Internode length in Pisum. III The effect and interaction of the Na/na and Le/le gene differences on endogenous gibberellin-like substances

In the garden pea ( Pisum sativum L.), shoots of the extremely short plants with the mutant na (phenotype nana) are found by bioassay to contain undetectable levels of gibberellin-like substances. This is confirmed by the use of near isogenic lines differing at the Na locus. Thus, mutant na appears to block a step early in the pathway of gibberellin synthesis. It is suggested that the polar gibberellin-like substance found in the apical portion of shoots of tall ( Le ) but not dwarf ( le ) peas could be GA₁. Extracts of shoots of na Le peas treated with GA₂₀ (the major active gibberellin in dwarf peas) possess a large amount of GA₁-like activity whereas extracts of shoots of na le peas treated with GA₂₀ possess a much reduced amount. Thus, gene Le may allow the conversion of a less active gibberellin (GA₂₀) into one more active in stimulating elongation in the pea (the GA₁-like compound). In contrast to their influence in the shoot, the na and Le genes do not appear to be operative in controlling the gibberellin content of developing seed, indicating that organ specific gibberellin biosynthesis and metabolism occur in peas.     

Internode length in Pisum. Action of gene lw

Jolly, C. J., Reid, J. B. and Ross, J. J. 1987. Internode length in Pisum. Action of gene lw.
Mutant K29 of Pisum sativum L. is shown to possess a recessive gene at a new locus, lw , which results in reduced internode length, delayed flowering and increased symptoms of water congestion compared with the parental cv. Torsdag. The interaction of gene lw with the internode length genes na, le, la and cry ⁵ is examined. Extracts from the shoots of Iw plants are shown to contain similar levels of gibberellin (GA)-like substances to comparable Lw plants, but Iw plants do not elongate to the same extent as Lw plants when treated with GA₁₉ GA₁₉, or GA₂₀. The effect of gene Iw is not graft-transmissible. Unlike essentially isogenic dwarf lines possessing the GA-synthesis genes le, Ih or Is, lw plants show a relative increase in elongation similar to Torsdag in response to photoperiod extensions from sources rich in far-red light. These results suggest that gene lw probably does not reduce elongation by influencing GA-synthesis and that the response to photoperiod extensions with far-red light may depend on the level of GA.     

Internode length in Pisum. IV. The effect of the Le gene on gibberellin metabolism

The highly active, polar gibberellin-like substance found in the apical region of shoots of tall (genotype Le ) peas ( Pisum sativum L.) is shown by combined gas chromatography-mass spectrometry (GC/MS) to be GA₁. This substance is either absent or present at only low levels in dwarf ( le ) plants. Multiple ion monitoring (MIM) tentatively suggests that GA₈ may also be present in shoot tissue of tall peas. Gibberellin A₁ is the first 3 β-hydroxylated gibberellin positively identified in peas, and its presence in shoot tissue demonstrates the organ specificity of gibberellin production since GA₁ has not been detected in developing seeds. Application of GA₁ can mask the Le/le gene difference. However, whilst Le plants respond equally to GA₂₀ and GA₁, le plants respond only weakly to GA₂₀, the major biologically active gibberellin found in dwarf peas. These results suggest that the Le gene controls the production of a 3 β-hydroxylase capable of converting GA₂₀ to GA₁. Further support for this view comes from feeds of [³H] GA₂₀ to Le and le plants. Plants with Le metabolise [³H] GA₂₀ to three major products whilst le plants produce only one major product after the same time. The metabolite common to Le and le plants co-chromatographs with GA₂₉. The additional two metabolites in Le peas co-chromatograph with GA₁ and GA₈.     

