Auxin Physiology of the Tomato Mutant diageotropica

The tomato (Lycopersicon esculentum, Mill.) mutant diageotropica (dgt) exhibits biochemical, physiological, and morphological abnormalities that suggest the mutation may have affected a primary site of auxin perception or action. We have compared two aspects of the auxin physiology of dgt and wild-type (VFN8) seedlings: auxin transport and cellular growth parameters. The rates of basipetal indole-3-acetic acid (IAA) polar transport are identical in hypocotyl sections of the two genotypes, but dgt sections have a slightly greater capacity for IAA transport. 2,3,5-Triiodobenzoic acid and ethylene reduce transport in both mutant and wild-type sections. The kinetics of auxin uptake into VFN8 and dgt sections are nearly identical. These results make it unlikely that an altered IAA efflux carrier or IAA uptake symport are responsible for the pleiotropic effects resulting from the dgt mutation. The lack of auxin-induced cell elongation in dgt plants is not due to insufficient turgor, as the osmotic potential of dgt cell sap is less (more negative) than that of VFN8. An auxin-induced increase in wall extensibility, as measured by the Instron technique, only occurs in the VFN8 plants. These data suggest dgt hypocotyls suffer a defect in the sequence of events culminating in auxin-induced cell wall loosening.     

Insensitivity of the diageotropica tomato mutant to auxin

The sensitivity of excised hypocotyl segments to indoleacetic acid (IAA) in two assays, ethylene production and elongation, was determined in the ethylene-requiring tomato (Lycopersicon esculentum Mill.) mutant, diageotropica (dgt), and its isogenic parent, cv VFN8. Endogenous (uninduced) ethylene synthesis rates were slightly lower in dgt hypocotyls than in VFN8 hypocotyls. Ethylene production was essentially unaffected by IAA in dgt, but was stimulated up to 10-fold by 10 micromolar IAA in VFN8. Elongation of dgt hypocotyls was also insensitive to concentrations of IAA as high as 100 micromolar, as compared to significant elongation of VFN8 hypocotyls in response to 0.1 micromolar IAA. A range of IAA analogs active in VFN8 was also ineffective in stimulating elongation of dgt hypocotyls, suggesting that the differences were not due to rapid metabolism of IAA by dgt tissues. Auxin-induced elongation of VFN8 hypocotyls was unaffected by 2,3,5-triiodobenzoic acid and naphthylphthalamic acid, indicating that polar auxin transport was not a factor in these experiments. Exogenous and auxin-induced ethylene had no effect on the elongation respone of either genotype, nor did exogenous ethylene restore the sensitivity of dgt hypocotyls to IAA. Despite their apparent insensitivity to auxin, dgt hypocotyls elongated dramatically and synthesized ethylene rapidly in response to 1.2 micromolar fusicoccin. These results suggest that the primary effect of the dgt mutation is to reduce the sensitivity of the tissue to auxin. As altered regulation of ethylene synthesis is only one symptom of this fundamental deficiency, dgt should more properly be considered to be the auxin-insensitive tomato mutant.     

Cytokinin inhibits a subset of diageotropica-dependent primary auxin responses in tomato

Many aspects of plant development are regulated by antagonistic interactions between the plant hormones auxin and cytokinin, but the molecular mechanisms of this interaction are not understood. To test whether cytokinin controls plant development through inhibiting an early step in the auxin response pathway, we compared the effects of cytokinin with those of the dgt (diageotropica) mutation, which is known to block rapid auxin reactions of tomato (Lycopersicon esculentum) hypocotyls. Long-term cytokinin treatment of wild-type seedlings phenocopied morphological traits of dgt plants such as stunting of root and shoot growth, reduced elongation of internodes, reduced apical dominance, and reduced leaf size and complexity. Cytokinin treatment also inhibited rapid auxin responses in hypocotyl segments: auxin-stimulated elongation, H(+) secretion, and ethylene synthesis were all inhibited by cytokinin in wild-type hypocotyl segments, and thus mimicked the impaired auxin responsiveness found in dgt hypocotyls. However, cytokinin failed to inhibit auxin-induced LeSAUR gene expression, an auxin response that is affected by the dgt mutation. In addition, cytokinin treatment inhibited the auxin induction of only one of two 1-aminocyclopropane-1-carboxylic acid synthase genes that exhibited impaired auxin inducibility in dgt hypocotyls. Thus, cytokinin inhibited a subset of the auxin responses impaired in dgt hypocotyls, suggesting that cytokinin blocks at least one branch of the DGT-dependent auxin response pathway.     

