Cyclic Nucleotides and In Vitro Plant Cultures: I. INDUCTION OF ORGANOGENESIS IN TOBACCO (NICOTIANA TABACUM CALLUS CULTURES

Mangat, B. S. and Janjua, S. 1987. Cyclic nucleotides and invitro plant cultures. I. Induction of organogenesis in tobacco(Nicotiana tabacum) callus cultures.—J. exp. Bot. 38:2059–2067. The possibility that cyclic nucleotides have a mediatory rolesimilar to cytokinins in plant tissue cultures was examined.Calli obtained from tobacco pith tissue were incubated on growthmedia supplemented with either cyclic AMP, cyclic GMP, adenosineor guanosine, in concentrations ranging from (mg dm^–3)0 to 2·0 together with 2·0 mg dm^–3 of IAA.Results were compared with identical calli grown on media containingcomparable amounts of kinetin and IAA. Increase in callus growthwas observed on all media containing cyclic AMP, cyclic GMP,adenosine, guanosine or kinetin. Adenosine or guanosine didnot promote organogenesis. Low concentrations (0·02 and0·05 mg dm^–3) of kinetin stimulated extensive rootdevelopment. Some root formation was also elicited with higheramounts of cyclic AMP (0·1 and 0·2 mg dm^–3)or cyclic GMP (0·2 and 0·5 mg dm^–3). Bothkinetin and cyclic GMP promoted shoot differentiation. However,in contrast to kinetin, cyclic GMP induced organogenesis atlower concentrations (0·02 and 0·1 mg dm^–3).The addition of 2·0 mg dm ^–3 of cyclic AM P toIAA-free growth media elicited shoot differentiation. This wasalso the case with a similar concentration of kinetin or cyclicGMP. Results suggest cytokinin activity for the two cyclic nucleotides. Key words: Tobacco, Nicotiana tabacum, tissue culture, cyclic nucleotides, cyclic AMP, cyclic GMP organogenesis     

The possible role of cyclic AMP in the control of genetic tumor induction

Young seedlings of the tumor-prone amphiploid Nicotiana suaveolensx N. langsdorffii, grown aseptically on nutrient medium, weretreated with 1x10^–2 M cyclic adenosine 3': 5'-monophosphate(cyclic AMP). The incidence of tumor formation was scored atregular intervals subsequent to exposure. Cyclic AMP causeda significant reduction in the rate of tumor formation. In addition,untreated plants grown on nutrient medium were harvested atregular intervals after the seeds had been sown. Cyclic AMPwas extracted, partially purified, and assayed radioimmunologically.The endogenous level of cyclic AMP in stem tissue was highestin young seedlings, rapidly fell reaching a low point in 18day-old plants, and eventually leveled off. The presence ofindoleacetic acid (IAA) in the growth medium at a final concentrationof 2x10^–5 M prevented the decline in cyclic AMP that occurredin seedlings grown on unsupplemented medium. (Received May 21, 1976; )     

Removal by chemicals of photoperiodic light requirements of Lemna gibba G3

Both the initial and the terminal 1 hr portions of the subjectiveday fraction, namely the L1- arid L2-phases, of a 24 hr daymust be illuminated in order for the day to be perceived asa long day in the min-LD determination by the long-day plant,Lemna gibba G3 (9). The light requirement of the L1-phase wassatisfied by a 10 min red light pulse given at the beginningof the phase. The red light effect was erased by a subsequent10 min far-red light, indicating phytochrome-mediated processesoccurring in the L1-phase. The light requirement of the L2-phasewas satisfied by blue or far-red light given during the terminal10 min period of the phase; there was no indication of phytochromeinvolvement. The light action on the L1-phase was replaced by10^–5 M of cyclic AMP or 10^–7 M of DL-isoproterenol.The isoproterenol action was antagonized by 10^–7 M ofDL-propranolol. Cyclic AMP (10^–5 M) combined with salicylicacid (10^–6 M), which can remove the light requirementof the L2-phase (10), rendered a completely dark day physiologicallyequivalent to a long day. Acetylcholine (10^–5 M) exertednyctomimetic action on the L1-phase of the second light day.The action of acetylcholine was antagonized by cyclic AMP (10^–5M). The L2-phase required no light in the presence of 10^–7M of DL-propranolol, and this propranolol action was not affectedby isoproterenol. These findings suggest changes in membranepermeability caused by the light given during the L1- and L2-phases. (Received July 7, 1976; )     

