Perrin H. Beatty

Fertilizing Nature: A Tragedy of Excess in the Commons

Why has nitrogen fertilizer use declined in some countries while increasing in others, despite significant environmental harm? Proper crop management strategies offer environmental and economic benefits without sacrificing yields.

Future Prospects for Cereals That Fix Nitrogen

What are the next steps in engineering crop plants that fix nitrogen and make their own fertilizer? Nitrogen availability is limiting to plant growth and has long been overcome through applications of synthetic nitrogen-rich fertilizer. This has revolutionized crop yield and food production worldwide, but at substantial economic and environmental cost (fixed N2; a

The APETALA-2-Like Transcription Factor OsAP2-39 Controls Key Interactions between Abscisic Acid and Gibberellin in Rice

100 billion per year global industry and environmental nitrogen pollution). Indeed, in April, the Edinburgh Declaration on Reactive Nitrogen was the most recent call for global action to address this source of nitrogen pollution. At the same time, the Bill and Melinda Gates Foundation convened a small meeting of international researchers to assess the way forward in reducing dependence on fertilizers through engineering crop plants that “fix” nitrogen themselves to sustain their growth and yield. The discussions of recent advances indicate that there are indeed viable options to achieve this goal.

Nitrogen use efficiencies of spring barley grown under varying nitrogen conditions in the field and growth chamber

The interaction between phytohormones is an important mechanism which controls growth and developmental processes in plants. Deciphering these interactions is a crucial step in helping to develop crops with enhanced yield and resistance to environmental stresses. Controlling the expression level of OsAP2-39 which includes an APETALA 2 (AP2) domain leads to phenotypic changes in rice. Overexpression of OsAP2-39 leads to a reduction in yield by decreasing the biomass and the number of seeds in the transgenic rice lines. Global transcriptome analysis of the OsAP2-39 overexpression transgenic rice revealed the upregulation of a key Abscisic Acid (ABA) biosynthetic gene OsNCED-I which codes for 9-cis-epoxycarotenoid dioxygenase and leads to an increase in the endogenous ABA level. In addition to OsNCED-1, the gene expression analysis revealed the upregulation of a gene that codes for the Elongation of Upper most Internode (EUI) protein, an enzyme that catalyzes 16α, 17-epoxidation of non-13-hydroxylated GAs, which has been shown to deactivate gibberellins (GAs) in rice. The exogenous application of GA restores the wild-type phenotype in the transgenic line and ABA application induces the expression of EUI and suppresses the expression of OsAP2-39 in the wild-type line. These observations clarify the antagonistic relationship between ABA and GA and illustrate a mechanism that leads to homeostasis of these hormones. In vivo and in vitro analysis showed that the expression of both OsNCED-1 and EUI are directly controlled by OsAP2-39. Together, these results reveal a novel mechanism for the control of the ABA/GA balance in rice which is regulated by OsAP2-39 that in turn regulates plant growth and seed production.

The Rice R2R3-MYB Transcription Factor OsMYB55 Is Involved in the Tolerance to High Temperature and Modulates Amino Acid Metabolism

BACKGROUND AND AIMS Nitrogen-use efficiency (NUE) of cereals needs to be improved by nitrogen (N) management, traditional plant breeding methods and/or biotechnology, while maintaining or, optimally, increasing crop yields. The aims of this study were to compare spring-barley genotypes grown on different nitrogen levels in field and growth-chamber conditions to determine the effects on N uptake (NUpE) and N utilization efficiency (NUtE) and ultimately, NUE. METHODS Morphological characteristics, seed yield and metabolite levels of 12 spring barley (Hordeum vulgare) genotypes were compared when grown at high and low nitrogen levels in field conditions during the 2007 and 2008 Canadian growing seasons, and in potted and hydroponic growth-chamber conditions. Genotypic NUpE, NUtE and NUE were calculated and compared between field and growth-chamber environments. KEY RESULTS Growth chamber and field tests generally showed consistent NUE characteristics. In the field, Vivar, Excel and Ponoka, showed high NUE phenotypes across years and N levels. Vivar also had high NUE in growth-chamber trials, showing NUE across complex to simplistic growth environments. With the high NUE genotypes grown at low N in the field, NUtE predominates over NUpE. N metabolism-associated amino acid levels were different between roots (elevated glutamine) and shoots (elevated glutamate and alanine) of hydroponically grown genotypes. In field trials, metabolite levels were different between Kasota grown at high N (elevated glutamine) and Kasota at low N plus Vivar at either N condition. CONCLUSIONS Determining which trait(s) or gene(s) to target to improve barley NUE is important and can be facilitated using simplified growth approaches to help determine the NUE phenotype of various genotypes. The genotypes studied showed similar growth and NUE characteristics across field and growth-chamber tests demonstrating that simplified, low-variable growth environments can help pinpoint genetic targets for improving spring barley NUE.

Transcriptome analysis of nitrogen‐efficient rice over‐expressing alanine aminotransferase

Temperatures higher than the optimum negatively affects plant growth and development. Tolerance to high temperature is a complex process that involves several pathways. Understanding this process, especially in crops such as rice, is essential to prepare for predicted climate changes due to global warming. Here, we show that OsMYB55 is induced by high temperature and overexpression of OsMYB55 resulted in improved plant growth under high temperature and decreased the negative effect of high temperature on grain yield. Transcriptome analysis revealed an increase in expression of several genes involved in amino acids metabolism. We demonstrate that OsMYB55 binds to the promoter regions of target genes and directly activates expression of some of those genes including glutamine synthetase (OsGS1;2) glutamine amidotransferase (GAT1) and glutamate decarboxylase 3 (GAD3). OsMYB55 overexpression resulted in an increase in total amino acid content and of the individual amino acids produced by the activation of the above mentioned genes and known for their roles in stress tolerance, namely L-glutamic acid, GABA and arginine especially under high temperature condition. In conclusion, overexpression of OsMYB55 improves rice plant tolerance to high temperature, and this high tolerance is associated with enhanced amino acid metabolism through transcription activation.