Internode length in Lathyrus odoratus. Effects of mutants l and lb on gibberellin metabolism and levels

After the application of [¹³C³H]-gibberellin A₂₀ to wild-type (tall) sweet peas ( Lathyrus odoratus L.) labelled gibberellin A₁ (GA₁), GA₈, GA₂₉ and 2-epiGA₂₉ were identified as major metabolities by gas chromatography-mass spectrometry after high performance liquid chromatography. By contrast in genetically comparable dwarf ( II ) plants only labelled GA₂₉ and 2-epiGA₂₉ were produced in significant amounts, although evidence was obtained for trace amounts of labelled GA₁ and GA₈. The apical portions of dwarf plants contained 8–10 times less GA₁ than those of tall plants but at least as much GA₂₀ (measured using di-deuterated internal standards). In conjunction with previous data these results strongly indicate that in genotype ll internode length is reduced and leaf growth altered by a reduction in GA1 levels attributable to a partial block in the 3β-hydroxylation of GA₂₀ to GA₁.
In contrast to dwarf plants, semidwarf plants (genotype lblb ) contained more GA₁ in the apical portion than wild-type counterparts. This is consistent with the suggestion that lb alters some aspect of GA sensitivity.     

Growth and gibberellin relations of the extreme dwarf dx tomato mutant

The extreme dwarf d ^x tomato ( Lycopersicon esculentum Mill.) mutant has very short internodes which were found to contain shorter and fewer epidermal cells. The leaves are highly abnormal. The mutant showed a substantial stem growth response to GA₃, without approaching normal stature or morphology. The active gibberellin GA₁ and its precursors GA₁₉ and GA₂₀ were identified by coupled gas chromatography-mass spectrometry (GC/MS) in d ^x shoots. Quantitative GC/MS revealed that GA₂₀ accumulated to far higher levels than normal in stems and leaves of the mutant.     

Internode length in Pisum. Comparison of genotypes in the light and dark

There is a strong relationship across the full range of gibberellin deficient, internode length genotypes ( le, lh, is, na ) between internode length in the dark and in red or white light. Further, the new, more severe allele at the le locus. Ie ^d, is shown to influence growth in the dark as well as in the light. These results suggest that darkeness does not specifically overcome any of the steps blocked by the gibberellin (GA) synthesis genes contrasting with the conclusions drawn by other workers. Supporting this conclusion in relation to the Ie gene are results which show that, at least at certain dosage rates, dark-grown Ie na plants respond better to GA₁ than to GA₂₀ similar to the response previously reported in light grown plants.
The greater response by plants of the nana line NGB1766 ( na ) to GA₁ in the dark than in the light suggests that light may influence internode length by altering GA-sensitivity. These results are discussed in relation to previous views on the control of stem elongation by light.     

Internode length in Pisum. A new allele at the Lh locus

A new allele at the Lh locus has been identified in Pisum sativum L. and named lhⁱ . This allele results in reduced GA levels in young shoots, and a dwarf phenotype. Gas chromatography-selected ion monitoring (GC-SIM) with dideuterated internal standards has been used to demonstrate a quantitative relationship between the level of endogenous GA₁ and internode length using the three alleles ( Lh, lh and lhⁱ ) at the Lh locus. These results are consistent with previous findings in peas (for alleles at the Le locus) and other species possessing a predominant early 13-hydroxylation pathway for GA biosynthesis and support the role of GA₁ as the major native GA in peas with biological activity in its own right. However, in contrast to alleles at the Le locus, GA₂₀ levels are also reduced in lh and lhⁱ plants. The lhⁱ allele also has possible pleiotropic effects on seed abortion, leading to a reduction in seed yield compared to plants homozygous for the previously characterised Lh or lh alleles.     

Internode length in Lathyrus odoratus. The involvement of gibberellins

Evidence was obtained by gas chromatography-mass spectrometry and gas chromatography-selected ion monitoring for the presence of gibberellin A₂₀), GA₁, GA₂₉, GA₈ and 2-epiGA₂₉ in vegetative shoots of tall sweet pea, Lathyrus odoratus L. Both tall (genotype L –) and dwarf (genotype II ) sweet peas elongated markedly in response to exogenous GA₁ attaining similar internode lengths at the highest dose levels. Likewise internode length in both genotypes was reduced by application of the GA biosynthesis inhibitor, PP333. The ratio of leaflet length to width was reduced by application of PP333 to tall plants and this effect was reversed by GA₁. When applied to plants previously treated with PP333, GA₂₀ promoted internode elongation of L – plants as effectively as GA₁, but GA₂₉ was not as effective as GA₁ when applied to II plants. In contrast, GA₂₀ and GA₁ were equally effective when applied to the semidwarf lb mutant but GA-treated lblb plants did not attain the same internode length as comparable GA-treated Lb – plants. The difference in stature between the tall and dwarf types persisted in dark-grown plants. It is concluded that GA₁ may be important for internode elongation and leaf growth in sweet pea. Mutant l may influence GA₁ synthesis by reducing 3β-hydroxylation of GA₂₀ whereas mutant lb appears to affect GA sensitivity.     