A possible mechanism for sugar inhibition of duckweed flowering

Sugar (ca. 1%) reduced the floral index in Lemna gibba G3 byca. 50%, and supplemental addition of cyclic AMP (ca. 10^–5M)removed nearly all of this sugar action. Inhibition by sugarof duckweed flowering may be explained in terms of cataboliterepression. (Received October 9, 1971; )     

Negative Phototropism in Vaucheria terrestris Regulated by Calcium III. The Role of Calcium Characterized by Use of a High-Power Argon-Ion Laser as the Source of Unilateral Blue Light

A tip-growing Xanthophycean algal coenocyte, Vaucheria terrestrissensu Gtz, is able to change the sign of its phototropic responsefrom positive to negative as a result of its ability to sensethe fluence rate (=intensity) of unilateral blue light (BL).The mechanism that determines the sign of phototropism was investigatedusing a high-power argon-ion laser (457.9 nm) as a source ofvery strong unilateral BL. The fluence-response relationshipwas determined by changing both the fluence rate and the durationof irradiation. Positive phototropic bending was induced whenthe fluence rate of BL from the laser was below 60 W m^–2.The positive bending obeyed the reciprocity law and was notaffected by the concentrations of external Ca²⁺ ions between0.4 mM and 4.4 mM. The positive curvature decreased when thealga was exposed to a unilateral pulse of BL with a durationof 10–300 s at fluence rates higher than 60 W m^–2.The alga finally showed a deep negative curvature when eitherthe fluence rate or the duration of irradiation was furtherincreased. The inversion of the phototropic response and developmentof the negative phototropic response was greatly enhanced inthe presence of 4.4 mM Ca²⁺ ions. However, the mechanism thatdetermine the sign of phototropism seemed to require a BL pulseof longer than several seconds, even when the fluence rate wassufficiently high. The role of cytoplasmic Ca²⁺ ions in positiveand negative phototropic responses is discussed. ¹This study was carried out as part of NIBB Cooperative ResearchProgram for the Okazaki Large Spectrograph (89-513 and 90-518). ²Part of this study was reported at the XXXII Yamada Conferenceon Plant Cell Walls as Biopolymers with Physiological Functions,May 5–8, 1992, Osaka (Kataoka and Watanabe 1992).     

Negative Phototropism of Vaucheria terrestris Regulated by Calcium II. Inhibition by Ca2+-Channel Blockers and Mimesis by A23187

As reported in a previous article [Kataoka (1988a) Plant CellPhysiol. 29: 1323], growing apices of the xanthophycean coenocyticalga, Vaucheria terrestris, bends away from a unilateral bluelight (BL) source, if they are simultaneously irradiated withstrong background BL in a solution containing 1–4 mM Ca²⁺.Since the negative bending is a function of the product of theexternal Ca²⁺ concentration and the fluence rate of backgroundBL, a BL-induced Ca²⁺-influx at the apex was hypothesized tobe the cause of the phototropic inversion. The present reportprovides strong evidence for this hypothesis. Addition of theCa²⁺ channel blockers, La³⁺, verapamil, nifedipine and nitrendipineto media containing 4 mM Ca²⁺ completely inhibited the phototropicinversion. By contrast, 1 µM A23187 [GenBank] (plus 4 mM Ca²⁺) notonly enhanced the phototropic inversion under background BL,but also mimicked the background BL; i.e. it caused negativebending under safe red light. Inhibition of phototropic inversionby La³⁺ is also observed under conditions where the algae areirradiated with unilateral BL for 1 week. A BL-dependent Ca²⁺influx and a consequential elevation of the cytoplasmic Ca²⁺-levelin the apical growth region must be involved in the early stepsof phototropic response. A BL-controlled opening of L-type Ca²⁺channels is also suggested. ¹A part of this study was reported at the 3rd Phycological Congressat Melbourne 1988 (Kataoka 1988b) and XXII Yamada Conferenceon Plant Water Relations and Growth Under Stress at Osaka 1989(Kataoka 1989) ²Dedicated to Prof. em. Dr. Noburo Kamiya on the occasion ofhis 77th birthday (Received May 23, 1990; Accepted July 13, 1990)     

