S. P. Vaughan

Molecular Characterization of Rht-1 Dwarfing Genes in Hexaploid Wheat

The introduction of the Reduced height (Rht)-B1b and Rht-D1b semidwarfing genes led to impressive increases in wheat (Triticum aestivum) yields during the Green Revolution. The reduction in stem elongation in varieties containing these alleles is caused by a limited response to the phytohormone gibberellin (GA), resulting in improved resistance to stem lodging and yield benefits through an increase in grain number. Rht-B1 and Rht-D1 encode DELLA proteins, which act to repress GA-responsive growth, and their mutant alleles Rht-B1b and Rht-D1b are thought to confer dwarfism by producing more active forms of these growth repressors. While no semidwarfing alleles of Rht-A1 have been identified, we show that this gene is expressed at comparable levels to the other homeologs and represents a potential target for producing novel dwarfing alleles. In this study, we have characterized additional dwarfing mutations in Rht-B1 and Rht-D1. We show that the severe dwarfism conferred by Rht-B1c is caused by an intragenic insertion, which results in an in-frame 90-bp insertion in the transcript and a predicted 30-amino acid insertion within the highly conserved amino-terminal DELLA domain. In contrast, the extreme dwarfism of Rht-D1c is due to overexpression of the semidwarfing Rht-D1b allele, caused by an increase in gene copy number. We show also that the semidwarfing alleles Rht-B1d and Rht-B1e introduce premature stop codons within the amino-terminal coding region. Yeast two-hybrid assays indicate that these newly characterized mutations in Rht-B1 and Rht-D1 confer “GA-insensitive” dwarfism by producing DELLA proteins that do not bind the GA receptor GA INSENSITIVE DWARF1, potentially compromising their targeted degradation.

Heterologous expression and transcript analysis of gibberellin biosynthetic genes of grasses reveals novel functionality in the GA3ox family

BackgroundThe gibberellin (GA) pathway plays a central role in the regulation of plant development, with the 2-oxoglutarate-dependent dioxygenases (2-ODDs: GA20ox, GA3ox, GA2ox) that catalyse the later steps in the biosynthetic pathway of particularly importance in regulating bioactive GA levels. Although GA has important impacts on crop yield and quality, our understanding of the regulation of GA biosynthesis during wheat and barley development remains limited. In this study we identified or assembled genes encoding the GA 2-ODDs of wheat, barley and Brachypodium distachyon and characterised the wheat genes by heterologous expression and transcript analysis.ResultsThe wheat, barley and Brachypodium genomes each contain orthologous copies of the GA20ox, GA3ox and GA2ox genes identified in rice, with the exception of OsGA3ox1 and OsGA2ox5 which are absent in these species. Some additional paralogs of 2-ODD genes were identified: notably, a novel gene in the wheat B genome related to GA3ox2 was shown to encode a GA 1-oxidase, named as TaGA1ox-B1. This enzyme is likely to be responsible for the abundant 1β-hydroxylated GAs present in developing wheat grains. We also identified a related gene in barley, located in a syntenic position to TaGA1ox-B1, that encodes a GA 3,18-dihydroxylase which similarly accounts for the accumulation of unusual GAs in barley grains. Transcript analysis showed that some paralogs of the different classes of 2-ODD were expressed mainly in a single tissue or at specific developmental stages. In particular, TaGA20ox3, TaGA1ox1, TaGA3ox3 and TaGA2ox7 were predominantly expressed in developing grain. More detailed analysis of grain-specific gene expression showed that while the transcripts of biosynthetic genes were most abundant in the endosperm, genes encoding inactivation and signalling components were more highly expressed in the seed coat and pericarp.ConclusionsThe comprehensive expression and functional characterisation of the multigene families encoding the 2-ODD enzymes of the GA pathway in wheat and barley will provide the basis for a better understanding of GA-regulated development in these species. This analysis revealed the existence of a novel, endosperm-specific GA 1-oxidase in wheat and a related GA 3,18-dihydroxylase enzyme in barley that may play important roles during grain expansion and development.

Interactions between root restriction, irrigation and rootstock treatments on the growth and cropping of ‘Queen Cox’ apple trees: Effects on orchard growth and cropping

Summary ‘Queen Cox’ apple trees on M.9 or MM.106 rootstocks were planted within root restrictive membranes buried in the soil and compared with trees planted without membranes. Half of the treated and control trees were irrigated using a trickle system and the other half did not have supplementary irrigation. Root restriction reduced greatly the extension shoot growth on the trees, by reducing the numbers and mean length of the shoots. Shoot growth on restricted trees terminated very early in the season. Irrigation, in contrast, increased the numbers of shoots and the mean individual shoot length. Root restriction reduced the mean individual size of spur and extension shoot leaves. Significant interactions between root restriction, irrigation and rootstock treatments were recorded over six seasons. Irrigation only partially alleviated the reductions in growth induced by root restriction. Root restriction increased the numbers of floral buds formed per unit trunk cross-sectional area in all years. The overall effect of root restriction was to reduce (by 50%) the annual and cumulative yields per tree, whilst supplementary irrigation had the reverse effect, increasing cumulative yields by approximately 100%. The treatments had similar effects on the yields of Class I (>65.mm diameter) fruits. Yield efficiency (yield per trunk cross-sectional area) was increased in all years by the irrigation treatment, but decreased in two of the four years by root restriction.

