Viral and chloroplastic signals essential for initiation and efficiency of translation in Agrobacterium tumefaciens

The construction of high-level protein expression vectors using the CaMV 35S promoter in concert with highly efficient translation initiation signals for Agrobacterium tumefaciens is a relatively less explored field compared to that of Escherichia coli. In the current study, we experimentally investigated the capacity of the CaMV 35S promoter to direct GFP gene expression in A. tumefaciens in the context of different viral and chloroplastic translation initiation signals. GFP expression and concomitant translational efficiency was monitored by confocal microscopy and Western blot analysis. Among all of the constructs, the highest level of translation was observed for the construct containing the phage T7 translation initiation region followed by the chloroplastic Rubisco Large Subunit (rbcL) 58-nucleotide 5′ leader region including its SD-like sequence (GGGAGGG). Replacing the SD-like (GGGAGGG) with non SD-like (TTTATTT) or replacing the remaining 52 nucleotides of rbcL with nonspecific sequence completely abolished translation. In addition, this 58 nucleotide region of rbcL serves as a translational enhancer in plants when located within the 5′ UTR of mRNA corresponding to GFP. Other constructs, including those containing sequences upstream of the coat proteins of Alfalfa Mosaic Virus, or the GAGG sequence of T4 phage or the chloroplastic atpI and/or PsbA 5′ UTR sequence, supported low levels of GFP expression or none at all. From these studies, we propose that we have created high expression vectors in A. tumefaciens and/or plants which contain the CaMV 35S promoter, followed by the translationally strong T7 SD plus RBS translation initiation region or the rbcL 58-nucleotide 5′ leader region upstream of the gene of interest.     

Fern leaves and cauliflower curds are not fractals

The popular demonstration of drawing a mature fern leaf as expressed by Barnsley''s fractal method is mathematically and visually very attractive but anatomically and developmentally misleading, and thus has limited, if any, biological significance. The same is true for the fractal demonstration of the external features of cauliflower curds. Actual fern leaves and cauliflower curds have a very small number of anatomically variable and non-iterating bifurcations, which superficially look self-similar, but do not allow for scaling down of their structure as real fractals do. Moreover, fern leaves and cauliflower curds develop from the inside out through a process totally different from fractal drawing procedures. The above cases demonstrate a general problem of using mathematical tools to investigate or illustrate biological phenomena in an irrelevant manner. A realistic set of mathematical equations to describe fern leaf or cauliflower curd development is needed.     

The rapid determination in controlled environments of parameters for predicting seedling growth rates in natural conditions

Parameters which characterise the response of seedling growth rate to temperature and mean daily radiant exposure were determined for cultivars of eight vegetable species and two ornamental bedding plant species using controlled environments. All species were grown for approximately four weeks in a series of four controlled environments consisting of factorial combinations of 20°C or 15°C with 7.78 or 0.98 MJ m^-2 radiant exposure per day to photosynthetically active radiation. Four of the vegetable cultivars had been used previously in a study of growth in the glasshouse at different seasons. The parameters derived from controlled environments were used, with temperature and light measurements from the glasshouse, to predict seedling shoot growth in the glasshouse. These predictions accounted for 92% of the observed variance in log shoot dry weight in a series of glasshouse experiments involving thirteen combinations of species and sowing date, each grown at four plant densities and sampled on four occasions. Equivalent parameters derived previously from the glasshouse data themselves accounted for 92% of the variance in the same predictive exercise. Differences in base temperature for growth, the responsiveness of relative growth rate to daily radiant exposure (light), and the potential relative growth rate were apparent between species and were quantitatively characterised by the parameters. To determine these parameters for a cultivar requires the equivalent of one square metre of controlled environment growing area for approximately sixteen days. Such parameters could be applied for scheduling and management in crop production, particularly in transplant production.     

Molecular and functional characterization of the plastid-localized Phosphoenolpyruvate enolase (ENO1) from Arabidopsis thaliana

The Arabidopsis thaliana gene At1g74030 codes for a putative plastid phosphoenolpyruvate (PEP) enolase (ENO1). The recombinant ENO1 protein exhibited enolase activity and its kinetic properties were determined. ENO1 is localized to plastids and expressed in most heterotrophic tissues including trichomes and non-root-hair cells, but not in the mesophyll of leaves. Two T-DNA insertion eno1 mutants exhibited distorted trichomes and reduced numbers of root hairs as the only visible phenotype. The essential role of ENO1 in PEP provision for anabolic processes within plastids, such as the shikimate pathway, is discussed with respect to plastid transporters, such as the PEP/phosphate translocator.     

AtCCS is a functional homolog of the yeast copper chaperone Ccs1/Lys7

In plant chloroplasts two superoxide dismutase (SOD) activities occur, FeSOD and Cu/ZnSOD, with reciprocal regulation in response to copper availability. This system presents a unique model to study the regulation of metal-cofactor delivery to an organelle. The Arabidopsis thaliana gene AtCCS encodes a functional homolog to yeast Ccs1p/Lys7p, a copper chaperone for SOD. The AtCCS protein was localized to chloroplasts where it may supply copper to the stromal Cu/ZnSOD. AtCCS mRNA expression levels are upregulated in response to Cu-feeding and senescence. We propose that AtCCS expression is regulated to allow the most optimal use of Cu for photosynthesis.