Effect of epibrassinolide on hypocotyl growth of the tomato mutant diageotropica

The effect of epibrassinolide (epiBR) on the growth and gravitropism of hypocotyls was investigated in diageotropica (dgt), a mutant of tomato (Lycopersicon esculentum Mill.). The elongation of (dgt) hypocotyls, which had been reported to be auxin-insensitive, was promoted by increasing concentrations of indole-3-acetic acid (IAA) in the presence of epiBR. α-(p-Chlorophenoxy)isobutyric acid, an inhibitor of auxin action, blocked the enhancement of growth by IAA and epiBR. Time course analysis of IAA-induced dgt hypocotyl elongation in the presence of epiBR revealed typical auxin response kinetics. These results suggest that epiBR restores the auxin responsiveness of dgt hypocotyls with respect to elongation. However, epiBR did not rescue the dgt phenotype with respect to shoot gravitropism. It was therefore concluded that brassinosteroid insensitivity or deficiency is not the primary defect of the dgt mutation. Received: 23 January 1998 / Accepted: 20 June 1998     

The auxin-resistant diageotropica mutant of tomato responds to gravity via an auxin-mediated pathway

 Hypocotyls of the diageotropica (dgt) mutant of tomato (Lycopersicon esculentum Mill.) do not elongate in response to exogenous auxin, but can respond to gravity. This appears paradoxical in light of the Cholodny-Went hypothesis, which states that shoot gravicurvature results from asymmetric stimulation of elongation by auxin. While light-grown dgt seedlings can achieve correct gravitropic reorientation, the response is slow compared to wild-type seedlings. The sensitivity of dgt seedlings to inhibition of gravicurvature by immersion in auxin or auxin-transport inhibitors is similar to that of wild-type plants, indicating that both an auxin gradient and auxin transport are required for the gravitropic response and that auxin uptake, efflux, and at least one auxin receptor are functional in dgt. Furthermore, dgt gravicurvature is the result of asymmetrically increased elongation as would be expected for an auxin-mediated response. Our results suggest differences between elongation in response to exogenous auxin (absent in dgt) and elongation in response to gravistimulation (present but attenuated in dgt) and confirm the presence of two phases during the gravitropic response, both of which are dependent on functional auxin transport. Received: 16 July 1999 / Accepted: 24 September 1999     

ZEA3: A Negative Modulator of Cytokinin Responses in Plant Seedlings

In Nicotiana plumbaginifolia cytokinins affect seedling development by inhibiting root growth and hypocotyl elongation and by stimulating cotyledon expansion. The zea3.1 mutant was selected for its inability to grow in conditions of low nitrogen and for its ability to grow independently on inhibitory concentrations of zeatin (J.D. Faure, M. Jullien, M. Caboche [1994] Plant J 5: 481-491). The zea3.1 growth response to cytokinins is reflected by an increase in cotyledon expansion due to cell division and by a swelling of the hypocotyl due to cell enlargement. An analysis of the seedling's root length and fresh weight over a wide range of benzyladenine concentrations showed that zea3.1 plants exhibit a higher sensitivity and an amplified response to cytokinins. A similar response of zea3.1 to benzyladenine was also seen in the expression of msr1, a cytokinin-regulated gene. Regulation of msr1 expression by protein phosphorylation was unaffected by the zea3.1 mutation. No significant differences in cytokinin and auxin levels were found between zea3.1 and wild-type seedlings, suggesting that the mutant phenotype is not caused by an alteration of these hormone levels. The data presented suggest that ZEA3 negatively modulates cytokinin responses and may function as a broad regulator of seedling development.     

