Plasma membrane protein composition and synthesis in soybean hypocotyl segments undergoing auxin-induced elongation

Summary The types and amount of plasma membrane proteins synthesized during cell elongation in response to auxin (2,4-dichlorophenoxyacetic acid) treatment were investigated. Auxin-treated and control soybean (Glycine max L.) hypocotyl segments were incubated with [³⁵S]methionine for various times, ranging from 0.5 to 18 h, prior to isolation of plasma membrane by aqueous two-phase partitioning. Protein accumulated in the plasma membrane after auxin treatment. Despite this accumulation, the protein incorporation rate, estimated by the amount of label in the plasma membrane following a 0.5 h [³⁵S]methionine pulse, was unaffected by auxin treatment at both 0.5 and 18 h of treatment. Protein apparently accumulated by a mechanism distinct from enhanced incorporation. The plasma membrane proteins synthesized by elongating segments differed from controls at 18 h, as evidenced by the pattern of fluorographs following a 0.5 h radiolabelling. However, auxin treatment did not alter the 2-D gel pattern of the polypeptides detectable by silver stain.Abbreviations 2,4-D 2,4-dichlorophenoxyacetic acid - IEF isoelectric focusing - PM plasma membrane - SDS-PAGE sodium dodecyl sulfate-polyacrylamide gel electrophoresis     

Plasma membrane phospholipid and sterol synthesis in soybean hypocotyl segments undergoing auxin-induced elongation

Summary Auxin-induced cell elongation necessitates plasma membrane enlargement. The effect of auxin (10 M 2,4-dichlorophenoxyacetic acid) treatment on amount, composition, and rate of synthesis of plasma membrane lipids was examined. Auxin-treated and control soybean (Glycine max L.) hypocotyl segments were incubated with [¹⁴C]acetate for times ranging from 0.5 to 18 h, prior to isolation of plasma membrane by aqueous two-phase partitioning. The composition of individual plasma membrane lipids in elongating segments did not differ from the composition in treatment time-matched control segments, except that after longer auxin treatments, phospholipids had more unsaturated fatty acids. Plasma membrane phospholipid and free sterol content both increased in elongating segments. The relative proportion of sterols and phospholipids in the plasma membrane primarily depended on time after segment excision, for both auxin-treated and control segments. Auxin enhanced the rate of lipid incorporation into the plasma membrane by 6 h, and stimulated the synthesis of some phospholipids and sterols.Abbreviations 2,4-D 2,4-dichlorophenoxyacetic acid - ER endoplasmic reticulum - GC gas chromatography - IAA indole-3-acetic acid - PA phosphatidic acid - PC phosphatidylcholine - PE phosphatidylethanolamine - PG phosphatidylglycerol - PI phosphatidylinositol - PM plasma membrane - PS phosphatidylserine     

Inhibition of chlorophyll synthesis by kinetin in Cucumis cotyledons

Kinetin is shown to inhibit the synthesis of chlorophyll inlight while causing expansion of isolated cotyledons of Cucumis.The inhibitory effect is more marked at higher concentrationsof kinetin. It has been suggested that specific kinins may beinvolved in different tissues for stimulating chlorophyll synthesis. (Received August 14, 1969; )     

Detection of endogenous gibberellins and their relationship to hypocotyl elongation in soybean seedlings

Four gibberellins, GA₅₃, GA₁₉, GA₂₀, and GA₁, were detected by bioassay, chromatography in two HPLC systems, and combined gas chromatography-mass spectroscopy-selected ion monitoring (GC-MS-SIM) in etiolated soybean (Glycine max [L.] Merr.) hypocotyls. GC-MS-SIM employed [²H₂]-labeled standards for each endogenous gibberellin detected, and quantities estimated from bioassays and GC-MS-SIM were similar. This result plus the tentative detection of GA₄₄ and GA₈ (standards not available) indicates that the early-C-13-hydroxylation pathway for gibberellin biosynthesis predominates in soybean hypocotyls. Other gibberellins were not detected. Growth rates decreased after transfer to low water potential (ψ_w) vermiculite and were completely arrested 24 hours after transfer. The GA₁ content in the elongating region of hypocotyls had declined to 38% of the 0 time value at 24 hours after transfer to low ψ_w vermiculite, a level which was only 13% of the GA₁ content in control seedlings at the same time (24 hours posttransfer). Rewatering seedlings following 24 hours growth in low ψ_w vermiculite resulted in a complete recovery in elongation rate, an increase in GA₁ (20% at 2 hours, two-fold at 8 hours, eightfold at 24 hours), and a decrease in ABA levels (tenfold at 2 hours). Treatment of well-watered seedlings with the GA-synthesis inhibitor tetcyclacis (TCY) resulted in lowered GA₁ levels and increased ABA levels. When seedlings grown 24 hours in low ψ_w vermiculite were rewatered with TCY, recovery of the elongation rate was delayed and reduced, and the decline in ABA levels was slowed. Addition of GA₃ restored the elongation rate inhibited by TCY. Seedlings were growth responsive to exogenous GA₃, and this GA₃-promoted growth was inhibited by exogenous ABA. The data are consistent with the hypothesis that changes in GA₁ and ABA levels play a role in adjusting hypocotyl elongation rates. However, the changes observed are not of sufficient magnitude nor do they occur rapidly enough to suggest they are the primary regulators of elongation rate responses to rapidly changing plant water status.     

