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 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.     

Internode length in Pisum. Gene lkd

A new, single gene, recessive internode length mutant in Pisum, lkd, is described. The internodes of lkd plants are ca. 40% shorter than comparable Lkd plants and this difference appears greater in the dark than in the light. The mutant does not appear to be dwarfed due to modified gibberellin (GA) levels, as determined by gas-chromatography-selected ion monitoring (GC-SIM) for GA₁ and GA₂₀. In relative terms, the mutant responds as well as the wild-type to applied GA₁. However due to its initial short stature it does not elongate to the same extent as the wild-type to high doses of GA₁ suggesting that some other factor, unrelated to GA levels or perception is probably limiting growth in this mutant. Author for correspondence     

Internode length in Zea mays L.

[¹³C, ³H]Gibberellin A₂₀ (GA₂₀) has been fed to seedlings of normal (tall) and dwarf-5 and dwarf-1 mutants of maize (Zea mays L.). The metabolites from these feeds were identified by combined gas chromatography-mass spectrometry. [¹³C, ³H]Gibberellin A₂₀ was metabolized to [¹³C, ³H]GA₂₉-catabolite and [¹³C, ³H]GA₁ by the normal, and to [¹³C, ³H]GA₂₉ and [¹³C, ³H]GA₁ by the dwarf-5 mutant. In the dwarf-1 mutant, [¹³C, ³H]GA₂₀ was metabolized to [¹³C, ³H]GA₂₉ and [¹³C, ³H]GA₂₉-catabolite; no evidence was found for the metabolism of [¹³C, ³H]GA₂₀ to [¹³C, ³H]GA₁. [¹³C, ³H]Gibberellin A₈ was not found in any of the feeds. In all feeds no dilution of ¹³C in recovered [¹³C, ³H]GA₂₀ was observed. Also in the dwarf-5 mutant, the [¹³C]label in the metabolites was apparently undiluted by endogenous [¹³C]GAs. However, dilution of the [¹³C]label in metabolites from [¹³C, ³H]GA₂₀ was observed in normal and dwarf-1 seedlings. The results from the feeding studies provide evidence that the dwarf-1 mutation of maize blocks the conversion of GA₂₀ to GA₁.Abbreviations GA_n gibberellin A_n - GC-MS combined gas chromatography-mass spectrometry - HPLC high-performance liquid chromatography - RP reverse phase     

Internode length in Pisum. Gibberellins and the slender phenotype

Pea plants ( Pisum sativum L.) possessing the slender phenotype (conferred by the gene combination la cry^s ) have extremely long, thin internodes and are phenotypically similar to dwarf plants (possessing genes La and/or Cry ) that have been treated with a non-limiting dose of gibberellin (GA₃). In contrast to tall and dwarf plants, slender plants are virtually insensitive to treatment with AMO 1618, PP333 or GA₃ and addition of the "gibberellin-less" mutant gene na does not alter the phenotype of slender plants. Na slender segregates possessed lower levels of gibberellin-like substances than comparable dwarf segregates when extracts from shoots were assayed using the lettuce hypocotyl or rice seedling bioassays. In addition, na slenders possessed little or no gibberellin-like activity even though they possessed a slender phenotype. Thus the gene combination la cry^s causes slender plants to respond as if they are saturated with gibberellins for growth. In addition, the gene combinations la cry^s and le la cry^c (allele cry^c is less extreme in effect than cry^s ) are shown to be almost completely epistatic to the alleles at the na locus. All these results suggest that gibberellin levels are not important in determining the internode length of slender peas (genotype la cry^s ). The possible mechanisms by which this could occur are discussed.     

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.     

