A High Power‐Density,Mediator‐Free,Microfluidic Biophotovoltaic Device for Cyanobacterial Cells

Biophotovoltaics has emerged as a promising technology for generating renewable energy because it relies on living organisms as inexpensive, self‐repairing, and readily available catalysts to produce electricity from an abundant resource: sunlight. The efficiency of biophotovoltaic cells, however, has remained significantly lower than that achievable through synthetic materials. Here, a platform is devised to harness the large power densities afforded by miniaturized geometries. To this effect, a soft‐lithography approach is developed for the fabrication of microfluidic biophotovoltaic devices that do not require membranes or mediators. Synechocystis sp. PCC 6803 cells are injected and allowed to settle on the anode, permitting the physical proximity between cells and electrode required for mediator‐free operation. Power densities of above 100 mW m^‐2 are demonstrated for a chlorophyll concentration of 100 μM under white light, which is a high value for biophotovoltaic devices without extrinsic supply of additional energy.     

2-O Heparan Sulfate Sulfation by Hs2st Is Required for Erk/Mapk Signalling Activation at the Mid-Gestational Mouse Telencephalic Midline

Heparan sulfate (HS) is a linear carbohydrate composed of polymerized uronate-glucosamine disaccharide units that decorates cell surface and secreted glycoproteins in the extracellular matrix. In mammals HS is subjected to differential sulfation by fifteen different heparan sulfotransferase (HST) enzymes of which Hs2st uniquely catalyzes the sulfation of the 2-O position of the uronate in HS. HS sulfation is postulated to be important for regulation of signaling pathways by facilitating the interaction of HS with signaling proteins including those of the Fibroblast Growth Factor (Fgf) family which signal through phosphorylation of extracellular signal-regulated kinases Erk1/2. In the developing mouse telencephalon Fgf2 signaling regulates proliferation and neurogenesis. Loss of Hs2st function phenocopies the thinned cerebral cortex of mutant mice in which Fgf2 or Erk1/2 function are abrogated, suggesting the hypothesis that 2-O-sulfated HS structures play a specific role in Fgf2/Erk signaling pathway in this context in vivo. This study investigated the molecular role of 2-O sulfation in Fgf2/Erk signaling in the developing telencephalic midline midway through mouse embryogenesis at E12.5. We examined the expression of Hs2st, Fgf2, and Erk1/2 activity in wild-type and Hs2st^-/- mice. We found that Hs2st is expressed at high levels at the midline correlating with high levels of Erk1/2 activation and Erk1/2 activation was drastically reduced in the Hs2st^-/- mutant at the rostral telencephalic midline. We also found that 2-O sulfation is specifically required for the binding of Fgf2 protein to Fgfr1, its major cell-surface receptor at the rostral telencephalic midline. We conclude that 2-O sulfated HS structures generated by Hs2st are needed to form productive signaling complexes between HS, Fgf2 and Fgfr1 that activate Erk1/2 at the midline. Overall, our data suggest the interesting possibility that differential expression of Hs2st targets the rostral telencephalic midline for high levels of Erk signaling by increasing the sensitivity of cells to an Fgf2 signal that is rather more widespread.     

Computational Identification of Post Translational Modification Regulated RNA Binding Protein Motifs

RNA and its associated RNA binding proteins (RBPs) mitigate a diverse array of cellular functions and phenotypes. The interactions between RNA and RBPs are implicated in many roles of biochemical processing by the cell such as localization, protein translation, and RNA stability. Recent discoveries of novel mechanisms that are of significant evolutionary advantage between RBPs and RNA include the interaction of the RBP with the 3’ and 5’ untranslated region (UTR) of target mRNA. These mechanisms are shown to function through interaction of a trans-factor (RBP) and a cis-regulatory element (3’ or 5’ UTR) by the binding of a RBP to a regulatory-consensus nucleic acid motif region that is conserved throughout evolution. Through signal transduction, regulatory RBPs are able to temporarily dissociate from their target sites on mRNAs and induce translation, typically through a post-translational modification (PTM). These small, regulatory motifs located in the UTR of mRNAs are subject to a loss-of-function due to single polymorphisms or other mutations that disrupt the motif and inhibit the ability to associate into the complex with RBPs. The identification of a consensus motif for a given RBP is difficult, time consuming, and requires a significant degree of experimentation to identify each motif-containing gene on a genomic scale. We have developed a computational algorithm to analyze high-throughput genomic arrays that contain differential binding induced by a PTM for a RBP of interest–RBP-PTM Target Scan (RPTS). We demonstrate the ability of this application to accurately predict a PTM-specific binding motif to an RBP that has no antibody capable of distinguishing the PTM of interest, negating the use of in-vitro exonuclease digestion techniques.     