Effects of tree age at planting, root manipulation and trickle irrigation on growth and cropping of apple (Malus pumila) cultivar Queen Cox on M.9 rootstock

Summary The apple cultivar Queen Cox on M.9 rootstock cropped more precociously when planted as two year old trees than when planted as one year old trees, even though there were no significant differences in the sizes (leader height and branch length) of the trees at the time of planting. However, the two year old trees had larger root systems at planting. As the trees aged, those planted as one year olds grew more vigorously and bore higher cumulative yields than those planted as two year olds. Annual root pruning of the trees, commencing 15 months after planting, reduced extension shoot growth, crown volume and grubbing weights (final fresh weights of scions) severely. In some seasons root pruning increased the number of spur and terminal floral buds produced and also the final sets and yield efficiencies on the treated trees. Planting trees within semi-permeable fabric membranes also reduced extension shoot growth and tree size, but less severely than the root-pruning treatment. Root restriction increased the efficiency of fruit set and yields and also improved the grades of fruits produced. Trickle irrigation treatments increased shoot growth and tree fresh weight at the time of grubbing, but had inconsistent and small effects on fruit set and yields. Interactions between tree age at the time of planting and the root manipulative treatments were significant.

Interactions between root restriction, irrigation and rootstock treatments on ‘Queen Cox’ apple trees: Effects of soil and plant water relations

Summary The purpose of this research was to determine if established, orchard-grown apple trees that were con®ned in root-restricting membranes received adequate water for growth when irrigated. Previous data had shown positive effects of root restriction on reducing shoot extension growth in apple. Soil matric potential (Ψsm), leaf stomatal conductance (g) and leaf water potentials (Ψl) were measured over daily cycles during the period of maximum tree water use (July and August). Measurements were also made of the Ψsm and fruit size throughout the growing season. Daily and seasonal Ψsm showed that the soil within the root restricting membranes (+R) when irrigated (+I) remained closed to field capacity (<100 HPa), which was not the case for unirrigated soil within the membranes (-I +R). Ψl measured before dawn, showed that similar levels of drought stress were evident between irrigated and restricted (+I +R) and unrestricted (+I -R and -I -R) trees. The Ψsm, and Ψl for trees with roots within restricting membranes were significantly more negative, in the absence of irrigation (-I +R). Measurement of g showed that root restricted trees were transpiring at similar rates in the presence of irrigation (+I +R) as unrestricted trees (+I -R) with or without irrigation. Stomatal closure could not explain the increased Ψl observed for the restricted irrigated (+I +R) treatment compared with unrestricted (-R) trees. A reduced stomatal aperture was the most likely explanation for the reduction in growth previously observed with the restricted unirrigated trees. Fruit size was also affected by root restriction and the effect became greater as the roots became more restricted with tree age. Similarly, there was also a negative effect, in one year, of root restriction on fruit size at harvest, even in the presence of irrigation. Data show that reductions in soil water availability, Ψl and g, for the root-restricted trees (+I +R), were unlikely to be the causes for the previously observed reductions in shoot growth (tree size). These results imply that other factors were in operation, among which root-synthesized chemical regulators of shoot growth are the most likely candidates.

Building the new international science of the agriculture-food-water-environment Nexus in China and the world

Abstract The multiple, complex and systemic problems of the agriculture–food–water–environment nexus (“Nexus”) are among the most significant challenges of the 21st century. China is a key site for Nexus research amidst profound socio‐environmental problems. The policy implications of these problems have been authoritatively summarized elsewhere. This study presents discussions at an international workshop in Guangzhou that asked instead “What science is needed to deliver the growing policy commitments regarding these challenges? And, What changes are needed to the science itself?” Understanding and effective intervention regarding the Nexus calls for a paradigm shift: to a new kind of science of (capacity for) international, interdisciplinary, and impactful research working with and within complex socio‐natural systems. We here argue that science must become proactive in approach, striving only for “minimal harm” not “silver bullet” solutions, and adopting an explicitly long‐term strategic perspective. Together, these arguments lead to calls for reorienting science and science policy in three ways: from short‐term remediation to longer‐term optimization; from a focus on environmental threats to one on the opportunities for international collaborative learning; and toward supporting new forms of scientific career. We bring these points together by recommending a new form of scientific institution: a global network of collaborative Nexus Centres, under the umbrella of a global Food Nexus Organization akin to those of the human genome and proteome.

Targeting transgene expression in apple tree tissues: Roles for heterologous and homologous promoters

Targeting transgene expression to specific cells and tissues is desirable for a number of reasons (Gittins et al., 2000) not least of which is the requirement to reduce perceived risks with GM technology. In fruit crops, where invariably the product is consumed fresh, this assumes unparalleled importance. To this end, the ability of several heterologous and homologous gene promoters to drive expression of the βglucuronidase (gusA) marker gene in the vegetative tissues of transgenic apple (Malus pumila, Mill.) have and are being tested. These promoters originally drove expression in leaves (Rubisco, small subunit (SSU), RBCS3C from tomato, SRS1 from soybean), in vascular tissue (rolC, and CoYMV) and in root tissue (extA from Brassica). Homologous promoters from apple were cloned for expression in fruit. Transgenic lines were produced by Agrobacterium-mediated transformation and the levels of gusA activity in the vegetative tissues of young plants were compared with those using the CaMV 35S promoter to drive expression of the same gene. Quantitative GUS data were related to the copy number of transgene loci assessed by Southern blotting. The precise location of GUS activity in each tissue was identified by staining of whole leaves and tissue sections with the chromogenic substrate X-Gluc. Light-regulation and patterns of expression are recorded in various vegetative tissues of apple.