Gene expression profile of zeitlupe/lov kelch protein1 T-DNA insertion mutants in Arabidopsis thaliana: Downregulation of auxin-inducible genes in hypocotyls

Elongation of hypocotyl cells has been studied as a model for elucidating the contribution of cellular expansion to plant organ growth. ZEITLUPE (ZTL) or LOV KELCH PROTEIN1 (LKP1) is a positive regulator of warmth-induced hypocotyl elongation under white light in Arabidopsis, although the molecular mechanisms by which it promotes hypocotyl cell elongation remain unknown. Microarray analysis showed that 134 genes were upregulated and 204 genes including 15 auxin-inducible genes were downregulated in the seedlings of 2 ztl T-DNA insertion mutants grown under warm conditions with continuous white light. Application of a polar auxin transport inhibitor, an auxin antagonist or an auxin biosynthesis inhibitor inhibited hypocotyl elongation of control seedlings to the level observed with the ztl mutant. Our data suggest the involvement of auxin and auxin-inducible genes in ZTL-mediated hypocotyl elongation.     

Transgene-mediated auxin overproduction in Arabidopsis: hypocotyl elongation phenotype and interactions with the hy6-1 hypocotyl elongation and axr1 auxin-resistant mutants

Transgenic Arabidopsis thaliana plants constitutively expressing Agrobacterium tumefaciens tryptophan monooxygenase (iaaM) were obtained and characterized. Arabidopsis plants expressing iaaM have up to 4-fold higher levels of free indole-3-acetic acid (IAA) and display increased hypocotyl elongation in the light. This result clearly demonstrates that excess endogenous auxin can promote cell elongation in a whole plant. Interactions of the auxin-overproducing transgenic plants with the phytochrome-deficient hy6-1 and auxin-resistant axrl-3 mutations were also studied. The effects of auxin overproduction on hypocotyl elongation were not additive to the effects of phytochrome deficiency in the hy6-1 mutant, indicating that excess auxin does not counteract factors that limit hypocotyl elongation in hy6-1 seedlings. Auxin-overproducing seedlings are also qualitatively indistinguishable from wild-type controls in their response to red, far-red, and blue light treatments, demonstrating that the effect of excess auxin on hypocotyl elongation is independent of red and blue light-mediated effects. All phenotypic effects of iaaM-mediated auxin overproduction (i.e. increased hypocotyl elongation in the light, severe rosette leaf epinasty, and increased apical dominance) are suppressed by the auxin-resistant axr1-3 mutation. The axr1-3 mutation apparently blocks auxin signal transduction since it does not reduce auxin levels when combined with the auxin-overproducing transgene.     

Callus,shoot and hairy root formation in vitro as affected by the sensitivity to auxin and ethylene in tomato mutants

We analyzed the impact of ethylene and auxin disturbances on callus, shoots and Agrobacterium rhizogenes-induced hairy root formation in tomato (Solanum lycopersicum L.). The auxin low-sensitivity dgt mutation showed little hairy root initiation, whereas the ethylene low-sensitivity Nr mutation did not differ from the control Micro-Tom cultivar. Micro-Tom and dgt hairy roots containing auxin sensitivity/biosynthesis rol and aux genes formed prominent callus onto media supplemented with cytokinin. Under the same conditions, Nr hairy roots did not form callus. Double mutants combining Rg1, a mutation conferring elevated shoot formation capacity, with either dgt or Nr produced explants that formed shoots with little callus proliferation. The presence of rol + aux genes in Rg1 hairy roots prevented shoot formation. Taken together, the results suggest that although ethylene does not affect hairy root induction, as auxin does, it may be necessary for auxin-induced callus formation in tomato. Moreover, excess auxin prevents shoot formation in Rg1.     