Calmodulin-Like Activity Associated with Chromatin from Pea Buds

NAD kinase and cyclic AMP phosphodiesterase were activated bya factor prepared from pea chromatin. About 62% of the originalamount of the factor in the purified chromatin was recoveredin the reassociated chromatin. The NAD kinase- and cyclic AMP phosphodiesterase-activatingfactor was released from the chromatin by heat treatment withethylene glycol-bis(ß-aminoethyl ether)- N,N,N',N'-tetraaceticacid (EGTA) then adsorbed on an affinity gel of phenothiazineagarosederivatives in the presence of excess Ca²⁺ over EGTA, afterwhich it was eluted by a flush of EGTA. Activation of NAD kinaseand cyclic AMP phosphodiesterase by this factor depended onthe presence of Ca²⁺. The NAD kinase-activating factor and chromatin were coelutedwhen soluble chromatin was applied to a Bio-Gel A50 column.When chromatin was chromatographed on the same column afterdigestion by DNase I, the factor was eluted in association withthe digested products of the chromatin. The activation propertiesof this factor indicate that a calmodulin-like activity existsin association with pea chromatin. The activation curves of cyclic AMP phosphodiesterase with thepea bud factor and with bovine brain calmodulin were compared.The amount of the factor in the chromatin fraction that correspondedto authentic calmodulin was calculated as 5.7 µg per mgDNA. (Received August 6, 1982; Accepted February 17, 1983)     

Swelling of Etiolated Oat Protoplasts Induced by cAMP and Red Light

Etiolated oat protoplasts were treated with dibutyryl cAMP tostudy possible function of cAMP in the development by measuringthe protoplast swelling. The mean diameter of protoplasts inthe absence of any chemical treatment was 33.58±1.26(SE) µm, which increased to 36.96±0.86 µmin the presence of 100 µM dibutyryl cAMP. Prostacyclin,a potent activator of adenyl cyclase, also showed a significantswelling effect (diameter 38.01±0.98 µm). Red lightalso elicited the swelling of protoplasts (40.26±0.8µm). ¹Present address: Department of Biology, Pusan National University,Pusan 607, Korea. ²Present address: Department of Horticulture, Cheju NationalUniversity, Cheju 590, Korea. ³Present address: Department of Biological Sciences, Texas TechUniversity, Lubbock, TX 79409, U.S.A. (Received June 29, 1985; Accepted November 18, 1985)     

Phototropism in a red alga, Griffithsia pacifica

In the red alga, Griffithsia pacifica, shoot portions of a plantare positively phototropic and rhizoids are negatively phototropic.We have studied the phototropic response of rhizoids which elongateby tip growth. For 45 min after the beginning of unilateralillumination a rhizoid grows straight, then phototropic curvaturebegins and continues rapidly until the rhizoid is growing awayfrom the light. Curvature is 70–80% complete after 3 hr.If the unilateral stimulus is given for a short time (15 min),curvature again begins at 45 min. However, within an additional30–45 min the rhizoid stops growing away from the lightand wanders back towards its original direction of growth. Phototropismis elicited by light of wavelengths from 350 nm to 500 nm; inlight of wavelengths above 550 nm, little, if any, responseoccurs. ¹Present address: Division of Natural Sciences, University ofCalifornia, Santa Cruz, California 95064, U.S.A. (Received December 10, 1976; )     

The Effect of Ethylene and Temperature on ATP Accumulation from Various Substrates in Peanut (Arachis hypogaea L. ) Seeds

Peanut (Arachis hypogaea L. ) seed powder accumulated ATP fromAMP and phosphoenolpyruvate (PEP) at a rate of approx. 100 pmolmin^–1mg^– powder at 35° C. When peanut seed powderwas incubated with various substrates, which may result in PEPor AMP (ADP) synthesis, then ATP accumulated. The best substratecombinations examined so far were AMP + succinate, NADH₂, andAMP + malate + NAD, with activities of 33, 12 and 12 pmol min^–mg^–1powder,respectively; AMP + malate showed very low activity. Some combinationsexhibited linear activities with time, while others had an exponential-typeprofile. The temperature dependence of the ATP accumulationdemonstrated by the Ahrrenius plot had a double phase with atransition point at 25° C. The Ea values between 15°C and 25° C were 25 000–50 000 cal/mol, while above25° C the Ea values fluctuated between 6000 and 8000 cal/mol(depending on the substrate). The AMP + PEP combination exhibiteda single-phase profile between 15° C and 40° C, withan Ea value of 22 000 cal/mol. In the presence of some substrates,ethephon (ethylene) had a stimulatory effect and caused an increasein the Ea values at the high temperature phase. A comparisonof seed powder from dormant seeds with that from non-dormantseeds revealed that some substrate combinations accumulate ATPfaster in non-dormant seeds and others do so in dormant seeds. Key words: Arachis hypogaea, ATP, Ethylene, Dormancy, Peanut, Seed     

