Maize Provitamin A Carotenoids, Current Resources, and Future Metabolic Engineering Challenges

Vitamin A deficiency is a serious global health problem that can be alleviated by improved nutrition. Development of cereal crops with increased provitamin A carotenoids can provide a sustainable solution to eliminating vitamin A deficiency worldwide. Maize is a model for cereals and a major staple carbohydrate source. Here, we discuss maize carotenogenesis with regard to pathway regulation, available resources, and current knowledge for improving carotenoid content and levels of provitamin A carotenoids in edible maize endosperm. This knowledge will be applied to improve the nutritional composition of related Poaceae crops. We discuss opportunities and challenges for optimizing provitamin A carotenoid biofortification of cereal food crops

A novel, on-line bioinformatics exercise in genomics

The teaching objective of the ‘Functional Genomics Lab’ is to engage the students by having them incorporate benchwork with web-based technology and analyse data in a biological context. The Functional Genomics practical involves sequencing and analysing an Arabidopsis gene. The on-line component introduces the undergraduates to a number of general and plant-specific databases, bioinformatics principles and tools. The practical is run over several sessions. The first two involve extracting and sequencing DNA to identify an ‘unknown’ gene. During the third session students analyse their sequence to predict a function and perform hypothetical experiments to test their hypotheses. Online exercises include PCR and RNAi vector design, microarray and promoter analysis using general and plant specific web-based tools

Regulation of Carotenoid Composition and Shoot Branching in Arabidopsis by a Chromatin Modifying Histone Methyltransferase, SDG8

Carotenoid pigments are critical for plant survival, and carotenoid composition is tuned to the developmental stage, tissue, and to environmental stimuli. We report the cloning of the CAROTENOID CHLOROPLAST REGULATORY1 (CCR1) gene. The ccr1 mutant has increased shoot branching and altered carotenoid composition, namely, reduced lutein in leaves and accumulation of cis-carotenes in dark-grown seedlings. The CCR1 gene was previously isolated as EARLY FLOWERING IN SHORT DAYS and encodes a histone methyltransferase (SET DOMAIN GROUP 8) that methylates histone H3 on Lys 4 and/or 36 (H3K4 and H3K36). ccr1 plants show reduced trimethyl-H3K4 and increased dimethyl-H3K4 surrounding the CAROTENOID ISOMERASE (CRTISO) translation start site, which correlates with low levels of CRTISO mRNA. Microarrays of ccr1 revealed the downregulation of 85 genes, including CRTISO and genes associated with signaling and development, and upregulation of just 28 genes. The reduction in CRTISO transcript abundance explains the altered carotenoid profile. The changes in shoot branching are additive with more axillary branching mutants, but the altered carotenoid profile may partially affect shoot branching, potentially by perturbed biosynthesis of the carotenoid substrates of strigolactones. These results are consistent with SDG8 regulating shoot meristem activity and carotenoid biosynthesis by modifying the chromatin surrounding key genes, including CRTISO. Thus, the level of lutein, the most abundant carotenoid in higher plants that is critical for photosynthesis and photoprotection, appears to be regulated by a chromatin modifying enzyme in Arabidopsis thaliana

Role of the Arabidopsis PIN6 auxin transporter in auxin homeostasis and auxin-mediated development

Plant-specific PIN-formed (PIN) efflux transporters for the plant hormone auxin are required for tissue-specific directional auxin transport and cellular auxin homeostasis. The Arabidopsis PIN protein family has been shown to play important roles in developmental processes such as embryogenesis, organogenesis, vascular tissue differentiation, root meristem patterning and tropic growth. Here we analyzed roles of the less characterised Arabidopsis PIN6 auxin transporter. PIN6 is auxin-inducible and is expressed during multiple auxin–regulated developmental processes. Loss of pin6 function interfered with primary root growth and lateral root development. Misexpression of PIN6 affected auxin transport and interfered with auxin homeostasis in other growth processes such as shoot apical dominance, lateral root primordia development, adventitious root formation, root hair outgrowth and root waving. These changes in auxin-regulated growth correlated with a reduction in total auxin transport as well as with an altered activity of DR5-GUS auxin response reporter. Overall, the data indicate that PIN6 regulates auxin homeostasis during plant development.Christopher I. Cazzonelli, Marleen Vanstraelen, Sibu Simon, Kuide Yin, Ashley Carron-Arthur, Nazia Nisar, Gauri Tarle, Abby J. Cuttriss¤, Iain R. Searle, Eva Benkova, Ulrike Mathesius, Josette Masle, Jiří Friml, Barry J. Pogso

Identifying chloroplast biogenesis and signalling mutants in Arabidopsis thaliana

The chloroplast is the largest and arguably the most complex of the three energy organelles in the plant cell. The biogenesis of the chloroplast requires a combination of thousands of proteins encoded by the chloroplastic and nuclear genomes. Chloroplas

The promoter of the Arabidopsis PIN6 auxin transporter enabled strong expression in the vasculature of roots, leaves, floral stems and reproductive organs

Cellular auxin homeostasis controls many aspects of plant growth, organogenesis and development. The existence of intracellular auxin transport mediated by endoplasmic reticulum (ER)-localized PIN 5, PIN 6 and PIN 8 proteins is a relatively recent discovery shaping a new era in understanding auxin-mediated growth processes. Here we summarize the importance of PIN 6 in mediating intracellular auxin transport during root formation, leaf vein patterning and nectar production. While, it was previously shown that PIN6 was strongly expressed in rosette leaf cell types important in vein formation, here we demonstrate by use a PIN6 promoter-reporter fusion, that PIN6 is also preferentially expressed in the vasculature of the primary root, cotyledons, cauline leaves, floral stem, sepals and the main transmitting tract of the reproductive silique. The strong, vein- specific reporter gene expression patterns enabled by the PIN6 promoter emphasizes that transcriptional control is likely to be a major regulator of PIN 6 protein levels, during vasculature formation, and supports the need for ER-localized PIN proteins in selecting specialized cells for vascular function in land plants

Regulation of Lutein Biosynthesis and Prolamellar Body Formation in Arabidopsis

Carotenoids are critical for photosynthetic function in chloroplasts, and are essential for the formation of the prolamellar body in the etioplasts of dark-grown (etiolated) seedlings. They are also precursors for plant hormones in both types of plastids. Lutein is one of the most abundant carotenoids found in both plastids. In this study we examine the regulation of lutein biosynthesis and investigate the effect of perturbing carotenoid biosynthesis on the formation of the lattice-like membranous structure of etioplasts, the prolamellar body (PLB). Analysis of mRNA abundance in wildtype and lutein-deficient mutants, lut2 and ccr2, in response to light transitions and herbicide treatments demonstrated that the mRNA abundance of the carotenoid isomerase (CRTISO) and epsilon-cyclase (εLCY) can be rate limiting steps in lutein biosynthesis. We show that accumulation of tetra-cis-lycopene and all-trans-lycopene correlates with the abundance of mRNA of several carotenoid biosynthetic genes. Herbicide treatments that inhibit carotenoid biosynthetic enzymes in wildtype and ccr2 etiolated seedlings were used to demonstrate that the loss of the PLB in ccr2 mutants is a result of perturbations in carotenoid accumulation, not indirect secondary effects, as PLB formation could be restored in ccr2 mutants treated with norflurazon