Juvenile hormone titres and winged offspring production do not correlate in the pea aphid, Acyrthosiphon pisum

Pea aphids, Acyrthosiphon pisum, reproduce parthenogenetically and are wing-dimorphic such that offspring can develop into winged (alate) or unwinged (apterous) adults. Alate induction is maternal and offspring phenotype is entirely determined by changes in the physiology and environment of the mother. Juvenile hormones (JHs) have been implicated in playing a role in wing differentiation in aphids, however until recently, methods were not available to accurately quantify these insect hormones in small insects such as aphids. Using a novel LC-MS approach we were able to quantify JH III in pea aphids that were either producing a high proportion of winged morphs among their offspring or mainly unwinged offspring. We measured JH III titres by pooling the hemolymph of 12 or fewer individuals (1 μL hemolymph) treated identically. Levels of JH ranged from 30 to 163 pg/μL. While aphids in the two treatments strongly differed in the proportion of winged morphs among their offspring, their JH III titres did not differ significantly. There was also no correlation between JH III titre and the proportion of winged offspring in induced aphids. This supports earlier findings that wing dimorphism in aphids may be regulated by other physiological mechanisms.     

Monolysocardiolipins accumulate in Barth syndrome but do not lead to enhanced apoptosis

Barth syndrome (BTHS) is an X-linked recessive disorder that is biochemically characterized by low cellular levels of the mitochondrial phospholipid cardiolipin (CL). Previously, we discovered that the yeast disruptant of the TAZ ortholog in Saccharomyces cerevisiae not only displays CL deficiency but also accumulates monolysocardiolipins (MLCLs), which are intermediates in CL remodeling. Therefore, we set out to investigate whether MLCL accumulation also occurs in BTHS. Indeed, we observed MLCL accumulation in heart, muscle, lymphocytes, and cultured lymphoblasts of BTHS patients; however, only very low levels of these lysophospholipids were found in platelets and fibroblasts of these patients. Although the fatty acid composition of the MLCLs was different depending on the tissue source, it did parallel the fatty acid composition of the (remaining) CLs. The possible implications of these findings for the two reported CL remodeling mechanisms, transacylation and deacylation/reacylation, are discussed. Because MLCLs have been proposed to be involved in the initiation of apoptosome-mediated cell death by the sequestration of the proapoptotic protein (t)BH3-interacting domain death agonist (Bid) to the mitochondrial membrane, we used control and BTHS lymphoblasts to investigate whether the accumulation of MLCLs results in higher levels of apoptosis. We found no differences in susceptibility to death receptor-mediated apoptosis or in cellular distribution of Bid, cytochrome c, and other parameters, implying that MLCL accumulation does not lead to enhanced apoptosis in cultured BTHS lymphoblasts.     

Internode length in Pisum. Further studies on the 'micro' gene, lm

Reid, J. B. and Ross, J. J. 1988. Internode length in Pisum. Further studies on the 'micro' gene, lm . - Physiol. Plant. 72: 547–554.
In the garden pea, Pisum sativum L., gene lm confers the micro phenotype. The shoots of lm plants may be described as scaled-down versions of comparable Lm plants, with reduced internode length, leaflet size and rate of leaf expansion. However, the first phenotypic effect of gene lm is on root morphology. The gene results in curling and reduced elongation of the roots and, eventually, degeneration of the root cortex. These changes commence prior to any major visible effects in the shoot. The primary action of the lm gene does not appear to be confined to the root system, however, since epicotyl grafts between Lm and lm plants showed no graft-transmissible effects. The effects of gene lm are also apparent in dark-grown plants. Microdwarf plants ( lm le ) respond well to gibberellin A₁ (GA₁), but do not elongate to the same extent as dwarf ( Lm le ) plants. The two genotypes contain the same complement of GA-like substances. It is argued that gene lm is unlikely to be directly involved with GA-metabolism or the reception of the GA signal, but rather reduces the GA response by influencing some aspect of normal cell development, which results in the wide range of pleiotropic effects observed. Consequently, it may be misleading to continue to classify this gene simply as an internode length 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. 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.     

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 Pisum. Biochemical expression of the le gene in darkness

Ross, J. J. and Reid, J. B. 1989. Internode length in Pisum. Biochemical expression of the Le gene in darkness.
The Le gene appears to be biochemically expressed in dark-grown pea ( Pisum sativum L.) plants since the previously reported difference in metabolism of [³H]-GA₃₀ between light-grown Le and Le plants was also observed in darkness. Furthermore, both light- and dark-grown Le plants contained more endogenous GA¹, -like substance than did comparable Le plants. Darkness did not appear to significantly increase the accumulation of GA¹, in either Le or Le plants, although confirmation of GA¹ levels by gas chromatography-selected ion monitoring is still required. The results support previous findings that the overall metabolism of [³H]-GA₂₀, is accelerated by darkness. The evidence presented here supports previous suggestions that darkness acts on internode length by increasing some aspect of GA sensitivity.     

