Ann E. Stapleton

Natural Genetic Variation in Lycopene Epsilon Cyclase Tapped for Maize Biofortification

Dietary vitamin A deficiency causes eye disease in 40 million children each year and places 140 to 250 million at risk for health disorders. Many children in sub-Saharan Africa subsist on maize-based diets. Maize displays considerable natural variation for carotenoid composition, including vitamin A precursors α-carotene, β-carotene, and β-cryptoxanthin. Through association analysis, linkage mapping, expression analysis, and mutagenesis, we show that variation at the lycopene epsilon cyclase (lcyE) locus alters flux down α-carotene versus β-carotene branches of the carotenoid pathway. Four natural lcyE polymorphisms explained 58% of the variation in these two branches and a threefold difference in provitamin A compounds. Selection of favorable lcyE alleles with inexpensive molecular markers will now enable developing-country breeders to more effectively produce maize grain with higher provitamin A levels.

Flavonoids can protect maize DNA from the induction of ultraviolet radiation damage

Diverse flavonoid compounds are widely distributed in angiosperm families. Flavonoids absorb radiation in the ultraviolet (UV) region of the spectrum, and it has been proposed that these compounds function as UV filters. We demonstrate that the DNA in Zea mays plants that contain flavonoids (primarily anthocyanins) is protected from the induction of damage caused by UV radiation relative to the DNA in plants that are genetically deficient in these compounds. DNA damage was measured with a sensitive and simple assay using individual monoclonal antibodies, one specific for cyclobutane pyrimidine dimer damage and the other specific for pyrimidine(6,4)pyrimidone damage.

Ultraviolet radiation and plants: burning questions

Plants use sunlight for photosynthesis and, as a consequence, are exposed to the ultraviolet (UV) radiation that is present in sunlight. UV radiation is generally divided into three classes: UV-C, UV-6, and UV-A. The UV-C region of the UV spectrum includes wavelengths below 280 nm; these highly energetic wavelengths are effectively absorbed by ozone in the stratosphere and, thus, are not present in sunlight at the earth’s surface. UV-C wavelengths will be removed from the light reaching the earth’s surface’so long as there is any ozone present (Caldwell et al., 1989). In contrast, UV radiation in the UV-B region, from 280 to 320 nm, does reach ground level. The UV-B portion of sunlight has received much attention in recent years because irradiation from this spectral region (especially 297 to 310 nm) will increase as the stratospheric ozone concentration decreases (Caldwell et al., 1989). Currently, ozone decreases result from chlorofluorocarbon contamination of the stratosphere (McFarland and Kaye, 1992). UV wavelengths from 320 to 390 nm, which make up the UV-A region of the spectrum, are not attenuated by ozone, so their fluence will be unaffected by ozone layer reduction. Like all living organisms, plants sense and respond to UV radiation, both the wavelengths present in sunlight (UV-A and UV-B) and the wavelengths below 280 nm (UV-C). AI1 types of UV radiation are known to damage various plant processes. Such damage can be classified into two categories: damage to DNA (which can cause heritable mutations) and damage to physiological processes. There has been much speculation about how increased UV radiation exposure will affect plants, but as yet, there are no definitive answers. In this review, I will discuss the kinds of damage that UV radiation can inflict on plants, the mechanisms plants use to perceive and respond to UV radiation, and the ecological relevance of UV light wavelengths that have been used in the experimental analysis of plant responses to UV radiation.

The iPlant Collaborative: Cyberinfrastructure for Plant Biology

The iPlant Collaborative (iPlant) is a United States National Science Foundation (NSF) funded project that aims to create an innovative, comprehensive, and foundational cyberinfrastructure in support of plant biology research (PSCIC, 2006). iPlant is developing cyberinfrastructure that uniquely enables scientists throughout the diverse fields that comprise plant biology to address Grand Challenges in new ways, to stimulate and facilitate cross-disciplinary research, to promote biology and computer science research interactions, and to train the next generation of scientists on the use of cyberinfrastructure in research and education. Meeting humanitys projected demands for agricultural and forest products and the expectation that natural ecosystems be managed sustainably will require synergies from the application of information technologies. The iPlant cyberinfrastructure design is based on an unprecedented period of research community input, and leverages developments in high-performance computing, data storage, and cyberinfrastructure for the physical sciences. iPlant is an open-source project with application programming interfaces that allow the community to extend the infrastructure to meet its needs. iPlant is sponsoring community-driven workshops addressing specific scientific questions via analysis tool integration and hypothesis testing. These workshops teach researchers how to add bioinformatics tools and/or datasets into the iPlant cyberinfrastructure enabling plant scientists to perform complex analyses on large datasets without the need to master the command-line or high-performance computational services.

CDC55, a Saccharomyces cerevisiae gene involved in cellular morphogenesis: identification, characterization, and homology to the B subunit of mammalian type 2A protein phosphatase.