Effect of Growth Regulators and Light on Pollen Germination and Pollen Tube Growth in Pinus roxburghii Sarg

Pine (Pinus roxburghii) pollen grown in suspension cultureswas used to study the effects of growth regulators and lightconditions on germination and pollen tube growth. Indol-3-ylacetic acid, gibberellic acid, ethylene, abscisic acid and cyclicAMP (cAMP) at low concentrations (1–10 mg 1^–1) promotedgermination and tube growth. Addition of 1 and 10 mg 1^–1cAMP to any of the growth regulators had a promotory effect.Pollen tube growth decreased in white light as compared to thedark, and was increased in red light. Far-red light counteractedthe effect of red light. The effect of growth regulators incausing the enhanced tube growth appears to be manifested throughsubstances such as cAMP, and phytochrome seems to be involved. Pinus roxburghii, pine, pollen germination, pollen tube growth, growth regulators, cyclic AMP, phytochrome     

Significance of Cyclic Nucleotides in Amaranthin Synthesis by Amaranthus caudatus Seedlings

Explants of 72–76 h old Amaranthus caudatus seedlingssynthesize the betalain pigment amaranthin in response to light.Light can be replaced with a cytokinin or a cyclic nucleotidewith an N⁶-substituent. Cyclic 3'5'-AMP shows only weak activityand that only at high unphysiological concentrations. Even cyclic2'3'-AMP, which docs not act as a ‘second messenger’,induces amaranthin synthesis to a greater degree than cyclic3',5'-AMP. But N⁶-monobutyryl-cyclic 3',5'-AMP and N⁶-2'-O-dibutyryl-cyclicAMPshow high activity, higher even than kinetin at its optimumconcentration of 10^–5 M. 2'-O-Monobutyryl-cyclicAMP, onthe other hand, is considerably less active, suggesting thatN⁶-substitution of the adenine ring is responsible for the enhancedactivity. N⁶-Propionyl, butyryl and valeryladenines are allhighly active, indicating that the cyclic monophosphate moietyis unnecessary for this response. All the compounds tested,including cyclic 3',5'-AMP, show additive effects, but thereis no amplification of the response, typical of second messengeraction. Inhibition of amaranthin synthesis imposed by hadacidin, isrelieved by kinetin, DBc AMP, N⁶-monobutyryl-cAMP and N⁶-butyryladenine. Cyclic 3',5'-AMP is weakly active in this regard. Asnatural cytokinins are N⁶-substituted adenine compounds, andas only N⁶-substituted cyclic nucleotides are able to mimicthe effect of cytokinin, it is concluded that these cyclic nucleotidesfunction as cytokinin analogues and not as ‘second messengers’'. Amaranthus caudatus, amaranthin, cytokinins, cyclic nucleotides     

Plasma Membrane H+-ATPase in Guard-Cell Protoplasts from Vicia faba L.

The activity of plasma membrane H⁺-ATPase was measured withmembrane fragments of guard-cell protoplasts isolated from Viciafaba L. ATP hydrolytic activity was slightly inhibited by oligomycinand ammonium molybdate, and markedly inhibited by NO₃^–and vanadate. In the presence of oligomycin, ammonium molybdateand NO₃^–, the ATP-hydrolyzing activity was strongly inhibitedby vanadate. It was also inhibited by diethylstilbestrol (DES),p-chloromercuribenzoic acid (PCMB) and Ca²⁺, but slightly stimulatedby carbonyl cyanide m-chlorophenylhydrazone (CCCP). The acitivityhad higher specificity for ATP as a substrate than other phosphoricesters such as ADP, AMP, GTP and p-nitrophenylphosphate; theK_m was 0.5 mM for ATP. The activity required Mg²⁺ but was notaffected by K⁺, and it was maximal around pH 6.8. When guard-cellprotoplasts were used instead of membrane fragments, the ATPaseactivity reached up to 800µmol Pi.(mg Chl)^–1.h^–1in the presence of lysolecithin. These results indicate thatthe guard cell has a high plasma membrane H⁺-ATPase activity. (Received December 23, 1986; Accepted April 28, 1987)     