Similarity between Cytokinin and Blue Light Inhibition of Cucumber Hypocotyl Elongation

The cytokinin benzyladenine inhibited endogenous hypocotyl elongation in intact etiolated seedlings of cucumber (Cucumis sativus L.). In hypocotyl segments, the inhibitory effect of benzyladenine on growth was clearly detectable in the presence of indoleacetic acid. Fusicoccin-induced elongation was unaffected by the presence of cytokinin. The effect of cytokinin on elongation of the segments was determined by measuring changes in fresh weight, a linear function of extension growth. The effect of benzyladenine on hypocotyl growth was at least as large in segments prepared from red-light-grown seedlings as in those from seedlings grown in total darkness. A comparison was made between the inhibitory effects of cytokinin and blue light. The use of the calcium chelator ethyleneglycol-bis(β-aminoethyl ether)-N, N′-tetraacetic acid indicated that calcium ions are required for manifestation of benzyladenine-induced inhibition.     

The Effects of Cytokinin and Light on Hypocotyl Elongation in Arabidopsis Seedlings Are Independent and Additive

Cytokinin has been reported to mimic some of the effects of light on de-etiolation responses in dark-grown Arabidopsis seedlings. The interaction between cytokinin and light was examined by analyzing cytokinin dose and light fluence effects on hypocotyl elongation in wild-type and mutant Arabidopsis seedlings with defects in light or hormone responses. It was found that (a) cytokinin and light-response systems have independent and additive effects on the inhibition of hypocotyl elongation and (b) either cytokinin or light can saturate the morphogenic responses. As a consequence, cytokinin has no effect on hypocotyl elongation under normal growth conditions because light levels saturate the hypocotyl inhibition response. To determine whether a functional light-response pathway is required for cytokinin responses, light-insensitive long hypocotyl (hy) mutants were tested for cytokinin responses. The hy mutants (hy1 to hy6) had normal cytokinin responses, except phyB-1 (hy3-1), in which hypocotyl elongation was insensitive to cytokinin. Cytokinin insensitivity in phyB-1 was attributed to an indirect effect of the mutation on cytokinin responses. The effects of cytokinin on the inhibition of hypocotyl elongation are largely mediated by ethylene, and blocking the ethylene-response pathway through the action of a cytokinin-resistant, ethylene-insensitive mutant (ckr1/ein2) had no effect on the light inhibition of hypocotyl elongation. These results do not support the idea that cytokinin mediates the action of light on hypocotyl elongation.     

Cytokinin-induced hypocotyl elongation in light-grown<Emphasis Type="Italic"> Arabidopsis</Emphasis> plants with inhibited ethylene action or indole-3-acetic acid transport

Cytokinins inhibit hypocotyl elongation in darkness but have no obvious effect on hypocotyl length in the light. However, we found that cytokinins do promote hypocotyl elongation in the light when ethylene action is blocked. A 50% increase in Arabidopsis thaliana (L.) Heynh. hypocotyl length was observed in response to N⁶-benzyladenine (BA) treatment in the presence of Ag⁺. The level of the ethylene precursor 1-aminocyclopropane-1-carboxylic acid was strongly increased, indicating that ethylene biosynthesis was up-regulated by treatment with cytokinin. Furthermore, the effects of cytokinins on hypocotyl elongation were also tested using a series of mutants in the cascade of the ethylene-signal pathway. In the ethylene-insensitive mutants etr1-3 and ein2-1, cytokinin treatment resulted in hypocotyl lengths comparable to those of wild-type seedlings treated with both Ag⁺ and BA. A similar phenotypical response to cytokinin was observed when auxin transport was blocked by -naphthylphthalamic acid (NPA). Applied cytokinin largely restored cell elongation in the basal and middle parts of the hypocotyls of NPA-treated seedlings and at the same time abolished the NPA-induced decrease in indole-3-acetic acid levels. Our data support the hypothesis that, in the light, cytokinins interact with the ethylene-signalling pathway and conditionally up-regulate ethylene and auxin synthesis.     