Microscopic screening of a collection of cold-sensitive mutants of Saccharomyces cerevisiae led to the identification of a new gene, CDC55, which appears to be involved in the morphogenetic events of the cell cycle. CDC55 maps between CDC43 and CHC1 on the left arm of chromosome VII. At restrictive temperature, the original cdc55 mutant produces abnormally elongated buds and displays a delay or partial block of septation and/or cell separation. A cdc55 deletion mutant displays a cold-sensitive phenotype like that of the original isolate. Sequencing of CDC55 revealed that it encodes a protein of about 60 kDa, as confirmed by Western immunoblots using Cdc55p-specific antibodies. This protein has greater than 50% sequence identity to the B subunits of rabbit skeletal muscle type 2A protein phosphatase; the latter sequences were obtained by analysis of peptides derived from the purified protein, a polymerase chain reaction product, and cDNA clones. An extragenic suppressor of the cdc55 mutation lies in BEM2, a gene previously identified on the basis of an apparent role in bud emergence.

Solar Ultraviolet-B Radiation Affects Seedling Emergence, DNA Integrity, Plant Morphology, Growth Rate, and Attractiveness to Herbivore Insects in Datura ferox.

To study functional relationships between the effects of solar ultraviolet-B radiation (UV-B) on different aspects of the physiology of a wild plant, we carried out exclusion experiments in the field with the summer annual Datura ferox L. Solar UV-B incident over Buenos Aires reduced daytime seedling emergence, inhibited stem elongation and leaf expansion, and tended to reduce biomass accumulation during early growth. However, UV-B had no effect on calculated net assimilation rate. Using a monoclonal antibody specific to the cyclobutane-pyrimidine dimer (CPD), we found that plants receiving full sunlight had more CPDs per unit of DNA than plants shielded from solar UV-B, but the positive correlation between UV-B and CPD burden tended to level off at high (near solar) UV-B levels. At our field site, Datura plants were consumed by leaf beetles (Coleoptera), and the proportion of plants attacked by insects declined with the amount of UV-B received during growth. Field experiments showed that plant exposure to solar UV-B reduced the likelihood of leaf beetle attack by one-half. Our results highlight the complexities associated with scaling plant responses to solar UV-B, because they show: (a) a lack of correspondence between UV-B effects on net assimilation rate and whole-plant growth rate, (b) nonlinear UV-B dose-response curves, and (c) UV-B effects of plant attractiveness to natural herbivores.

Ultraviolet Radiation Alters Maize Phyllosphere Bacterial Diversity

Epiphytic bacteria are subjected to very stressful environments, including UV radiation. Bacterial assemblages on Zea mays (maize) leaves exposure were examined with and without UV-B radiation. Culture-independent molecular techniques were utilized for bacterial identification, diversity analysis and selection of putative UV exposure marker sequences. Few sequences corresponded to previously characterized phyllosphere bacteria. There was a strong tendency toward increased 16S rDNA sequence diversity in UV samples. Overall community structure was assessed using denaturing gel gradient electrophoresis; significant alterations in community structure were found in comparisons of phyllosphere bacterial samples from control and solar UV-B exposed plants.

Maize Leaf Epiphytic Bacteria Diversity Patterns Are Genetically Correlated with Resistance to Fungal Pathogen Infection

Plant leaves host a specific set of microbial epiphytes. Plant genetic and solar UV-B radiation effects on the diversity of the phyllosphere were examined by measuring epiphytic bacterial ribosomal DNA diversity in a maize recombinant inbred (RI) mapping population. Several chromosomal quantitative trait loci (QTL) with significant effects on bacterial diversity were identified, some of which had effects only in the presence of UV-B radiation and others that had effects both with and without UV-B. Candidate genes with allele-specific effects were mapped to the bacterial diversity chromosomal regions. A glutamate decarboxylase candidate gene was located at a UV-B-specific chromosomal locus, and in a comparison between two RI lines with contrasting bacterial diversity phenotypes, high bacterial diversity was associated with high levels of glutamate decarboxylase enzyme activity, a component of the gamma-aminobutyric acid (GABA) pathway. The bacterial diversity loci exhibited a significant overlap with loci connected with Southern leaf blight (SLB) susceptibility in the field. A SLB-resistant inbred genotype had less beta bacterial diversity, and antibiotic treatment of inbreds increased this diversity. These results suggest that the GABA pathway is genetically associated with phyllosphere bacterial diversity. Furthermore, the colocalization of QTL between low bacterial diversity and fungal blight-resistance and the increase in beta diversity in antibiotic-treated leaves suggest that occupation of leaf habitats by a particular set of suppressive bacteria may restrict phyllosphere bacterial variability and increase resistance to fungal infection.

The maize epicuticular wax layer provides UV protection

As surface ultraviolet-B (UV-B) radiation levels increase due to the decline in the protective stratospheric ozone layer, ultraviolet radiation sunscreens will become more important for all plant species that grow in sunlight. Epicuticular waxes, which cover the aerial portions of all terrestrial plants, are ideally located to be sunscreens. The sun-screening ability of maize (Zea mays L.) epicuticular waxes was tested using the glossy1 mutant, which is specifically defective in juvenile wax production. A significant difference between the glossy1 mutant and wild type was seen in UV-induced leaf rolling and in some measurements of UV-induced DNA damage levels under enhanced UV. Isolated epicuticular wax layers absorbed significant amounts of UV, and leaves with wax absorbed more UV than leaves with little wax. Thus, by some measures, the epicuticular waxy layer acts as an ultraviolet radiation protectant in maize.

A simple and sensitive antibody-based method to measure UV-induced DNA damage in Zea mays

We describe a western blotting procedure in which immobilized DNA is reacted with monoclonal antibodies specific to individual types of DNA damage. This method is applicable to small DNA samples and is more sensitive than the standard assay.