Reversal of sucrose inhibition of Lemna flowering by adenine derivatives

Sucrose inhibition of flowering in Lemna perpusilla 6746 wasat least partially reversed by 5'-AMP, cyclic 3',5'-AMP, 5'-ADP,5'-ATP and K₂HPO₄. These results are in contrast to those reportedfor L. gibba in which reversal was effected by cyclic AMP, butnot by other adenine derivatives. ¹ This work was supported by National Science Foundation GrantGB-12955. (Received June 11, 1973; )     

Effects of Electrolyte and Non-Electrolyte Solutions on the Tropic Responses of Avena Coleoptiles

Solutions of mannitol and carbowax 1540 reduced the geotropicresponse of Avena sativa coleoptiles to at least the same extentas a potassium chloride solution of equal osmotic potential.Similar results were obtained with the phototropic response.The magnitude of these decreases increased almost linearly withthe osmotic concentration of the solutions over the range 0.025–0.45J cm^–3. In contrast, the geotropic and phototropic responseswere scarcely affected by exposing the coleoptiles to solutionsof glycerol. The absence of an effect with glycerol is probably due to thepenetration of the coleoptile cells by this solute. The similaror greater reduction in tropic curvature brought about by mannitoland carbowax 1540 solutions, as compared with a potassium chloridesolution of equal osmotic potential, makes untenable a previoussuggestion attributing the inhibitory effects of electrolytesolutions to a shunting of the electro-potential differenceson the plant surface.     

Photosynthetic electron transport in Dunaliella tertiolecta (Chlorophyceae) measured by fast repetition rate fluorometry: relation to carbon assimilation

A comparison of photosynthesis-irradiance response curves (P–Eresponse curves) obtained through fast repetition rate (FRR)fluorometry and radiocarbon (¹⁴C) tracer method was made inthe chlorophyte, Dunaliella tertiolecta, grown under differentirradiance conditions. In FRR-based P–E response curveexperiments, actinic light provided by white light-emittingdiodes (LEDs) was increased gradually from 0 to 1500 µmolquanta m^–2 s^–1 and the rate of photosyntheticelectron transport was determined at each light level. Short-termexperiments (20 min) of ¹⁴C-based P–E response curvewere carried out with an improved photosynthetron, which containswhite LEDs as the light source. Irrespective of growth irradiance,the ratios of FRR to ¹⁴C-based initial slopes were almost uniform.The ratios of FRR- to ¹⁴C-based maximum rates were 25–36%higher than those of FRR- to ¹⁴C-based initial slopes. The relationshipbetween electron transport and carbon assimilation was non-linearwith increasing discrepancy towards high actinic light. Thisnon-linear relationship between FRR- and ¹⁴C-based estimatesis primarily due to the effect of physiological processes stimulatedat high levels of light, such as cyclic electron flow and theMehler reaction. The results of this study indicate that theFRR fluorometry can be used as a good indicator of photosyntheticrates from low to middle light levels, but becomes increasinglyquestionable as the maximum photosynthetic rate is approached.The degree to which this relationship is further affected bynutrient-status warrants investigation.     

Effects of Cyclic AMP on the Activity of Soluble Protein Kinases in Lemna paucicostata

Three protein kinases which phosphorylate histone were isolatedfrom cellular extract of Lemna plants. They were separated byelution from DEAE-Sephacel column and referred to as PI, PITand PHI. The PI protein kinase activity was partially inhibitedby 10µM cyclic AMP, cyclic GMP or cyclic IMP, while thePII enzyme was activated in the presence of these cyclic nucleotides.The PIII enzyme was cAMPindependent, but slightly inhibitedby cyclic CMP and cyclic UMP. The molecular weights of thesethree protein kinases were 165,000, 85,000 and 145,000, respectively,as estimated from Sephacryl S-300 gel filtration. A single cyclicAMP-binding protein was detected in the PII enzyme fractionby using the photoaffinity cAMP-analogue, 8-N₃-cAMP. The proteinwhich specifically bound [³H]-8-N₃-cAMP had an apparent molecularweight of 48,000 as determined by SDS-polyacrylamide gel electrophoresis.The phosphorylation of cellular proteins in Lemna was examinedby SDS-polyacrylamide gel electrophoresis. Four phosphorylatedpolypeptides were detected, the phosphorylations of which werestimulated by cAMP. The molecular weights of these four polypeptideswere 59,000, 19,000, 16,000 and 14,000, respectively. (Received January 26, 1983; Accepted April 13, 1983)     