Influence of auxin and incubation on the relative level of polyribosomes in excised soybean hypocotyl

The influence of incubation and auxin (2,4-D) on polyribosome level in soybean hypocotyl was studied.     

Change in XET activities,cell wall extensibility and hypocotyl elongation of soybean seedlings at low water potential

In dark-grown soybean (Glycine max [L.] Merr.) seedlings, exposing the roots to water-deficient vermiculite (_w=–0.36 MPa) inhibited hypocotyl (stem) elongation. The inhibition was associated with decreased extensibility of the cell walls in the elongation zone. A detailed spatial analysis showed xyloglucan endotransglucosylase (XET; EC 2.4.1.207) activity on the basis of unit cell wall dry weight was decreased in the elongation region after seedlings were transplanted to low _w. The decrease in XET activity was at least partially due to an accumulation of cell wall mass. Since cell number was only slightly altered, wall mass had increased per cell and probably led to increased wall thickness and decreased cell wall extensibility. Alternatively, an increase in cell wall mass may represent a mechanism for regulating enzyme activity in cell walls, XET in this case, and therefore cell wall extensibility. Hypocotyl elongation was partially recovered after seedlings were grown in low-_w vermiculate for about 80 h. The partial recovery of hypocotyl elongation was associated with a partial recovery of cell wall extensibility and an enhancement of XET activity in the hypocotyl elongation zone. Our results indicate XTH proteins may play an important role in regulating cell wall extensibility and thus cell elongation in soybean hypocotyls. Our results also showed an imperfect correlation of spatial elongation and XET activity along the hypocotyls. Other potential functions of XTH and their regulation in soybean hypocotyl growth are discussed.     

Cell elongation and cell division in elongating lettuce hypocotyl sections

The roles of cell division and cell elongation in the growth of sections excised from hypocotyls of lettuce (Lactuca sativa L. cv. Arctic) were investigated. Elongation of sections incubated in the light is inhibited compared to dark-grown sections and this inhibition is reversed by gibberellic acid (GA₃). The elongation of both dark-grown and GA₃-treated, light-grown sections can be enhanced by 10mM KCl. Under all conditions of incubation, elongation growth is greatest in the uppermost quarter of the hypocotyl section while the basal quarter does not elongate. In darkness the two apical segments of sections marked into four equal parts grow at the same rate, while in light, growth of the apical segment exceeds that of the second segment. Cell division in cortical or epidermal cells, as measured by mitotic index or cell number, is not affected by illumination conditions nor by GA₃ or KCl treatments. Although -irradiation and FUDR pretreatment eliminate or cause a marked reduction in cell division in the excised hypocotyl, sections from seeds irradiated with -rays or incubated in 5-fluorodeoxyuridine elongate in response to GA₃ and KCl treatment as do sections from non-pretreated controls. Therefore, since neither GA₃ nor darkness affect celldivision activity and since treatments which eliminate or significantly reduce cell division do not affect growth, we conclude that the effect of GA₃ and darkness in this material is to increase cell elongation.Abbreviations FUDR 5-fluorodeoxyuridine - GA(s) gibberellin(s) - GA₃ gibberellic acid     

Metabolism of kinetin by soybean callus

Soybean callus metabolised applied 6-furfurylamino (8^-14C) purine very rapidly to polar compounds, some of which were retained on a Dowex 50 cation exchange resin, and were unaffected by alkaline phosphatase; while others were apparently phosphorylated and were detected in the aqueous fraction. Adenine was detected as an intermediate and it can be concluded that it was formed as a result of the rapid and efficient removal of the furfuryl side chain. The formed adenine was rapidly incorporated into the polar metabolites. Exactly how the presence of this cytokinin stimulates cell division is not apparent from the results.The financial support of the C.S.I.R., Pretoria, and The Natal University Development Fund is gratefully acknowledged.     

