Gibberellin: inhibitor of an inhibitor of...?

Gibberellin is an endogenous plant growth regulator. Here, we describe our present understanding of how gibberellin regulates plant growth, using recent results gained from studies of gibberellin-signalling mutants of Arabidopsis. These results show that a signalling pathway represses plant growth and that gibberellin releases this repression. In consequence, the well-known growth-promoting properties of gibberellin are due to its activity as an "inhibitor of an inhibitor" [Brian Pw. Sym Soc. Exp Bio 1957; 11:166-182 (Ref. 1)] of plant growth.     

Regenerant Arabidopsis lineages display a distinct genome-wide spectrum of mutations conferring variant phenotypes

Multicellular organisms can be regenerated from totipotent differentiated somatic cell or nuclear founders [1-3]. Organisms regenerated from clonally related isogenic founders might a priori have been expected to be phenotypically invariant. However, clonal regenerant animals display variant phenotypes caused by defective epigenetic reprogramming of gene expression [2], and clonal regenerant plants exhibit poorly understood heritable phenotypic ("somaclonal") variation [4-7]. Here we show that somaclonal variation in regenerant Arabidopsis lineages is associated with genome-wide elevation in DNA sequence mutation rate. We also show that regenerant mutations comprise?a distinctive molecular spectrum of base substitutions, insertions, and deletions that probably results from decreased DNA repair fidelity. Finally, we show that while regenerant base substitutions are a likely major genetic cause of the somaclonal variation of regenerant Arabidopsis lineages, transposon movement is unlikely to contribute substantially to that variation. We conclude that the phenotypic variation of regenerant plants, unlike that of regenerant animals, is substantially due to DNA sequence mutation.     

ROS-mediated vascular homeostatic control of root-to-shoot soil Na delivery in Arabidopsis

Sodium (Na) is ubiquitous in soils, and is transported to plant shoots via transpiration through xylem elements in the vascular tissue. However, excess Na is damaging. Accordingly, control of xylem-sap Na concentration is important for maintenance of shoot Na homeostasis, especially under Na stress conditions. Here we report that shoot Na homeostasis of Arabidopsis thaliana plants grown in saline soils is conferred by reactive oxygen species (ROS) regulation of xylem-sap Na concentrations. We show that lack of A. thaliana respiratory burst oxidase protein F (AtrbohF; an NADPH oxidase catalysing ROS production) causes hypersensitivity of shoots to soil salinity. Lack of AtrbohF-dependent salinity-induced vascular ROS accumulation leads to increased Na concentrations in root vasculature cells and in xylem sap, thus causing delivery of damaging amounts of Na to the shoot. We also show that the excess shoot Na delivery caused by lack of AtrbohF is dependent upon transpiration. We conclude that AtrbohF increases ROS levels in wild-type root vasculature in response to raised soil salinity, thereby limiting Na concentrations in xylem sap, and in turn protecting shoot cells from transpiration-dependent delivery of excess Na.     

Transient gibberellin application promotes Arabidopsis thaliana hypocotyl cell elongation without maintaining transverse orientation of microtubules on the outer tangential wall of epidermal cells

The phytohormone gibberellin (GA) promotes plant growth by stimulating cellular expansion. Whilst it is known that GA acts by opposing the growth-repressing effects of DELLA proteins, it is not known how these events promote cellular expansion. Here we present a time-lapse analysis of the effects of a single pulse of GA on the growth of Arabidopsis hypocotyls. Our analyses permit kinetic resolution of the transient growth effects of GA on expanding cells. We show that pulsed application of GA to the relatively slowly growing cells of the unexpanded light-grown Arabidopsis hypocotyl results in a transient burst of anisotropic cellular growth. This burst, and the subsequent restoration of initial cellular elongation rates, occurred respectively following the degradation and subsequent reappearance of a GFP-tagged DELLA (GFP-RGA). In addition, we used a GFP-tagged α-tubulin 6 (GFP-TUA6) to visualise the behaviour of microtubules (MTs) on the outer tangential wall (OTW) of epidermal cells. In contrast to some current hypotheses concerning the effect of GA on MTs, we show that the GA-induced boost of hypocotyl cell elongation rate is not dependent upon the maintenance of transverse orientation of the OTW MTs. This confirms that transverse alignment of outer face MTs is not necessary to maintain rapid elongation rates of light-grown hypocotyls. Together with future studies on MT dynamics in other faces of epidermal cells and in cells deeper within the hypocotyl, our observations advance understanding of the mechanisms by which GA promotes plant cell and organ growth.     

Gibberellins control Arabidopsis hypocotyl growth via regulation of cellular elongation

The gibberellins (GAs) are endogenous regulators of plant growth. Experiments are described here that test the hypothesis that GA regulates hypocotyl growth by altering the extent of hypocotyl cell elongation. These experiments use GA-deficient and altered GA-response mutants of Arabidopsis thaliana (L.) Heyhn. It is shown that GA regulates elongation, in both light- and dark-grown hypocotyls, by influencing the rate and final extent of cellular elongation. However, light- and dark-grown hypocotyls exhibit markedly different GA dose-response relationships. The length of dark-grown hypocotyls is relatively unaffected by exogenous GA, whilst light-grown hypocotyl length is significantly increased by exogenous GA. Further analysis suggests that GA control of hypocotyl length is close to saturation in dark-grown hypocotyls, but not in light grown hypocotyls. The results show that a large range of possible hypocotyl lengths is achieved via dose-dependent GA-regulated alterations in the degree of elongation of individual hypocotyl cells.Key words: Arabidopsis, cell elongation, gibberellin (GA), GA mutants, hypocotyl.      

