Identification and Profiling of MicroRNAs from Skeletal Muscle of the Common Carp

The common carp is one of the most important cultivated species in the world of freshwater aquaculture. The cultivation of this species is particularly productive due to its high skeletal muscle mass; however, the molecular mechanisms of skeletal muscle development in the common carp remain unknown. It has been shown that a class of non-coding ∼22 nucleotide RNAs called microRNAs (miRNAs) play important roles in vertebrate development. They regulate gene expression through sequence-specific interactions with the 3′ untranslated regions (UTRs) of target mRNAs and thereby cause translational repression or mRNA destabilization. Intriguingly, the role of miRNAs in the skeletal muscle development of the common carp remains unknown. In this study, a small-RNA cDNA library was constructed from the skeletal muscle of the common carp, and Solexa sequencing technology was used to perform high throughput sequencing of the library. Subsequent bioinformatics analysis identified 188 conserved miRNAs and 7 novel miRNAs in the carp skeletal muscle. The miRNA expression profiling showed that, miR-1, miR-133a-3p, and miR-206 were specifically expressed in muscle-containing organs, and that miR-1, miR-21, miR-26a, miR-27a, miR-133a-3p, miR-206, miR-214 and miR-222 were differentially expressed in the process of skeletal muscle development of the common carp. This study provides a first identification and profiling of miRNAs related to the muscle biology of the common carp. Their identification could provide clues leading towards a better understanding of the molecular mechanisms of carp skeletal muscle development

Molecular adaptation of a plant-bacterium outer membrane protease towards plague virulence factor Pla

<p>Abstract</p> <p>Background</p> <p>Omptins are a family of outer membrane proteases that have spread by horizontal gene transfer in Gram-negative bacteria that infect vertebrates or plants. Despite structural similarity, the molecular functions of omptins differ in a manner that reflects the life style of their host bacteria. To simulate the molecular adaptation of omptins, we applied site-specific mutagenesis to make Epo of the plant pathogenic <it>Erwinia pyrifoliae </it>exhibit virulence-associated functions of its close homolog, the plasminogen activator Pla of <it>Yersinia pestis</it>. We addressed three virulence-associated functions exhibited by Pla, i.e., proteolytic activation of plasminogen, proteolytic degradation of serine protease inhibitors, and invasion into human cells.</p> <p>Results</p> <p>Pla and Epo expressed in <it>Escherichia coli </it>are both functional endopeptidases and cleave human serine protease inhibitors, but Epo failed to activate plasminogen and to mediate invasion into a human endothelial-like cell line. Swapping of ten amino acid residues at two surface loops of Pla and Epo introduced plasminogen activation capacity in Epo and inactivated the function in Pla. We also compared the structure of Pla and the modeled structure of Epo to analyze the structural variations that could rationalize the different proteolytic activities. Epo-expressing bacteria managed to invade human cells only after all extramembranous residues that differ between Pla and Epo and the first transmembrane β-strand had been changed.</p> <p>Conclusions</p> <p>We describe molecular adaptation of a protease from an environmental setting towards a virulence factor detrimental for humans. Our results stress the evolvability of bacterial β-barrel surface structures and the environment as a source of progenitor virulence molecules of human pathogens.</p

Current understanding of the human microbiome

Author Posting. © The Author(s), 2018. This is the author's version of the work. It is posted here by permission of Nature Publishing Group for personal use, not for redistribution. The definitive version was published in Nature Medicine 24 (2018): 392–400, doi:10.1038/nm.4517.Our understanding of the link between the human microbiome and disease, including obesity, inflammatory bowel disease, arthritis and autism, is rapidly expanding. Improvements in the throughput and accuracy of DNA sequencing of the genomes of microbial communities associated with human samples, complemented by analysis of transcriptomes, proteomes, metabolomes and immunomes, and mechanistic experiments in model systems, have vastly improved our ability to understand the structure and function of the microbiome in both diseased and healthy states. However, many challenges remain. In this Review, we focus on studies in humans to describe these challenges, and propose strategies that leverage existing knowledge to move rapidly from correlation to causation, and ultimately to translation.Many of the studies described here in our laboratories were supported by the NIH, NSF, DOE, and the Alfred P. Sloan Foundation.2018-10-1

Outer membrane protein folding from an energy landscape perspective

The cell envelope is essential for the survival of Gram-negative bacteria. This specialised membrane is densely packed with outer membrane proteins (OMPs), which perform a variety of functions. How OMPs fold into this crowded environment remains an open question. Here, we review current knowledge about OFMP folding mechanisms in vitro and discuss how the need to fold to a stable native state has shaped their folding energy landscapes. We also highlight the role of chaperones and the β-barrel assembly machinery (BAM) in assisting OMP folding in vivo and discuss proposed mechanisms by which this fascinating machinery may catalyse OMP folding

From sucrose to starch granule to starch physical behaviour: a focus on rice starch