Gibberellins and auxin regulate soybean hypocotyl elongation under low light and high-temperature interaction

Soybean is an important oilseed crop grown globally. However, two examples of environmental stresses that drastically regulate soybean growth are low light and high-temperature. Emerging evidence suggests a possible interconnection between these two environmental stimuli. Low light and high-temperature as individual factors have been reported to regulate plant hypocotyl elongation. However, their interactive signal effect on soybean growth and development remains largely unclear. Here, we report that gibberellins (GAs) and auxin are required for soybean hypocotyl elongation under low light and high-temperature interaction. Our analysis indicated that low light and high-temperature interaction enhanced the regulation of soybean hypocotyl elongation and that the endogenous GA₃, GA₇, indole-3-acetic acid (IAA), and indole-3-pyruvate (IPA) contents significantly increased. Again, analysis of the effect of exogenous phytohormones and biosynthesis inhibitors treatments showed that exogenous GA, IAA, and paclobutrazol (PAC), 2, 3, 5,-triiodobenzoic acid (TIBA) treatments significantly regulated soybean seedlings growth under low light and high-temperature interaction. Further qRT-PCR analysis showed that the expression level of GA biosynthesis pathway genes (GmGA3ox1, GmGA3ox2 and GmGA3) and auxin biosynthesis pathway genes (GmYUCCA3, GmYUCCA5 and GmYUCCA7) significantly increased under (i) low light and high-temperature interaction and (ii) exogenous GA and IAA treatments. Altogether, these observations support the hypothesis that gibberellins and auxin regulate soybean hypocotyl elongation under low light and high-temperature stress interaction.     

Inhibition of auxin-induced cell elongation by galactose

Galactose at concentrations higher than 3 m M inhibited specifically auxin-induced elongation of oat, wheat and rice coleoptile segments but not of pea and mung bean stem segments. Glucose, arabinose, rhamnose, xylose, mannose and glucosamine did not inhibit auxin-induced elongation of coleoptile segments. Galactose inhibited auxin-induced but not hydrogen ion-induced growth.     

Arabidopsis SMALL AUXIN UP RNA63 promotes hypocotyl and stamen filament elongation

Auxin regulates plant growth and development in part by activating gene expression. Arabidopsis thaliana SMALL AUXIN UP RNAs (SAURs) are a family of early auxin-responsive genes with unknown functionality. Here, we show that transgenic plant lines expressing artificial microRNA constructs (aMIR-SAUR-A or -B) that target a SAUR subfamily (SAUR61-SAUR68 and SAUR75) had slightly reduced hypocotyl and stamen filament elongation. In contrast, transgenic plants expressing SAUR63:GFP or SAUR63:GUS fusions had long hypocotyls, petals and stamen filaments, suggesting that these protein fusions caused a gain of function. SAUR63:GFP and SAUR63:GUS seedlings also accumulated a higher level of basipetally transported auxin in the hypocotyl than did wild-type seedlings, and had wavy hypocotyls and twisted inflorescence stems. Mutations in auxin efflux carriers could partially suppress some SAUR63:GUS phenotypes. In contrast, SAUR63:HA plants had wild-type elongation and auxin transport. SAUR63:GFP protein had a longer half-life than SAUR63:HA. Fluorescence imaging and microsomal fractionation studies revealed that SAUR63:GFP was localized mainly in the plasma membrane, whereas SAUR63:HA was present in both soluble and membrane fractions. Low light conditions increased SAUR63:HA protein turnover rate. These results indicate that membrane-associated Arabidopsis SAUR63 promotes auxin-stimulated organ elongation.     

Auxin-gibberellin relationships in their effects on hypocotyl elongation of light-grown cucumber seedlings VI. Promotive and inhibitory effects of kinetin