Synthetic lipid (DOPG) vesicles accumulate in the cell plate region but do not fuse

The cell plate is the new cell wall, with bordering plasma membrane, that is formed between two daughter cells in plants, and it is formed by fusion of vesicles (approximately 60 nm). To start to determine physical properties of cell plate forming vesicles for their transport through the phragmoplast, and fusion with each other, we microinjected fluorescent synthetic lipid vesicles that were made of 1,2-dioleoyl-sn-glycero-3-[phospho-rac-(1-glycerol)] (DOPG) into Tradescantia virginiana stamen hair cells. During interphase, the 60-nm wide DOPG vesicles moved inside the cytoplasm comparably to organelles. During cytokinesis, they were transported through the phragmoplast and accumulated in the cell plate region together with the endogenous vesicles, even inside the central cell plate region. Because at this stage microtubules are virtually absent from that region, while actin filaments are present, actin filaments may have a role in the transport of vesicles toward the cell plate. Unlike the endogenous vesicles, the synthetic DOPG vesicles did not fuse with the developing cell plate. Instead, they redistributed into the cytoplasm of the daughter cells upon completion of cytokinesis. Because the redistribution of the vesicles occurs when actin filaments disappear from the phragmoplast, actin filaments may be involved in keeping the vesicles inside the developing cell plate region.     

Mutants of Escherichia coli that do not contain 1,4-diaminobutane (putrescine) or spermidine.

Strains of Escherichia coli K12 have been constructed which do not contain any of the polyamines normally present in a wild type strain, namely, 1,4-diaminobutane (putrescine) and spermidine. This phenotype arises as a consequence of the assembly into these strains of deletion mutations in speA (arginine decarboxylase), speB (agmatine ureohydrolase), speC (ornithine decarboxylase), and speD (adenosylmethionine decarboxylase). The polyamine-deficient strains grow indefinitely in the absence of polyamines but with a growth rate one-third of that found in the presence of polyamines. These strains can act as hosts for bacteriophages T4, T7, and f2, although the latter phage is poorly adsorbed; they can also maintain F' factors, ColE1 and P1 plasmids, and lysogeny by bacteriophage lambda. In contrast, the production of bacteriophage lambda in the absence of polyamines is strikingly decreased (greater than 99%) either after infection of a nonlysogen or after induction of a lysogen. A polyamine-deficient Hfr strain can transfer its chromosome to a recipient at a normal rate, but the number of recombinants observed in a cross is decreased approximately 300-fold. No such effect is observed when only the F- recipient strain in a cross is polyamine deficient.     

Mutations causing defects in the biosynthesis and response to gibberellins, abscisic acid and phytochrome B do not inhibit vernalization in Arabidopsis fca-1

 The roles of gibberellins, abscisic acid and phytochrome B in the vernalization response were investigated by combining mutations causing defects in their biosynthesis and response with the Arabidopsis thaliana (L.) Heynh. fca-1 mutation. The fca-1 mutation confers a very late-flowering phenotype which can be reversed to wild-type flowering if the seedlings are vernalized. Vernalization was unaffected in ga1-3, gai, abi1-1, abi2-1, abi3-1 and phyB-1 backgrounds, suggesting that gibberellin action mediated via GA1 and GAI, abscisic acid action mediated through ABI1 and ABI2, and phytochrome B, function independently of vernalization. However, the mutations did interact with fca-1 to change flowering time in the absence of vernalization. The abi1 fca-1 and abi2 fca-1 double mutants flowered earlier than fca-1 implying a role for abscisic acid in floral repression. Combination of ga1-3 or gai with fca-1 unexpectedly resulted in opposite interactions, with gai partially suppressing the late flowering of fca-1. Received: 17 July 1999 / Accepted: 11 October 1999     

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.