Tissue stretch induces nuclear remodeling in connective tissue fibroblasts

Studies in cultured cells have shown that nuclear shape is an important factor influencing nuclear function, and that mechanical forces applied to the cell can directly affect nuclear shape. In a previous study, we demonstrated that stretching of whole mouse subcutaneous tissue causes dynamic cytoskeletal remodeling with perinuclear redistribution of α-actin in fibroblasts within the tissue. We have further shown that the nuclei of these fibroblasts have deep invaginations containing α-actin. In the current study, we hypothesized that tissue stretch would cause nuclear remodeling with a reduced amount of nuclear invagination, measurable as a change in nuclear concavity. Subcutaneous areolar connective tissue samples were excised from 28 mice and randomized to either tissue stretch or no stretch for 30 min, then examined with histochemistry and confocal microscopy. In stretched tissue (vs. non-stretched), fibroblast nuclei had a larger cross-sectional area (P < 0.001), smaller thickness (P < 0.03) in the plane of the tissue, and smaller relative concavity (P < 0.005) indicating an increase in nuclear convexity. The stretch-induced loss of invaginations may have important influences on gene expression, RNA trafficking and/or cell differentiation.     

Early seed fall and seedling emergence: precursors to tropical restoration

We explore processes of seed immigration and seedling recruitment before an experimental rainforest restoration matures enough to affect either. Twenty-four 30 × 30-m plots were fenced in 12 ha of pasture in 2006. Seeds were collected in ninety-six 1-m⁻² seed traps; recruits were censused in ~12,000 m² in which establishment was allowed. We tested effects of distance from forest, living trees, and stumps of trees cut during site preparation on seed rain in 2007 and effects of these and soil depth on recruits through June 2008. Seed fall and recruitment were not correlated with distance to forest 90–400 m away, nor to living shade trees outside the 160 × 485-m experimental grid. Recruitment differed for animal- and wind-dispersed species in a topographically complex landscape. Recruitment of wind-dispersed species was random with respect to soil depth or distance to recent stumps. Recruitment of animal-dispersed species was multimodal; partial correlations with number of stumps within 30 m of plots were significant with soil depth held constant (P < 0.025), as were correlations of recruitment with soil depth with number of stumps held constant (P < 0.01). Animal-dispersed recruits were often not conspecifics of adults that had been cut, indicating a legacy of attraction by fruiting trees of animals bearing seeds from distant sources. Ecological implications are that recruitment in pastures released from grazing reflects a mix of widely scattered wind-dispersed pioneers and, where animal-dispersed trees exist, multi-modal and decidedly non-random recruitment of pioneer and later successional animal-dispersed trees from seed banks.     

Stability of plant defense proteins in the gut of insect herbivores

Plant defense against insect herbivores is mediated in part by enzymes that impair digestive processes in the insect gut. Little is known about the evolutionary origins of these enzymes, their distribution in the plant kingdom, or the mechanisms by which they act in the protease-rich environment of the animal digestive tract. One example of such an enzyme is threonine (Thr) deaminase (TD), which in tomato (Solanum lycopersicum) serves a dual role in isoleucine (Ile) biosynthesis in planta and Thr degradation in the insect midgut. Here, we report that tomato uses different TD isozymes to perform these functions. Whereas the constitutively expressed TD1 has a housekeeping role in Ile biosynthesis, expression of TD2 in leaves is activated by the jasmonate signaling pathway in response to herbivore attack. Ingestion of tomato foliage by specialist (Manduca sexta) and generalist (Trichoplusia ni) insect herbivores triggered proteolytic removal of TD2's C-terminal regulatory domain, resulting in an enzyme that degrades Thr without being inhibited through feedback by Ile. This processed form (pTD2) of TD2 accumulated to high levels in the insect midgut and feces (frass). Purified pTD2 exhibited biochemical properties that are consistent with a postingestive role in defense. Shotgun proteomic analysis of frass from tomato-reared M. sexta identified pTD2 as one of the most abundant proteins in the excrement. Among the other tomato proteins identified were several jasmonate-inducible proteins that have a known or proposed role in anti-insect defense. Subtilisin-like proteases and other pathogenesis-related proteins, as well as proteins of unknown function, were also cataloged. We conclude that proteomic analysis of frass from insect herbivores provides a robust experimental approach to identify hyperstable plant proteins that serve important roles in defense.