RcRR1, a Rosa canina Type-A Response Regulator Gene,Is Involved in Cytokinin-Modulated Rhizoid Organogenesis

In vitro, a new protocol of plant regeneration in rose was achieved via protocorm-like bodies (PLBs) induced from the root-like organs named rhizoids that developed from leaf explants. The development of rhizoids is a critical stage for efficient regeneration, which is triggered by exogenous auxin. However, the role of cytokinin in the control of organogenesis in rose is as yet uncharacterized. The aim of this study was to elucidate the molecular mechanism of cytokinin-modulated rhizoid formation in Rosa canina. Here, we found that cytokinin is a key regulator in the formation of rhizoids. Treatment with cytokinin reduced callus activity and significantly inhibited rhizoid formation in Rosa canina. We further isolated the full-length cDNA of a type-A response regulator gene of cytokinin signaling, RcRR1, from which the deduced amino acid sequence contained the conserved DDK motif. Gene expression analysis revealed that RcRR1 was differentially expressed during rhizoid formation and its expression level was rapidly up-regulated by cytokinin. In addition, the functionality of RcRR1 was tested in Arabidopsis. RcRR1 was found to be localized to the nucleus in GFP-RcRR1 transgenic plants and overexpression of RcRR1 resulted in increased primary root length and lateral root density. More importantly, RcRR1 overexpression transgenic plants also showed reduced sensitivity to cytokinin during root growth; auxin distribution and the expression of auxin efflux carriers PIN genes were altered in RcRR1 overexpression plants. Taken together, these results demonstrate that RcRR1 is a functional type-A response regulator which is involved in cytokinin-regulated rhizoid formation in Rosa canina.     

The Inhibition by Cytokinin of Auxin-Promoted Elongation in Excised Soybean Hypocotyl

The experiments characterize the inhibition by kinetin of auxin-promoted elongation in excised hypocotyl sections of 3-day soybean seedlings (Glycine max cv. Hawkeye 63). It was found that concentrations of kinetin above 4.2 μM did not further inhibit auxin-promoted elongation. Kinetin is as potent an inhibitor of elongation as actinomycin D or cycloheximide. Tissue incubated for 3 or 5 h in the absence of auxin or cytokinin would, upon addition of auxin, exhibit a new growth rate similar to that of tissue grown in auxin for the entire incubation period. Similarly, tissue grown for 3 and 5 h in the presence of auxin would revert to the control rate of elongation upon addition of kinetin. A 10 to 30 min preincubation in kinetin yielded the tissue incapable, for the ensuing 6 h, of increasing its rate of elongation in response to auxin. Zeatin and isopentenyladenine were more potent than kinetin and benzyladenine in the inhibition of elongation. Levels of ethylene produced in the presence of auxin plus cytokinin indicated that it was not involved in this auxin-cytokinin interaction. Kinetin by itself did not promote elongation; nor did it enhance auxin-promoted elongation at low auxin concentrations.     

The diageotropica gene of tomato encodes a cyclophilin: a novel player in auxin signaling

The single gene, auxin-resistant diageotropica (dgt) mutant of tomato displays a pleiotropic auxin-related phenotype that includes a slow gravitropic response, lack of lateral roots, reduced apical dominance, altered vascular development, and reduced fruit growth. Some auxin responses are unaltered in dgt plants, however, and the levels, metabolism, and transport of auxin appear normal, indicating that the Dgt gene encodes a component of a specific auxin signaling pathway. By combining map-based cloning with comparative microsynteny, we determined that the Dgt gene encodes a cyclophilin (CYP) (LeCYP1; gi:170439) that has not previously been identified as a component of auxin signaling and plant development. Each of the three known dgt alleles contains a unique mutation in the coding sequence of LeCyp1. Alleles dgt ^1-1 and dgt ^1-2 contain single nucleotide point mutations that generate an amino acid change (G₁₃₇R) and a stop codon (W₁₂₈stop), respectively, while dgt ^dp has an amino acid change (W₁₂₈CΔ129–133) preceding a 15 bp deletion. Complementation of dgt plants with the wild-type LeCyp1 gene restored the wild-type phenotype. Each dgt mutation reduced or nullified the peptidyl–prolyl isomerase activity of the GST–LeCYP1 fusion proteins in vitro. RT-PCR and immunoblot analyses indicated that the dgt mutations do not affect the expression of LeCyp1 mRNA, but the accumulation of LeCYP1 protein is greatly reduced for all three mutant alleles. The CYP inhibitor, cyclosporin A, partially mimics the effects of the dgt mutation in inhibiting auxin-induced adventitious root initiation in tomato hypocotyl sections and reducing the auxin-induced expression of the early auxin response genes, LeIAA10 and 11. These observations confirm that the PPIase activity of the tomato CYP, LeCYP1, encoded by the Dgt gene is important for specific aspects of auxin regulation of plant growth, development, and environmental responses.     

