Engineering photosynthetic light capture: impacts on improved solar energy to biomass conversion

The main function of the photosynthetic process is to capture solar energy and to store it in the form of chemical 'fuels'. Increasingly, the photosynthetic machinery is being used for the production of biofuels such as bio-ethanol, biodiesel and bio-H₂. Fuel production efficiency is directly dependent on the solar photon capture and conversion efficiency of the system. Green algae (e.g. Chlamydomonas reinhardtii ) have evolved genetic strategies to assemble large light-harvesting antenna complexes (LHC) to maximize light capture under low-light conditions, with the downside that under high solar irradiance, most of the absorbed photons are wasted as fluorescence and heat to protect against photodamage. This limits the production process efficiency of mass culture. We applied RNAi technology to down-regulate the entire LHC gene family simultaneously to reduce energy losses by fluorescence and heat. The mutant Stm3LR3 had significantly reduced levels of LHCI and LHCII mRNAs and proteins while chlorophyll and pigment synthesis was functional. The grana were markedly less tightly stacked, consistent with the role of LHCII. Stm3LR3 also exhibited reduced levels of fluorescence, a higher photosynthetic quantum yield and a reduced sensitivity to photoinhibition, resulting in an increased efficiency of cell cultivation under elevated light conditions. Collectively, these properties offer three advantages in terms of algal bioreactor efficiency under natural high-light levels: (i) reduced fluorescence and LHC-dependent heat losses and thus increased photosynthetic efficiencies under high-light conditions; (ii) improved light penetration properties; and (iii) potentially reduced risk of oxidative photodamage of PSII.     

Development of a method for the purification and culture of rodent astrocytes

The inability to purify and culture astrocytes has long?hindered studies of their function. Whereas astrocyte progenitor cells can be cultured from neonatal brain, culture of mature astrocytes from postnatal brain has not been possible. Here, we report a new method to prospectively purify astrocytes by immunopanning. These astrocytes undergo apoptosis in culture, but vascular cells and HBEGF promote their survival in serum-free culture. We found that some developing astrocytes normally undergo apoptosis in?vivo and that the vast majority of astrocytes contact blood vessels, suggesting that?astrocytes are matched to blood vessels by competing for vascular-derived trophic factors such as HBEGF. Compared to traditional astrocyte cultures, the gene profiles of the cultured purified postnatal astrocytes much more closely resemble those of in?vivo astrocytes. Although these astrocytes strongly promote synapse formation and function, they do not secrete glutamate in response to stimulation.     

Microbiome Influences Prenatal and Adult Microglia in a Sex-Specific Manner

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Association of Interleukin-1 gene polymorphisms with central obesity and metabolic syndrome in a coronary heart disease population

The objective of this study was to determine whether single nucleotide polymorphisms (SNPs) in the Interleukin-1 (IL-1) gene family are associated with central obesity and metabolic syndrome in a coronary heart disease population. The IL-1α C-889T (rs1800587) and IL-1β +3954 (rs1143634) SNPs were studied in a Western Australian coronary heart disease (CHD) population (N = 556). Subjects who were TT homozygous at either SNP had larger waist circumference (IL-1α: 1.8 cm greater, P = 0.04; IL-1β: 4 cm greater, P = 0.0004) compared with major allele homozygotes. Individuals with two copies of the IL-1α:IL-1β T:T haplotype had greater waist circumference (4.7 cm greater, P = 0.0001) compared to other haplotypes. There was a significant interaction between the IL-1β SNP and BMI level on waist circumference (P = 0.01). When the cohort was stratified by median BMI, TT carriers for IL-1β with above median BMI had greater waist circumference (6.1 cm greater, P = 0.007) compared to baseline carriers, whilst no significant association was seen in the below median group. Similarly, when the cohort was stratified by median fibrinogen level (IL-1α interaction P = 0.01; IL-1β interaction P = 0.04), TT carriers for both SNPs in the above median fibrinogen group had greater waist circumference (IL-1α 2.7 cm greater, P = 0.007; IL-1β 3.3 cm greater, P = 0.003) compared with major allele homozygotes. This association was not seen in the below median group. Also, we found a trend of increased metabolic syndrome for IL-1β TT homozygotes (P = 0.07). In conclusion, our findings suggest that in a CHD population IL-1 gene polymorphisms may be involved in increased central obesity, and the genetic influences are more evident among patients who have a higher level of obesity or inflammatory markers.     

Intratumor heterogeneity in evolutionary models of tumor progression

With rare exceptions, human tumors arise from single cells that have accumulated the necessary number and types of heritable alterations. Each such cell leads to dysregulated growth and eventually the formation of a tumor. Despite their monoclonal origin, at the time of diagnosis most tumors show a striking amount of intratumor heterogeneity in all measurable phenotypes; such heterogeneity has implications for diagnosis, treatment efficacy, and the identification of drug targets. An understanding of the extent and evolution of intratumor heterogeneity is therefore of direct clinical importance. In this article, we investigate the evolutionary dynamics of heterogeneity arising during exponential expansion of a tumor cell population, in which heritable alterations confer random fitness changes to cells. We obtain analytical estimates for the extent of heterogeneity and quantify the effects of system parameters on this tumor trait. Our work contributes to a mathematical understanding of intratumor heterogeneity and is also applicable to organisms like bacteria, agricultural pests, and other microbes.     

