It is proposed that the terminal parts act by regulating thesupply of cytokinins and gibberellins from the basal parts,and that IAA does not mediate this funotion in this species. The results are compared with those reported for other species.     

Photocontrol of the hypocotyl hook opening of Sinapis alba seedlings. Involvement of phytochrome and a high irradiance response

Baumgartner, N. and Fondeville, J. C. 1989. Photocontrol of the hypocotyl hook opening of Sinapis alba seedlings. Involvement of phytochrome and a high irradiance response.
A statistical evaluation of the hypocotyl hook opening (hook opening index) was used for measurement of the hook angle in lots of etiolated Sinapis alba L. cv. Albatros seedlings. Studies of the kinetics for hook opening were carried out in continuous fluorescent white, blue and red light (6, 15 and 40 μmol m^-2s^-1) with 2-day-old dark-grown seedlings. At the beginning of the irradiation period the photoresponse in red light was the opposite to that in blue (low photon fluences). Blue rapidly induced the hook opening (in less than 20 min), while red produced hook tightening (photon fluences up to 70 mmol m^-2), which precedes the normal progressive hook opening. For low fluences, the data were consistent with the involvement of phytochrome and a specific blue light photoreceptor. A phytochrome effect was observed in the hook opening, dependent upon a high irradiance response (HIR). This HIR (like that for the inhibition of the hypocotyl elongation) was characterized by a wavelength response curve with maxima in the blue and far-red regions of the spectrum.