Internode length in Pisum. Two further mutants, lh and ls, with reduced gibberellin synthesis, and a gibberellin insensitive mutant, lk

Three further internode length mutants in peas ( Pisum sativum L.), lh , ls and lk , were examined to determine if they influenced gibberellin synthesis or sensitivity. Two mutants, lh and ls , showed pronounced elongation in response to applied GA₁ and extracts from their shoots contained little gibberellin-like activity when assayed on the rice seedling (cv. Tan ginbozu) bioassay compared with similar extracts from essentially isogenic Lh and Ls plants. The third mutant, lk , was almost insensitive to applied GA₁ and at no dose rate did it become a phenocopy of normal Lk plants. Extracts from the shoots of lk and Lk segregants contained similar levels of gibberellinlike substances. All three mutants influenced growth in both the light and the dark, although only the effect of genes Lh and Ls were graft transmissible. These results suggest that lh and ls are mutants with reduced gibberellin synthesis, while lk is the first gibberellin-insensitive dwarfing gene identified in peas.     

Response of genetic dwarf apple plants to GA3

High temperature has been implicated as the major factor responsible for dwarfing of selected apple ( Malus domestica Borkh.) trees of a hybrid population of cv. Goldspur Delicious x cv. Redspur Delicious. Dwarf plants grew only 2.2 cm in 63 days under a ramped temperature regime (night 15°C, day ramped up to 38°C, held for 2 h and ramped down to 15°C—14 h daylength), whereas semi-dwarf plants grew 26.3 cm. At a constant 27°C (14 h daylength), both dwarf and semi-dwarf plants grew 26.3 cm. At a constant 27°C (14 h daylength), both dwarf and semi-dwarf plants grew nearly 50 cm. The gibberellin biosynthesis inhibitor, paclobutrazol, retarded growth of semi-dwarf plants in both ramped and constant environments and dwarf plants in the constant 27°C environment. It did not further reduce the size of dwarf plants growing under the ramped regime. Gibberellin (GA₃) treatment reversed the inhibition of growth caused by paclobutrazol for all plants except it did not restore growth of dwarf plants in the ramped environment. These data suggest that neither pacobutrazoltreated nor untreated dwarf plants growing in the ramped environment (or in the orchard during hot summer months) are able to respond to GA₃. In constrast, GA₃ was utilized by the paclobutrazol-inhibited dwarf plants growing at constant 27°C, enabling shoot elongation to take place. It appears that high temperature may have caused alterations in GA target tissues in dwarf plants so that they no longer had the capacity to respond to GA.     

Internode length in Pisum. Estimation of GA1 levels in genotypes Le, le and led

The levels of GA₁, 3-epiGA₁ and GA₈ in genotypes Le, le and le^d of Pisum sativum L. were determined by gas chromatography-selected ion monitoring (GC-SIM) after feeds of [³H, ¹³C]-GA₂₀ to each genotype. The levels of endogenous and [¹³C]-labelled metabolites were determined by reverse isotope dilution with unlabelled GA₁, 3-epiGA₁ and GA₈. The results demonstrate a quantitative relationship between the level of GA₁ and the extent of elongation both on a per plant and a per g fresh weight basis. These results are consistent with previous findings in peas and other species possessing a predominant early 13-hydroxylation pathway for GA biosynthesis.
The levels of 3-epiGA₁ also decreased in the genotypic sequence Le, le, le^d although not as rapidly as for the level of GA₁. This may suggest that the alleles at the le locus also influence the formation of 3-epiGA₁.     