The interaction of kinetin with IAA and GA₃ on the elongationof hypocotyl sections of Cucumis sativus L. cv. National Picklingwas studied. Kinetin in the concentration range of 10^–7M to 10^–4 M markedly inhibited IAA-induced elongation,while in a lower range from 10^–10 M to 10^–8 M, itsynergistically enhanced IAA-induced elongation. Kinetin alonein this range had no effect. A 5-to 15-min pulse treatment seemsenough to induce the maximum effect for both inhibition andpromotion. Since the magnitude of the maximum inhibition dependedon the concentration and not on the duration of treatment, thereaction in the cell caused by kinetin seemed to be completedwithin a short period. Washing of the sections with distilledwater after kinetin treatment (30 min) did not significantlyeliminate the kinetin effect. This probably indicates that thebinding of kinetin molecules to a supposed acceptor is not reversible.Interaction of kinetin with GA₃ in their pretreatment effectson IAA-induced elongation shows that in the inhibitory concentrationrange, the kinetin effect was partly overcome by GA₃, and thatin the promotive range, the magnitude of the enhancement wasdetermined by kinetin regardless of the presence of GA₃. Theeffect of kinetin seems to dominate over that of GA₃ indicatingthat the modes of their pretreatment effects differ from oneanother. (Received June 24, 1977; )     

Inhibition of hypocotyl elongation by ultraviolet-B radiation in de-etiolating tomato seedlings. I. The photoreceptor

Broad-band UV-B radiation inhibited hypocotyl elongation in etiolated tomato ( Lycopersicon esculentum Mill. cv. Alisa Craig) seedlings. This inhibition could be elicited by < 3 μmol m⁻² s⁻¹ of UV-B radiation provided against a background of white light (> 620 μmol m⁻² s⁻¹ between 320 and 800 nm), and was similar in wild-type and phytochrome-1-deficient aurea mutant seedlings. These observations suggest that the effect of UV-B radiation is not mediated by phytochrome. An activity spectrum obtained by delivering 1 μmol m⁻² s⁻¹ of monochromatic UV radiation against a while light background (63 μmol m⁻² s⁻¹ showed maximum effectiveness around 300 nm, which suggests that DNA or aromatic residues in proteins are not the chromophores mediating UV-B induced inhibition of elongation. Chemicals that affect the normal (photo)chemistry of flavins and possibly pterins (KI, NaN, and phenylacetic acid) largely abolished the inhibitor) effect of broad-hand UV-B radiation when applied to the root zone before irradiation. KI was effective at concentrations < 10⁻⁴ M , which have been shown in vitro to be effective in quenching the triplet excited stales of flavins but not fluorescence from pterine or singlet states of flavins. Elimination of blue light or reduction of UV-A, two sources of flavin excitation, promoted hypocotyl elongation, but did not affect the inhibition of elongation evened by UV-B. Kl applied after UV-B irradiation had no effect on the inhibition response. Taken together these findings suggest that the chromophore of the photoreceptor system invoked in UV-B perception by tomato seedlings during de-etiolation may be a flavin.     

Inhibition by ricin of protein synthesis in vitro. Inhibition of the binding of elongation factor 2 and of adenosine diphosphate-ribosylated elongation factor 2 to ribosomes.

The binding of EF2 (elongation factor 2) and of ADP-ribosyl-EF 2 to rat liver ribosomes is inhibited by ricin. This result suggests that the native enzyme and its ADP-ribose derivative have the same or closely related binding sites on the ribosome. The inhibition by ricin of the binding of EF 2 to ribosomes is consistent with the previous observation that ricin affects EF 2-catalysed translocation during polypeptide chain elongation.     

Brassinazole represses tomato hypocotyl elongation via inhibition of cell division

Plant Growth Regulation - Hypocotyl length is determined by cell division and elongation, which are associated with canonical cell cycle and endoreduplication. Plant hormone brassinosteroid (BR) is...     

Inhibition of arabidopsis hypocotyl elongation by jasmonates is enhanced under red light in phytochrome B dependent manner

Jasmonates are phytohormones derived from oxygenated fatty acids that regulate a broad range of plant defense and developmental processes. In Arabidopsis, hypocotyl elongation under various light conditions was suppressed by exogenously supplied methyl jasmonate (MeJA). Moreover, this suppression by MeJA was particularly effective under red light condition. Mutant analyses suggested that SCF^COI1-mediated proteolysis was involved in this function. However, MeJA action still remained in the coi1 mutant, and (+)-7-iso-JA-L-Ile, a well-known active form of jasmonate, had a weaker effect than MeJA under the red light condition, suggesting that unknown signaling pathway are present in MeJA-mediated inhibition of hypocotyl elongation. EMS mutant screening identified two MeJA-insensitive hypocotyl elongation mutants, jasmonate resistance long hypocotyl 1 (jal1) and jal36, which had mutations in the phytochrome B (PHYB) gene. These analyses suggested that inhibition of hypocotyl elongation by jasmonates is enhanced under red light in phyB dependent manner.