Synthesis of oleyl oleate wax esters in Arabidopsis thaliana and Camelina sativa seed oil

Seed oil composed of wax esters with long‐chain monoenoic acyl moieties represents a high‐value commodity for industry. Such plant‐derived sperm oil‐like liquid wax esters are biodegradable and can have excellent properties for lubrication. In addition, wax ester oil may represent a superior substrate for biodiesel production. In this study, we demonstrate that the low‐input oil seed crop Camelina sativa can serve as a biotechnological platform for environmentally benign wax ester production. Two biosynthetic steps catalysed by a fatty alcohol‐forming acyl‐CoA reductase (FAR) and a wax ester synthase (WS) are sufficient to achieve wax ester accumulation from acyl‐CoA substrates. To produce plant‐derived sperm oil‐like liquid wax esters, the WS from Mus musculus (MmWS) or Simmondsia chinensis (ScWS) were expressed in combination with the FAR from Mus musculus (MmFAR1) or Marinobacter aquaeolei (MaFAR) in seeds of Arabidopsis thaliana and Camelina sativa. The three analysed enzyme combinations Oleo3:mCherry:MmFAR1?c/Oleo3:EYFP:MmWS, Oleo3:mCherry:MmFAR1?c/ScWS and MaFAR/ScWS showed differences in the wax ester molecular species profiles and overall biosynthetic performance. By expressing MaFAR/ScWS in Arabidopsis or Camelina up to 59% or 21% of the seed oil TAGs were replaced by wax esters, respectively. This combination also yielded wax ester molecular species with highest content of monounsaturated acyl moieties. Expression of the enzyme combinations in the Arabidopsis fae1 fad2 mutant background high in oleic acid resulted in wax ester accumulation enriched in oleyl oleate (18:1/18:1 > 60%), suggesting that similar values may be obtained with a Camelina high oleic acid line.     

The TGF-alpha precursor expressed on the cell surface binds to the EGF receptor on adjacent cells, leading to signal transduction

The 50 amino acid form of TGF-alpha is cleaved from a conserved integral membrane glycoprotein by a protease that, in many tumor cells, appears to be limiting. To test whether the membrane-bound precursor has biological activity in the absence of processing, we introduced amino acid substitutions at the proteolytic cleavage sites. BHK cells transfected with expression vectors containing these altered sequences do not secrete detectable levels of mature TGF-alpha into the medium, but express high levels of proTGF-alpha at the cell surface. Coincubation of these BHK cells with A431 cells demonstrates that membrane-bound proTGF-alpha may bind to EGF receptors on the surface of contiguous cells, induce receptor autophosphorylation, and thereby produce a rapid rise in A431 intracellular calcium levels. Thus, proTGF-alpha can be biologically active in the absence of processing, a fact that may have implications for the integral membrane precursors of related growth factors.     

Neural Dynamics of Learning Sound—Action Associations

A motor component is pre-requisite to any communicative act as one must inherently move to communicate. To learn to make a communicative act, the brain must be able to dynamically associate arbitrary percepts to the neural substrate underlying the pre-requisite motor activity. We aimed to investigate whether brain regions involved in complex gestures (ventral pre-motor cortex, Brodmann Area 44) were involved in mediating association between novel abstract auditory stimuli and novel gestural movements. In a functional resonance imaging (fMRI) study we asked participants to learn associations between previously unrelated novel sounds and meaningless gestures inside the scanner. We use functional connectivity analysis to eliminate the often present confound of ‘strategic covert naming’ when dealing with BA44 and to rule out effects of non-specific reductions in signal. Brodmann Area 44, a region incorporating Broca''s region showed strong, bilateral, negative correlation of BOLD (blood oxygen level dependent) response with learning of sound-action associations during data acquisition. Left-inferior-parietal-lobule (l-IPL) and bilateral loci in and around visual area V5, right-orbital-frontal-gyrus, right-hippocampus, left-para-hippocampus, right-head-of-caudate, right-insula and left-lingual-gyrus also showed decreases in BOLD response with learning. Concurrent with these decreases in BOLD response, an increasing connectivity between areas of the imaged network as well as the right-middle-frontal-gyrus with rising learning performance was revealed by a psychophysiological interaction (PPI) analysis. The increasing connectivity therefore occurs within an increasingly energy efficient network as learning proceeds. Strongest learning related connectivity between regions was found when analysing BA44 and l-IPL seeds. The results clearly show that BA44 and l-IPL is dynamically involved in linking gesture and sound and therefore provides evidence that one of the mechanisms required for the evolution of human communication is found within these motor regions.     

Comparative analysis of stress responses of H9c2 rat cardiomyoblasts following treatment with doxorubicin and tBOOH

Cardiotoxicity is the major dose-limiting adverse effect of anthracyclines and is hypothesized to result from damage induced by reactive oxygen species (ROS) or inhibition of topoisomerase II. Here, we comparatively analyzed the effect of doxorubicin and the organic peroxide tertiary-butylhydroperoxide (tBOOH) on stress responses of rat cardiomyblast cells (H9c2). Moreover, we investigated the impact of serum factors and the novel prototypical protein kinase CK2 inhibitor resorufin on the sensentivity of H9c2 cells exposed to doxorubicin or tBOOH. Measuring cell viability by use of the WST assay as well as cell cycle progression and apoptotic death by FACS-based methods, we found that the sensitivity of H9c2 cells to doxorubicin and tBOOH was differently affected by both serum factors and resorufin. Formation of reactive oxygen species was observed after exposure of H9c2 cells to high doses (i.e. ≥5 μM) of doxorubicin only. Moreover, the antioxidant N-acetylcysteine protected H9c2 cells from cytotoxicity provoked by tBOOH but not doxorubicin. Analyzing the phosphorylation level of genotoxic stress responsive protein kinases and histone H2AX, which is indicative of an activated DNA damage response (DDR), we found that resorufin modulates doxorubicin- and tBOOH-induced responses in an agent specific manner. Taken together, the data indicate that (i) oxidative injury is not the most relevant type of damage triggering cell death of H9c2 cells following doxorubicin treatment, (ii) serum factors differently influence the sensitivity of cardiomyoblasts to doxorubicin and tBOOH and (iii) inhibition of CK2 unequally affects doxorubicin- and tBOOH-induced DDR of rat cardiomyoblasts.     

Sequence length bounds for resolving a deep phylogenetic divergence

In evolutionary biology, genetic sequences carry with them a trace of the underlying tree that describes their evolution from a common ancestral sequence. The question of how many sequence sites are required to recover this evolutionary relationship accurately depends on the model of sequence evolution, the substitution rate, divergence times and the method used to infer phylogenetic history. A particularly challenging problem for phylogenetic methods arises when a rapid divergence event occurred in the distant past. We analyse an idealised form of this problem in which the terminal edges of a symmetric four-taxon tree are some factor (λ) times the length of the interior edge. We determine an order λ² lower bound on the growth rate for the sequence length required to resolve the tree (independent of any particular branch length). We also show that this rate of sequence length growth can be achieved by existing methods (including the simple ‘maximum parsimony’ method), and compare these order λ² bounds with an order λ growth rate for a model that describes low-homoplasy evolution. In the final section, we provide a generic bound on the sequence length requirement for a more general class of Markov processes.