Geibberllin and temperature influence carbohydrate content and flowering in Phalaenopsis

When Phalaenopsis amabilis is grown under high temperature (30/25°C, day/night), flowering is blocked, and this can be reversed by gibberellin A₃ (GA₃) treatment. Associated with GA₃ treatment under high temperature are increases in sucrose, glucose and fructose as compared with warm-treated plants. Spraying with sucrose solution alone caused leaf epinasty in plants grown under high temperature. Epinasty was released by about 9 days of GA₃ treatment. In GA₃-treated plants under high temperatures, sucrose application to the source leaves led to an increase in sugar content in both leaves and inflorescence. In contrast, although in warm-treated plants sucrose application to the source leaves increased sugar content in the leaves, it did not increase sucrose content in the inflorescence. These results corroborate our hypothesis that in Phalaenopsis GA₃ stimulates sink activity in the apical meristem and promotes the translocation of sucrose from source leaves to the apex of the inflorescence, where it accumulates. GA₃ treatment led to an increase in sucrose synthase activity and had no effect on invertase activity.     

Internode length in Pisum. Two further gibberellin-insensitivity genes, lka and lkb

Two further internode length genes are identified in Pisum sativum L. and named lka (identified from line NGB5865) and lkb (from NGB5862). These genes result in a similar phenotype, which includes reduced stem elongation, peduncle length and basal branching, and 'banding' of the stem. These effects are similar to, but less severe than, those of gene lk . Genes lka and lkb influence gibberellin (GA) sensitivity, since mutants NGB5865 and NGB5862 possess similar levels of endogenous GA-like substances to the wild-type parental cultivar Torsdag and respond less to applied GA₁ than do wild-type plants or GA-synthesis mutants of a similar stature. The action of genes lka and lkb is localised in the young apical tissue but is not thought to involve GA-perception, since plants possessing genes lka and lkb are not true phenocopies of GA-deficient plants. The genetic interaction of genes lka and lkb is examined and the action of gene lkb on a le gene background determined.     

Internode length in Pisum. The lrs mutation reduces gibberellin response and levels

We describe a new mutation, lrs , which reduces internode length in Pisum sativum L. The mutation appears to act by reducing both GA synthesis and the response to GA₁. The levels of the 13‐hydroxylated GAs, GA₅₃, GA₄₄, GA₁₉, GA₂₀, GA₁, and GA₈ in the lrs mutant were greatly reduced compared with the wild‐type. The extent of the reduction in GA₁ content in the apical tissues would, at least in part, account for the dwarf phenotype of the mutant. The reduced GA responsiveness of the new mutant was indicated by the inability of applied GA₁ to remove the difference in elongation between lrs and LRS plants. The lrs mutant appears to be unique amongst internode length genotypes, possessing characteristics of both GA synthesis and GA response mutants.     

Constitutive expression of a fruit phytoene synthase gene in transgenic tomatoes causes dwarfism by redirecting metabolites from the gibberellin pathway

Tomato plants transformed with a copy of the fruit-expressed phytoene synthase cDNA under control of the CaMV 35S promoter showed ectopic production of carotenoids. High expressers were reduced in stature. The dwarf character was inherited with an inverse relationship between expression of phytoene synthase and plant height. Severely affected plants also showed reduced chlorophyll content in young leaves. These dwarfs showed a 30-fold reduction in levels of gibberellin A₁ (GA₁) and growth was partially restored by treatment with exogenous GA₃. Qualitative and quantitative changes in carotenoids were also found. It is proposed that the dwarf phenotype results from the over-production of phytoene synthase, which converts geranylgeranyl diphosphate to phytoene and thereby diverts this intermediate away from the gibberellin (GA) and phytol biosynthetic pathways.     

Role of endogenous gibberellins in germination of melon (Cucumis melo) seeds

The effect of the plant growth retardants ancymidol. mefluidide and uniconazole on germination of two melon accessions differing in their ability to germinate at 14°C was examined. The accessions were the cold sensitive Noy Yizre'el and the cold tolerant Persia 202. The three growth retardants were able to delay the germination of intact Noy Yizre'el seeds, but did not affect that of intact Persia 202 seeds. On the other hand germination of decoated seeds of both accessions was unaffected by these inhibitors at normal oxygen concentration, but was inhibited at 5% oxygen. When gibberellin-like activity was measured by a dwarf rice biological assay following HPLC fractionation, it was found that seeds of Persia 202 contained much more gibberellin-like activity than Noy Yizre'el seeds. Among the extracted compounds several endogenous gibberellins were identified by combined gas chromatography-mass spectrometry (GC-MS). They included GA₄, GA₂₀, GA₁ and GA₃ in Noy Yizre'el and GA₃₄, GA₂₀, GA₁ and GA₈ in Persia 202. It is suggested that the better germination of intact Persia 202 seeds, compared to Noy Yizre'el seeds at low temperature and low oxygen concentration, is due to a higher endogenous level of GA and a better seed coat permeability to oxygen.     