Auxin increases the hydrogen peroxide (H2O2) concentration in tomato (Solanum lycopersicum) root tips while inhibiting root growth

Background and Aims

The hormone auxin and reactive oxygen species (ROS) regulate root elongation, but the interactions between the two pathways are not well understood. The aim of this study was to investigate how auxin interacts with ROS in regulating root elongation in tomato, Solanum lycopersicum.

Methods

Wild-type and auxin-resistant mutant, diageotropica (dgt), of tomato (S. lycopersicum ‘Ailsa Craig’) were characterized in terms of root apical meristem and elongation zone histology, expression of the cell-cycle marker gene Sl-CycB1;1, accumulation of ROS, response to auxin and hydrogen peroxide (H₂O₂), and expression of ROS-related mRNAs.

Key Results

The dgt mutant exhibited histological defects in the root apical meristem and elongation zone and displayed a constitutively increased level of hydrogen peroxide (H₂O₂) in the root tip, part of which was detected in the apoplast. Treatments of wild-type with auxin increased the H₂O₂ concentration in the root tip in a dose-dependent manner. Auxin and H₂O₂ elicited similar inhibition of cell elongation while bringing forth differential responses in terms of meristem length and number of cells in the elongation zone. Auxin treatments affected the expression of mRNAs of ROS-scavenging enzymes and less significantly mRNAs related to antioxidant level. The dgt mutation resulted in resistance to both auxin and H₂O₂ and affected profoundly the expression of mRNAs related to antioxidant level.

Conclusions

The results indicate that auxin regulates the level of H₂O₂ in the root tip, so increasing the auxin level triggers accumulation of H₂O₂ leading to inhibition of root cell elongation and root growth. The dgt mutation affects this pathway by reducing the auxin responsiveness of tissues and by disrupting the H₂O₂ homeostasis in the root tip.     

A role for ABCB19‐mediated polar auxin transport in seedling photomorphogenesis mediated by cryptochrome 1 and phytochrome B

During seedling establishment, blue and red light suppress hypocotyl growth through the cryptochrome 1 (cry1) and phytochrome B (phyB) photosensory pathways, respectively. How these photosensory pathways integrate with growth control mechanisms to achieve the appropriate degree of stem elongation was investigated by combining cry1 and phyB photoreceptor mutations with genetic manipulations of a multidrug resistance‐like membrane protein known as ABCB19 that influenced auxin distribution within the plant, as evidenced by a combination of reporter gene assays and direct auxin measurements. Auxin signaling and ABCB19 protein levels, hypocotyl growth rates, and apical hook opening were measured in mutant and wild‐type seedlings exposed to a range of red and blue light conditions. Ectopic/overexpression of ABCB19 (B19OE) greatly increased auxin in the hypocotyl, which reduced the sensitivity of hypocotyl growth specifically to blue light in long‐term assays and red light in high‐resolution, short‐term assays. Loss of ABCB19 partially suppressed the cry1 hypocotyl growth phenotype in blue light. Hypocotyl growth of B19OE seedlings in red light was very similar to phyB mutants. Altered auxin distribution in B19OE seedlings also affected the opening of the apical hook. The cry1 and phyB photoreceptor mutations both increased ABCB19 protein levels at the plasma membrane, as measured by confocal microscopy. The B19OE plant proved to be a useful tool for determining aspects of the mechanism by which light, acting through cry1 or phyB, influences the auxin transport process to control hypocotyl growth during de‐etiolation.     

