The interplay of cyclic stretch and vascular endothelial growth factor in regulating the initial steps for angiogenesis

Angiogenesis is regulated by chemical and mechanical factors in vivo. The regulatory role of mechanical factors and how chemical and mechanical angiogenic regulators work in concert remains to be explored. We investigated the effect of cyclic uniaxial stretch (20%, 1 Hz), with and without the stimulation of vascular endothelial growth factor (VEGF), on sprouting angiogenesis by employing a stretchable three‐dimensional cell culture model. When compared to static controls, stretch alone significantly increased the density of endothelial sprouts, and these sprouts aligned perpendicular to the direction of stretch. The Rho‐associated kinase (ROCK) inhibitor Y27632 suppressed stretch‐induced sprouting angiogenesis and associated sprout alignment. While VEGF is a potent angiogenic stimulus through ROCK‐dependent pathways, the combination of VEGF and stretch did not have an additive effect on angiogenesis. In the presence of VEGF stimulation, the ROCK inhibitor suppressed stretch‐induced sprout alignment but did not affect stretch‐induced sprout density; in contrast, the receptor tyrosine kinase (RTK) inhibitor sunitinib had no effect on stretch‐induced alignment but trended toward suppressed stretch‐induced sprout density. Our results suggest that the formation of sprouts and their directionality do not have completely identical regulatory pathways, and thus it is possible to separately manipulate the number and pattern of new sprouts. © 2014 American Institute of Chemical Engineers Biotechnol. Prog., 31:248–257, 2015     

Large amounts of labile organic carbon in permafrost soils of northern Alaska

Permafrost‐affected soils of the northern circumpolar region represent 50% of the terrestrial soil organic carbon (SOC) reservoir and are most strongly affected by climatic change. There is growing concern that this vast SOC pool could transition from a net C sink to a source. But so far little is known on how the organic matter (OM) in permafrost soils will respond in a warming future, which is governed by OM composition and possible stabilization mechanisms. To investigate if and how SOC in the active layer and adjacent permafrost is protected against degradation, we employed density fractionation to separate differently stabilized SOM fractions. We studied the quantity and quality of OM in different compartments using elemental analysis, ¹³C solid‐phase nuclear magnetic resonance (¹³C‐NMR) spectroscopy, and ¹⁴C analyses. The soil samples were derived from 16 cores from drained thaw lake basins, ranging from 0 to 5500 years of age, representing a unique series of developing Arctic soils over time. The normalized SOC stocks ranged between 35.5 and 86.2 kg SOC m^?3, with the major amount of SOC located in the active layers. The SOC stock is dominated by large amounts of particulate organic matter (POM), whereas mineral‐associated OM especially in older soils is of minor importance on a mass basis. We show that tremendous amounts of over 25 kg OC per square meter are stored as presumably easily degradable OM rich in carbohydrates. Only about 10 kg OC per square meter is present as presumably more stable, mineral‐associated OC. Significant amounts of the easily degradable, carbohydrate‐rich OM are preserved in the yet permanently frozen soil below the permafrost table. Forced by global warming, this vast labile OM pool could soon become available for microbial degradation due to the continuous deepening of the annually thawing active layer.     

Divergent target recognition by coexpressed 5′-isomiRs of miR-142-3p and selective viral mimicry

Sequence heterogeneity at the ends of mature microRNAs (miRNAs) is well documented, but its effects on miRNA function are largely unexplored. Here we studied the impact of miRNA 5′-heterogeneity, which affects the seed region critical for target recognition. Using the example of miR-142-3p, an emerging regulator of the hematopoietic lineage in vertebrates, we show that naturally coexpressed 5′-variants (5′-isomiRs) can recognize largely distinct sets of binding sites. Despite this, both miR-142-3p isomiRs regulate exclusive and shared targets involved in actin dynamics. Thus, 5′-heterogeneity can substantially broaden and enhance regulation of one pathway. Other 5′-isomiRs, in contrast, recognize largely overlapping sets of binding sites. This is exemplified by two herpesviral 5′-isomiRs that selectively mimic one of the miR-142-3p 5′-isomiRs. We hypothesize that other cellular and viral 5′-isomiRs can similarly be grouped into those with divergent or convergent target repertoires, based on 5′-sequence features. Taken together, our results provide a detailed characterization of target recognition by miR-142-3p and its 5′-isomiR-specific viral mimic. We furthermore demonstrate that miRNA 5′-end variation leads to differential targeting and can thus broaden the target range of miRNAs.     