Internode length in Pisum. The involvement of ethylene with the gibberellin-insensitive erectoides phenotype

The gene lk in peas ( Pisum sativum L.) confers the erectoides phenotype. This phenotype possesses much reduced internode and petiole lengths and is practically insensitive to applied GA₁, compared with Lk plants. Application of the ethylene synthesis inhibitor, aminoethoxyvinylglycine (AVG), resulted in increased internode elongation and increased GA-sensitivity in lk plants, but not in the Lk line, L53. The ethylene-releasing compound, ethephon, had the reverse effect when applied to the Lk line, L58, reducing internode length and GA-sensitivity. Ethylene production was higher in lk segregates than in Lk segregates under the conditions used, and the shoot anatomy of lk segregates was consistent with these higher ethylene levels.
These results suggest that the phenotypic effects of gene lk may be due, at least in part, to increased ethylene production in erectoides plants. However, AVG application to lk plants did not produce true phenocopies of comparable lk plants and ethephon application to Lk plants did not produce the erectoides phenotype. Further work is therefore required to determine whether the effect of the gene lk on ethylene production is the primary action of this gene or merely a secondary consequence.     

Interaction of potassium ions and gibberellin in the control of hypocotyl growth in Amaranthus caudatus

Potassium promotes growth in several plant tissues. Elongation growth of the hypocotyls of Amaranthus caudatus L. ev. Lalsag is mainly controlled by gibberellins, but K⁺ also promotes growth. In the present study the interaction of K⁺ with gibberellin (GA₃) and chlorocholine chloride (CCC) has been investigated. When K⁺ was applied externally in the dark, hypocotyl growth was promoted in the seedlings. External application of GA₃ did not promote growth in the dark. GA₃ was effective in the light and K⁺ was synergistic with GA₃ in promoting elongation. Application of CCC in the dark makes the seedlings sensitive to GA₃. The inhibition of growth by CCC was also reversed by K⁺. The results indicate a possible role of K⁺ in GA₃ induced elongation of hypocotyls.     

Elongation rates and endogenous indoleacetic acid levels in roots of pea mutants differing in internode length

Growth of the primary root of 12 genotypes of peas ( Pisum sativum ) differing in their stem height was recorded for 14 days. The growth rate of roots of wild-type tall, gibberellin (GA)-deficient le dwarf or slender mutants (with la cry^s ) was similar (3 cm day⁻¹); that of severely GA-deficient nana ( na-1 ) plants was 50% of wild-type but elongation ceased after 8 days; moderately severe dwarf GA-deficient lines ls-1 and lh-1 had a 15% reduction in elongation rate but displayed no time-dependent slowing of the growth rate and brassinosteroid-insensitive and -deficient dwarfs lka and lkb showed slightly decreased root elongation. GA (levels reported in Yaxley et al. 2001 ) is not substantially limiting to root growth until it is severely deficient. The terminal 3 cm of roots of tall plants contained about 25 or 35 ng g⁻¹ fresh weight indole-3-acetic acid (IAA), depending on the genetic background, and le-1 dwarfs were similar. Nana ( na-1 ) had less than 50% the level of IAA of tall, all the moderately severe dwarfs had reductions of about 30% and the slender plants had about 40% more IAA than the corresponding wild-type. With the exception of slender plants, IAA level in the root tips correlated with root elongation. Root growth seems to be promoted by IAA within the range of the internal concentrations detected. Nana plants had a reduced amount of IAA and a lower root-growth rate. Whereas external application of IAA always inhibits root growth, even at very low concentrations, root growth is not similarly inhibited by internal IAA as slender plants had the highest IAA level and growth rate similar to wild-type, regardless of the shoot GA content.