Arabidopsis cytokinin-resistant mutant, cnr1, displays altered auxin responses and sugar sensitivity

Based upon the phenotype of young, dark-grown seedlings, a cytokinin-resistant mutant, cnr1, has been isolated, which displays altered cytokinin- and auxin-induced responses. The mutant seedlings possess short hypocotyls and open apical hooks (in dark), and display agravitropism, hyponastic cotyledons, reduced shoot growth, compact rosettes and short roots with increased adventitious branching and reduced number of root hairs. A number of these features invariably depend upon auxin/cytokinin ratio but the cnr1 mutant retains normal sensitivity towards auxin as well as auxin polar transport inhibitor, TIBA, although upregulation of primary auxin-responsive Aux/IAA genes is reduced. The mutant shows resistance towards cytokinin in hypocotyl/root growth inhibition assays, displays reduced regeneration in tissue cultures (cytokinin response) and decreased sensitivity to cytokinin for anthocyanin accumulation. It is thus conceivable that due to reduced sensitivity to cytokinin, the cnr1 mutant also shows altered auxin response. Surprisingly, the mutant retains normal sensitivity to cytokinin for induction of primary response genes, the type-A Arabidopsis response regulators, although the basal level of their expression was considerably reduced as compared to the wild-type. The zeatin and zeatin riboside levels, as estimated by HPLC, and the cytokinin oxidase activity were comparable in the cnr1 mutant and the wild-type. The hypersensitivity to red light (in hypocotyl growth inhibition assay), partial photomorphogenesis in dark, and hypersensitivity to sugars, are some other features displayed by the cnr1 mutant. The lesion in the cnr1 mutant has been mapped to the top of chromosome 1 where no other previously known cytokinin-resistant mutant has been mapped, indicating that the cnr1 mutant defines a novel locus involved in hormone, light and sugar signalling.     

Effects of Brassin-Complex on Auxin and Gibberellin Mediated Events in the Morphogenesis of the Etiolated Bean Hypocotyl

The excised, hooked bean hypocotyl was the system used to determine wheiher the ‘auxin- and gibberellin like’ effect of the lipoidal pollen extract, Brass in-complex (Br), were mediated through, or independent of, auxin and gibberellin. The morphogenetic events of hook opening and hypocotyl elongation in this system are regulated by auxin and gibberellin, respectively. Brassin complex, like IAA, elicited a book closure in (he dark and retarded its opening in red light. This effect was synergized by T1BA, IAA and the presence of the auxin-producing organs, the epicotyl and cotyledons. Br-elicited hook closure was inhibited by the antiauxin. PCIB. Both GA₃ and Br totally reversed the light inhibition of hypocotyl elongation. The GA₃-effect, but nol the Br elicited elongation, was overcome by Ancymidol. Hypocotyl elongation was partially inhibited by TIBA and PCIB. suggesting a possible auxin involvement also in this effect of Br. Br may elicit its growth responses through an effect on endogenous auxin levels, In this way it is different from other lipoidat growth regulators, such as the oleanimins which require the presence of exogenous growth regulators for activity.     

Interaction of ethylene and a cytokinin in promoting hypocotyl elongation in a dwarf strain of watermelon

Experiments were conducted to study the interaction of ethylene and the cytokinin N⁶-benzyladenine (BA) in promoting hypocotyl elongation in a dwarf strain of watermelon (Citrullus lanatus [Thunb] Matsu. and Nakai). Optimum promotion of hypocotyl elongation is elicited by an apical treatment with 0.2 microgram BA. At dosages above 0.3 microgram per apex, BA-enhancement of elongation is reduced concomitant with stimulation of ethylene production and lateral expansion of hypocotyls. Application of the ethylene biogenesis inhibitor, aminoethoxyvinylglycine, at dosages from 0.3 to 10 micrograms per apex inhibited BA-induced ethylene production. In seedlings treated with 0.2 microgram BA, 10 micrograms aminoethoxyvinylglycine per apex reduced ethylene production to about one-third of control levels and reduced BA stimulation of hypocotyl elongation by 74%. Exposure of watermelon seedlings to 60 ± 10 nanoliters per liter of ethylene in a flowing system nearly eliminated aminoethoxyvinylglycine inhibition of BA-promoted growth. The results suggest that physiological levels of internal ethylene are required for cytokinin promotion of hypocotyl elongation in watermelon.