Construction, characterization, and preliminary BAC-end sequencing analysis of a bacterial artificial chromosome library of white clover (Trifolium repens L.).

White clover (Trifolium repens L.) is a forage legume widely used in combination with grass in pastures because of its ability to fix nitrogen. We have constructed a bacterial artificial chromosome (BAC) library of an advanced breeding line of white clover. The library contains 37 248 clones with an average insert size of approximately 85 kb, representing an approximate 3-fold coverage of the white clover genome based on an estimated genome size of 960 Mb. The BAC library was pooled and screened by polymerase chain reaction (PCR) amplification using both white clover microsatellites and PCR-based markers derived from Medicago truncatula, resulting in an average of 6 hits per marker; this supports the estimated 3-fold genome coverage in this allotetraploid species. PCR-based screening of 766 clones with a multiplex set of chloroplast primers showed that only 0.5% of BAC clones contained chloroplast-derived inserts. The library was further evaluated by sequencing both ends of 724 of the clover BACs. These were analysed with respect to their sequence content and their homology to the contents of a range of plant gene, expressed sequence tag, and repeat element databases. Forty-three microsatellites were discovered in the BAC-end sequences (BESs) and investigated as potential genetic markers in white clover. The BESs were also compared with the partially sequenced genome of the model legume M. truncatula with the specific intention of identifying putative comparative-tile BACs, which represent potential regions of microsynteny between the 2 species; 14 such BACs were discovered. The results suggest that a large-scale BAC-end sequencing strategy has the potential to anchor a significant proportion of the genome of white clover onto the gene-space sequence of M. truncatula.     

Condensed tannins: co-occurrence of procyanidins,prodelphinidins and profisetinidins in the heartwood of Acacia baileyana

The flavonoids and condensed tannins of the heartwood of Acacia baileyana var. purpurea are described. In conformity with other Acacia species, the hydroxylation pattern of the flavonoids is of the resorcinol type but, in sharp contrast, the tannins are heterogeneous consisting of a mixture of the resorcinol and phtoroglucinol series. Dimeric proanthocyanidins of the phloroglucinol type were absent and this exception to the general observation that they invariably co-occur with the polymers may be explained by the relative nucleophilicity of the aromatic A-rings.     

Recognition of mitochondrial targeting sequences by the import receptors Tom20 and Tom22

The Tom20 and Tom22 receptor subunits of the TOM (translocase of the outer mitochondrial membrane) complex recognize N-terminal presequences of proteins that are to be imported into the mitochondrion. In plants, Tom20 is C-terminally anchored in the mitochondrial membrane, whereas Tom20 is N-terminally anchored in animals and fungi. Furthermore, the cytosolic domain of Tom22 in plants is smaller than its animal/fungal counterpart and contains fewer acidic residues. Here, NMR spectroscopy was used to explore presequence interactions with the cytosolic regions of receptors from the plant Arabidopsis thaliana and the fungus Saccharomyces cerevisiae (i.e., AtTom20, AtTom22, and ScTom22). It was found that AtTom20 possesses a discontinuous bidentate hydrophobic binding site for presequences. The presequences on plant mitochondrial proteins comprise two or more hydrophobic binding regions to match this bidentate site. NMR data suggested that while these presequences bind to ScTom22, they do not bind to AtTom22. AtTom22, however, binds to AtTom20 at the same binding site as presequences, suggesting that this domain competes with the presequences of imported proteins, thereby enabling their progression along the import pathway.     

A continuous spectrophotometric assay and nonlinear kinetic analysis of methionine γ-lyase catalysis

In this article, we present a new, easy-to-implement assay for methionine γ-lyase (MGL)-catalyzed γ-elimination reactions of l-methionine and its analogues that produce α-ketobutyrate (α-KB) as product. The assay employs ultraviolet–visible (UV–Vis) spectrophotometry to continuously monitor the rate of formation of α-KB by its absorbance at 315 nm. We also employ a nonlinear data analysis method that obviates the need for an “initial slope” determination, which can introduce errors when the progress curves are nonlinear. The spectrophotometric assay is validated through product analysis by ¹H NMR (nuclear magnetic resonance), which showed that under the conditions of study l-methionine (l-met) and l-methionine sulfone (l-met sulfone) substrates were converted to α-KB product with greater than 99% yield. Using this assay method, we determined for the first time the Michaelis–Menten parameters for a recombinant form of MGL from Porphyromonas gingivalis, obtaining respective k_cat and K_m values of 328 ± 8 min⁻¹ and 1.2 ± 0.1 mM for l-met γ-elimination and 2048 ± 59 min⁻¹ and 38 ± 2 mM for l-met sulfone γ-elimination reactions. We envisage that this assay method will be useful for determining the activity of MGL γ-elimination reactions that produce α-KB as the end product.