Brucella suis vaccine strain S2-infected immortalized caprine endometrial epithelial cell lines induce non-apoptotic ER-stress

Brucella, which is regarded as an intracellular pathogen responsible for a zoonotic disease called brucellosis, survives and proliferates within several types of phagocytic and non-phagocytic cells. Brucella infects not only their preferred hosts but also other domestic and wild animal species, inducing abortion and infertility. Therefore, the interaction between uterine cells and Brucella is important for understanding the pathogenesis of this disease. In this study, we describe the Brucella suis vaccine strain S2 (B.suis.S2) infection and replication in the immortalized caprine endometrial epithelial cell line hTERT-EECs and the induced cellular and molecular response modulation in vitro. We found that B.suis S2 was able to infect and replicate to high titers and inhibit the proliferation of EECs and induce non-apoptotic pathways, as determined by B.suis.S2 detection using MTT and acridine orange/ethidium bromide (AO/EB) staining and flow cytometry. We explored the evidence of non-apoptotic pathways using real-time quantitative RT-PCR and by western blot analysis. Finally, we discovered the over-expression of GRP78, ATF4, ATF6, PERK, eIF2α, CHOP, and cytochrome c (Cyt-c) but not IRE1, xbp-1, and caspase-3 in B.suis.S2 (HK)-attacked and B.suis.S2-infected cells, suggesting that the molecular mechanism of ER stress sensor activation by B.suis.S2 is basically concomitant with that by B.suis.S2 (HK) and that ER stress, especially the PERK pathway, plays an important role in the process of B.suis.S2 infecting EEC, which may, in part, explain the role of the uterus in the pathogenesis of B.suis.S2.

Electronic supplementary material

The online version of this article (doi:10.1007/s12192-014-0564-x) contains supplementary material, which is available to authorized users.     

Engineering temporal accumulation of a low recalcitrance polysaccharide leads to increased C6 sugar content in plant cell walls

Reduced cell wall recalcitrance and increased C6 monosaccharide content are desirable traits for future biofuel crops, as long as these biomass modifications do not significantly alter normal growth and development. Mixed‐linkage glucan (MLG), a cell wall polysaccharide only present in grasses and related species among flowering plants, is comprised of glucose monomers linked by both β‐1,3 and β‐1,4 bonds. Previous data have shown that constitutive production of MLG in barley (Hordeum vulgare) severely compromises growth and development. Here, we used spatio‐temporal strategies to engineer Arabidopsis thaliana plants to accumulate significant amounts of MLG in the cell wall by expressing the rice CslF6 MLG synthase using secondary cell wall and senescence‐associated promoters. Results using secondary wall promoters were suboptimal. When the rice MLG synthase was expressed under the control of a senescence‐associated promoter, we obtained up to four times more glucose in the matrix cell wall fraction and up to a 42% increase in saccharification compared to control lines. Importantly, these plants grew and developed normally. The induction of MLG deposition at senescence correlated with an increase of gluconic acid in cell wall extracts of transgenic plants in contrast to the other approaches presented in this study. MLG produced in Arabidopsis has an altered structure compared to the grass glucan, which likely affects its solubility, while its molecular size is unaffected. The induction of cell wall polysaccharide biosynthesis in senescing tissues offers a novel engineering alternative to enhance cell wall properties of lignocellulosic biofuel crops.     

Deep sequencing reveals a novel closterovirus associated with wild rose leaf rosette disease

A bizarre virus‐like symptom of a leaf rosette formed by dense small leaves on branches of wild roses (Rosa multiflora Thunb.), designated as ‘wild rose leaf rosette disease’ (WRLRD), was observed in China. To investigate the presumed causal virus, a wild rose sample affected by WRLRD was subjected to deep sequencing of small interfering RNAs (siRNAs) for a complete survey of the infecting viruses and viroids. The assembly of siRNAs led to the reconstruction of the complete genomes of three known viruses, namely Apple stem grooving virus (ASGV), Blackberry chlorotic ringspot virus (BCRV) and Prunus necrotic ringspot virus (PNRSV), and of a novel virus provisionally named ‘rose leaf rosette‐associated virus’ (RLRaV). Phylogenetic analysis clearly placed RLRaV alongside members of the genus Closterovirus, family Closteroviridae. Genome organization of RLRaV RNA (17 653 nucleotides) showed 13 open reading frames (ORFs), except ORF1 and the quintuple gene block, most of which showed no significant similarities with known viral proteins, but, instead, had detectable identities to fungal or bacterial proteins. Additional novel molecular features indicated that RLRaV seems to be the most complex virus among the known genus members. To our knowledge, this is the first report of WRLRD and its associated closterovirus, as well as two ilarviruses and one capilovirus, infecting wild roses. Our findings present novel information about the closterovirus and the aetiology of this rose disease which should facilitate its control. More importantly, the novel features of RLRaV help to clarify the molecular and evolutionary features of the closterovirus.