The Genetics of Nitrogen Use Efficiency in Crop Plants

Crop plants require nitrogen for key macromolecules, such as DNA, proteins and metabolites, yet they are generally inefficient at acquiring nitrogen from the soil. Crop producers compensate for this low nitrogen utilization efficiency by applying nitrogen fertilizers. However, much of this nitrogen is unavailable to the plants as a result of microbial uptake and environmental loss of nitrogen, causing air, water and soil pollution. We engineered rice over-expressing alanine aminotransferase (AlaAT) under the control of a tissue-specific promoter that showed a strong nitrogen use efficiency phenotype. In this study, we examined the transcriptome response in roots and shoots to the over-expression of AlaAT to provide insights into the nitrogen-use-efficient phenotype of these plants. Transgenic and control rice plants were grown hydroponically and the root and shoot gene expression profiles were analysed using Affymetrix Rice GeneChip microarrays. Transcriptome analysis revealed that there was little impact on the transgenic transcriptome compared with controls, with 0.11% and 0.07% differentially regulated genes in roots and shoots, respectively. The most up-regulated transcripts, a glycine-rich cell wall (GRP) gene and a gene encoding a hypothetical protein (Os8823), were expressed in roots. Another transgenic root-specific up-regulated gene was leucine rich repeat (LRR). Genes induced in the transgenic shoots included GRP, LRR, acireductone dioxygenase (OsARD), SNF2 ATP-translocase and a putative leucine zipper transcription factor. This study provides a genome-wide view of the response to AlaAT over-expression, and elucidates some of the genes that may play a role in the nitrogen-use-efficient phenotype.

Two Sets of Paralogous Genes Encode the Enzymes Involved in the Early Stages of Clavulanic Acid and Clavam Metabolite Biosynthesis in Streptomyces clavuligerus

In the past 50 years, the application of synthetic nitrogen (N) fertilizer to farmland resulted in a dramatic increase in crop yields but with considerable negative impacts on the environment. New solutions are therefore needed to simultaneously increase yields while maintaining, or preferably decreasing, applied N to maximize the nitrogen use efficiency (NUE) of crops. In this review, we outline the definition of NUE, the selection and development of NUE crops, and the factors that interact with NUE. In particular, we emphasize the challenges of developing crop plants with enhanced NUE, using more classical genetic approaches based on utilizing existing allelic variation for NUE traits. The challenges of phenotyping, mapping quantitative trait loci (QTLs), and selecting candidate genes for NUE improvement are described. In addition, we highlight the importance of different factors that lead to changes in the NUE components of nitrogen uptake efficiency (NUpE) and nitrogen utilization efficiency (NUtE).

Five Additional Genes Are Involved in Clavulanic Acid Biosynthesis in Streptomyces clavuligerus

ABSTRACT Recently, a second copy of a gene encoding proclavaminate amidinohydrolase (pah1), an enzyme involved in the early stages of clavulanic acid and clavam metabolite biosynthesis in Streptomyces clavuligerus, was identified and isolated. Using Southern analysis, we have now isolated second copies of the genes encoding the carboxyethylarginine synthase (ceaS) and β-lactam synthetase (bls) enzymes. These new paralogues are given the gene designations ceaS1 and bls1 and are located immediately upstream of pah1 on the chromosome. Furthermore, sequence analysis of the region downstream of pah1 revealed a second copy of a gene encoding ornithine acetyltransferase (oat1), thus indicating the presence of a cluster of paralogue genes. ceaS1, bls1, and oat1 display 73, 60, and 63% identities, respectively, at the nucleotide level to the original ceaS2, bls2, and oat2 genes from the clavulanic acid gene cluster. Single mutants defective in ceaS1, bls1, or oat1 were prepared and characterized and were found to be affected to variable degrees in their ability to produce clavulanic acid and clavam metabolites. Double mutants defective in both copies of the genes were also prepared and tested. The ceaS1/ceaS2 and the bls1/bls2 mutant strains were completely blocked in clavulanic acid and clavam metabolite biosynthesis. On the other hand, oat1/oat2 double mutants still produced some clavulanic acid and clavam metabolites. This may be attributed to the presence of the argJ gene in S. clavuligerus, which encodes yet another ornithine acetyltransferase enzyme that may be able to compensate for the lack of OAT1 and -2 in the double mutants.

Sulfolane degradation by mixed cultures and a bacterial isolate identified as a Variovorax sp.

ABSTRACT An approximately 12.5-kbp region of DNA sequence from beyond the end of the previously described clavulanic acid gene cluster was analyzed and found to encode nine possible open reading frames (ORFs). Involvement of these ORFs in clavulanic acid biosynthesis was assessed by creating mutants with defects in each of the ORFs. orf12 and orf14 had been previously reported to be involved in clavulanic acid biosynthesis. Now five additional ORFs are shown to play a role, since their mutation results in a significant decrease or total absence of clavulanic acid production. Most of these newly described ORFs encode proteins with little similarity to others in the databases, and so their roles in clavulanic acid biosynthesis are unclear. Mutation of two of the ORFs, orf15 and orf16, results in the accumulation of a new metabolite, N-acetylglycylclavaminic acid, in place of clavulanic acid. orf18 and orf19 encode apparent penicillin binding proteins, and while mutations in these genes have minimal effects on clavulanic acid production, their normal roles as cell wall biosynthetic enzymes and as targets for β-lactam antibiotics, together with their clustered location, suggest that they are part of the clavulanic acid gene cluster.