The Ultrastructural Study on the Early Embryonic Development of Radish

The ultrastructure of the mature embryo sac, the early stages of the embryo and endosperm development of common radish, Raphanur sativus was examined. The embryo sac consists of 7 cells with antipodal ceils disappeared when it matures. The egg cell is highly polarized. The wall surrounded the chalazal end of the egg cell is incomplete, showing a discontinuous structure of an electron dense material deposited intermittently in the space between the two plasma membranes of the egg cell and central cell. The synergid has filiform apparatus, rich in organelles and well developed ER. The two polar nuclei of the central cell are located near the egg apparatus because of the big vacuole, and the finger-like protrutions from the cell wall, as that in synergid, are found. The first division of the zygote occurs 4–5 days after pollination and the development of the embryo follows the Onagrad type, and the structure of the embryo cell is quite simple for containing small quantity of ER, plastids and other organelles. The primary endosperm nucleus deviates 2 days earlier than zygote. The endosperm is of nuclear-endosperm containing chloroplasts, well developed ER, and plentiful of mitochondria and golgi bodies and the nodule-like aggregation in both. the chalazal and micropylar ends of the embryo sac during the early development appeared, and cell wall starting at the micropylar end by freely-growing forms about 16 days after pollination.     

Enantioselective metabolism of primaquine by human CYP2D6

Given the threat of resistance of human malaria parasites, including to artemisinin derivatives, new agents are needed. Chloroquine (CQ) has been the most widely used anti-malarial, and new analogs (CQAns) presenting alkynes and side chain variations with high antiplasmodial activity were evaluated. Six diaminealkyne and diaminedialkyne CQAns were evaluated against CQ-resistant (CQ-R) (W2) and CQ-sensitive (CQ-S) (3D7) Plasmodium falciparum parasites in culture. Drug cytotoxicity to a human hepatoma cell line (HepG2) evaluated, allowed to calculate the drug selectivity index (SI), a ratio of drug toxicity to activity in vitro. The CQAns were re-evaluated against CQ-resistant and -sensitive P. berghei parasites in mice using the suppressive test. Docking studies with the CQAns and the human (Hss LDH) or plasmodial lactate dehydrogenase (Pf LDH) enzymes, and, a β-haematin formation assay were performed using a lipid as a catalyst to promote crystallization in vitro. All tested CQAns were highly active against CQ-R P. falciparum parasites, exhibiting half-maximal inhibitory concentration (IC50) values below 1 μΜ. CQAn33 and CQAn37 had the highest SIs. Docking studies revealed the best conformation of CQAn33 inside the binding pocket of Pf LDH; specificity between the residues involved in H-bonds of the Pf LDH with CQAn37. CQAn33 and CQAn37 were also shown to be weak inhibitors of Pf LDH. CQAn33 and CQAn37 inhibited β-haematin formation with either a similar or a 2-fold higher IC50 value, respectively, compared with CQ. CQAn37 was active in mice with P. berghei, reducing parasitaemia by 100%. CQAn33, -39 and -45 also inhibited CQ-resistant P. berghei parasites in mice, whereas high doses of CQ were inactive. The presence of an alkyne group and the size of the side chain affected anti-P. falciparum activity in vitro. Docking studies suggested a mechanism of action other than Pf LDH inhibition. The β-haematin assay suggested the presence of an additional mechanism of action of CQAn33 and CQAn37. Tests with CQAn34, CQAn37, CQAn39 and CQAn45 confirmed previous results against P. berghei malaria in mice, and CQAn33, 39 and 45 were active against CQ-resistant parasites, but CQAn28 and CQAn34 were not. The result likely reflects structure-activity relationships related to the resistant phenotype.     

Gibberellin deficiency and response mutations suppress the stem elongation phenotype of phytochrome-deficient mutants of Arabidopsis.

Plant growth and development are regulated by numerous internal and external factors. Among these, gibberellin (GA) (an endogenous plant growth regulator) and phytochrome (a photoreceptor) often influence the same processes. For example, in plants grown in the light Arabidopsis thaliana hypocotyl elongation is reduced by GA deficiency and increased by phytochrome deficiency. Here we describe experiments in which the phenotypes of Arabidopsis plants doubly homozygous for GA-related and phytochrome-related mutations were examined. The double mutants were studied at various stages in the plant life cycle, including the seed germination, young seedling, adult, and reproductive phases of development. The results of these experiments are complex, but indicate that a fully functional GA system is necessary for full expression of the elongated phenotypes conferred by phytochrome deficiency.     

Fruit Growth in Arabidopsis Occurs via DELLA-Dependent and DELLA-Independent Gibberellin Responses

Fruit growth and development depend on highly coordinated hormonal activities. The phytohormone gibberellin (GA) promotes growth by inducing degradation of the growth-repressing DELLA proteins; however, the extent to which DELLA proteins contribute to GA-mediated gynoecium and fruit development remains to be clarified. Here, we provide an in-depth characterization of the role of DELLA proteins in Arabidopsis thaliana fruit growth. We show that DELLA proteins are key regulators of reproductive organ size and important for ensuring optimal fertilization. We demonstrate that the seedless fruit growth (parthenocarpy) observed in della mutants can be directly attributed to the constitutive activation of GA signaling. It has been known for >75 years that another hormone, auxin, can induce formation of seedless fruits. Using mutants with complete lack of DELLA activity, we show here that auxin-induced parthenocarpy occurs entirely through GA signaling in Arabidopsis. Finally, we uncover the existence of a DELLA-independent GA response that promotes fruit growth. This response requires GIBBERELLIN-INSENSITIVE DWARF1–mediated GA perception and a functional 26S proteasome and involves the basic helix-loop-helix protein SPATULA as a key component. Taken together, our results describe additional complexities in GA signaling during fruit development, which may be particularly important to optimize the conditions for successful reproduction.