The diversity of the physical and consequently the functional behaviour of starches, isolated from different rice varieties, is related to their specific structures. The latter are directly related to the starch biosynthetic pathway. To fully take advantage of the different functionalities of starches from different rice varieties and to design tailor-made starches, it is important to gain insight into biosynthesis-structure-physical behaviour-functionality relations. In a first part of this review, starch composition is described with a focus on rice starch. Secondly, current knowledge on starch biosynthesis is discussed. This more specifically includes (i) the function of the rice biosynthetic enzymes (i.e. adenosine diphosphate glucose pyrophosphorylases, synthases, branching and debranching enzymes), (ii) the effect of mutations on rice starch structure and, (iii) models for amylose and amylopectin synthesis. Thirdly, starch structure [i.e. from granule (2-100 p,m), to growth ring (120-500 nm), blocklet (20-500 nm), amorphous and crystalline lamellae (9 nm), and amylopectin and amylose chain levels (0.1-1.0 nm)] is dealt with. Finally, relations between rice starch structural aspects [i.e. amylopectin (core) chain length distributions] and physical behaviour (i.e. gelatinisation and amylopectin retrogradation as measured by differential scanning calorimetry) are studied. (C) 2004 Elsevier Ltd. All rights reserved.status: publishe

Isolation and characterisation of rye starch

Starch was isolated from rye flour by alkaline extraction or Pronase treatments. Both procedures led to starches of comparable purity. The present first report on the use of proteolytic enzymes in rye starch isolation showed that such treatment resulted in the highest starch yields. The rye starches had A-type granules with average particle size of 31 mum. The Pronase isolated starch had lower differential scanning calorimetry gelatinisation temperatures than the alkaline extracted starch. Defatted rye flour resulted in a Pronase isolated starch with lower peak and higher end viscosities. Wheat and rye starches isolated by the Pronase procedure were of comparable purity, but a higher yield of the former was obtained. The wheat starch had lower levels of A-type granules with an average particle size of 26 mum. When compared pairwise to the rye starches, the wheat starches had higher gelatinisation temperatures, a higher pasting temperature, a higher peak viscosity and a lower end viscosity. (C) 2003 Elsevier Ltd. All rights reserved.status: publishe

Rice starches. I. Structural aspects provide insight into crystallinity characteristics and gelatinisation behaviour of granular starch

To gain an understanding of the contribution of structural aspects to the gelatinisation behaviour of rice starch, five waxy and 10 normal rice starches, isolated from 15 milled rice samples, were analysed for structural features [absolute (AAM), free (FAM) and lipid-complexed (LAM) amylose contents, amylopectin chain length distribution and relative crystallinity] and gelatinisation behaviour as evaluated by differential scanning calorimetry. In doing so, the normal rice starches were classified according to their peak gelatinisation temperatures (T-p) as low, intermediate and high T-p rice starches. Higher AAM and FAM contents went hand in hand with decreased onset (T-o), T-p and conclusion (T-c) gelatinisation temperatures and increased gelatinisation range (T-c - T-o) of normal intermediate and high T-p starches, whereas LAM contents increased with T-o, T-p and T-c and decreased with T-c - T-o of all investigated starches. For all starches, the relative amounts of amylopectin chains with degree of polymerisation (DP) 6-9 were negatively correlated with T-o, T-p and T-c and positively with T-c-T-o. On the contrary, chains of DP 12 - 22 increased gelatinisation temperatures and decreased T-c - T-o. Furthermore, T-o, T-p and T-c of the normal rice starches increased with relative crystallinity. (C) 2003 Elsevier Science Ltd. All rights reserved.status: publishe

Rice starches. II. Structural aspects provide insight into swelling and pasting properties

The influence of starch structure (absolute (AAM), free (FAM), lipid complexed (LAM) amylose contents and amylopectin chain length distribution) on swelling behaviour (swelling power (SP), close packing concentration (C), total starch solubility (S) and amylose leaching (AML) in a temperature range between 55 and 125 degreesC) and pasting properties (6, 8 and 10% dry matter starch) of five waxy and 10 normal (low (three), intermediate (four) and high (three) peak gelatinisation temperature) rice starches was investigated. At temperatures between 55 and 85 degreesC, waxy starches had higher SP than normal starches. In this temperature range, however, SP and C* of the normal starches were independent of AAM and FAM. On the contrary, at temperatures between 95 and 125 degreesC, SP and C* of all starches decreased and increased with AAM and FAM contents, respectively. For the normal starches, SP decreased and C* increased with LAM contents at 65 degreesC. For all starches, relative amounts of short amylopectin chains with degrees of polymerisation (DP) 6-9 led to increased SP and decreased C* at 55 and 65 degreesC. Opposite results were observed for relative amounts of longer amylopectin chains with DP 12-22. AAM and FAM contents decreased with starting gel point temperatures (6, 8 and 10% dry matter starch) of normal intermediate and high peak gelatinisation temperature starches. Peak and breakdown viscosities (6% dry matter starch) decreased, whereas setback and final viscosities (10% dry matter starch) increased with AAM and FAM contents of all investigated starches. Chains of DP 6-9 and 12-22, respectively, decreased and increased starting gel point temperatures. No significant correlations between amylopectin chain length distribution and peak, breakdown, setback and final viscosities were observed. (C) 2003 Elsevier Science Ltd. All rights reserved.status: publishe