Involvement of Cyclic AMP in ABA- and Ca2--Mediated Signal Transduction of Stomatal Regulation in Vicia faba

The focus of this study is to investigate possible involvementof cyclic AMP in regulation of Vicia stomatal movements. Thepresence of 0.1 mM 8-Br-cAMP, a membrane-permeable analogueof cAMP, alone in the incubation medium did not affect stomatalopening in the light in leaf epidermal peel experiments. However,addition of 0.1 mM 8-Br-cAMP completely reversed exogenous ABA-and C^a2+-induced inhibition of stomatal opening. Consistentwith these results, patch-clamping experiments showed that intracellularaddition of 0.5 mM or 1 mM cAMP significantly reversed the inhibitionof whole-cell inward K⁺ currents by internally supplied 13 µMCa²⁺ or 10 µM ABA in stomatal guard cell protoplasts,respectively. Furthermore, intracellular addition of either10 µM prostaglandin E₁ (PGE1, an adenylate cyclase activator)or 1 mM 3-isobutyl-1-methylxanthine (IBMX, a phosphodiesteraseinhibitor) mimicked the effect of exogenous cAMP on the removalof ABA- or Ca²⁺ inhibition of inward K⁺-current. These resultssuggest that a cAMP signaling pathway is involved in signaltransduction in stomatal regulation by interacting with ABAand Ca²⁺ signaling cascades. A hypothetical mechanism by whichcAMP may regulate K⁺ in stomatal guard cells is also discussed. (Received May 6, 1999; Accepted August 27, 1999)     

Photoinhibition and light-induced cyclic electron transport in ndhB(-) and psaE(-) mutants of Synechocystis sp. PCC 6803

The ndhB^– and psaE^– mutants of the cyanobacteriumSynechocystis sp. PCC 6803 are partly deficient in PSI-drivencyclic electron transport. We compared photoinhibition in thesemutants to the wild type to test the hypothesis that PSI cyclicelectron transport protects against photoinhibition. Photoinhibitorytreatment greatly accelerated PSI cyclic electron transportin the wild type and also in both the mutants. The psaE^–mutant showed rates of PSI cyclic electron transport similarto the wild type under all conditions tested. The ndhB^–mutant showed much lower rates of PSI cyclic electron transportthan the wild type following brief dark adaptation but exceededwild type rates after exposure to photoinhibitory light. Thewild type and both mutants showed similar rates of photoinhibitiondamage and photoinhibition repair at PSII. Photoinhibition atPSI was much slower than at PSII and was also similar betweenthe wild type and both mutants, despite the known instabilityof PSI in the psaE^– mutant. We conclude that photoinhibitorylight induces sufficient PSI-driven cyclic electron transportin both the ndhB^– and psaE^– mutants to fulfill anyrole that cyclic electron transport plays in protection againstphotoinhibition. ⁴ Corresponding author: E-mail, sherbert@uwyo.edu; Fax, +1-307-766-2851;Phone, +1-307-766-4353.     

Variations in the Amounts of 3', 5'-Cyclic AMP in Plant Tissues5

Tissue concentrations of 3', 5'-cyclic AMP have been measuredin soybean callus in liquid suspension. After transfer to freshmedium there was a sharp decline in cyclic-AMP concentrationwithin 30 min. This effect was much reduced if sucrose was omittedfrom the new medium. The presence of cytokinin had no effecton the fluctuations observed during the 20 h immediately followingsubculture but after that time cytokinin tended to produce muchhigher concentrations of cyclic AMP. In Avena coleoptiles theapplication of auxin caused a sharp increase in cyclic-AMP concentrationwithin 15 min. The amount of cyclic AMP in auxin-treated tissueremained higher than the values obtained for untreated samplesover the next 3 h. Neither cyclic AMP nor N⁶, O²-dibutyryl cyclicAMP had any significant effect on the rate of coleoptile extensiongrowth.