The absence of sharks from abyssal regions of the world's oceans

The oceanic abyss (depths greater than 3000 m), one of the largest environments on the planet, is characterized by absence of solar light, high pressures and remoteness from surface food supply necessitating special molecular, physiological, behavioural and ecological adaptations of organisms that live there. Sampling by trawl, baited hooks and cameras we show that the Chondrichthyes (sharks, rays and chimaeras) are absent from, or very rare in this region. Analysis of a global data set shows a trend of rapid disappearance of chondrichthyan species with depth when compared with bony fishes. Sharks, apparently well adapted to life at high pressures are conspicuous on slopes down to 2000 m including scavenging at food falls such as dead whales. We propose that they are excluded from the abyss by high-energy demand, including an oil-rich liver for buoyancy, which cannot be sustained in extreme oligotrophic conditions. Sharks are apparently confined to ca 30% of the total ocean and distribution of many species is fragmented around sea mounts, ocean ridges and ocean margins. All populations are therefore within reach of human fisheries, and there is no hidden reserve of chondrichthyan biomass or biodiversity in the deep sea. Sharks may be more vulnerable to over-exploitation than previously thought.     

A global view of protein expression in human cells,tissues, and organs

Defining the protein profiles of tissues and organs is critical to understanding the unique characteristics of the various cell types in the human body. In this study, we report on an anatomically comprehensive analysis of 4842 protein profiles in 48 human tissues and 45 human cell lines. A detailed analysis of over 2 million manually annotated, high‐resolution, immunohistochemistry‐based images showed a high fraction (>65%) of expressed proteins in most cells and tissues, with very few proteins (<2%) detected in any single cell type. Similarly, confocal microscopy in three human cell lines detected expression of more than 70% of the analyzed proteins. Despite this ubiquitous expression, hierarchical clustering analysis, based on global protein expression patterns, shows that the analyzed cells can be still subdivided into groups according to the current concepts of histology and cellular differentiation. This study suggests that tissue specificity is achieved by precise regulation of protein levels in space and time, and that different tissues in the body acquire their unique characteristics by controlling not which proteins are expressed but how much of each is produced.     

The reg4 Gene,Amplified in the Early Stages of Pancreatic Cancer Development,Is a Promising Therapeutic Target

Background

The aim of our work was to identify the genes specifically altered in pancreatic adenocarcinoma and especially those that are altered early in cancer development.

Methodology/Principal Findings

Gene copy number was systematically assessed with an ultra-high resolution CGH oligonucleotide microarray in DNA from samples of pancreatic cancer. Several new cancer-associated variations were observed. In this work we focused on one of them, involving the reg4 gene. Gene copy number gain of the reg4 gene was confirmed by qPCR in 14 cancer samples. It was also found with increased copy number in most PanIN3 samples. The relationship betweena gain in reg4 gene copy number and cancer development was investigated on the human pancreatic cancer cell line Mia-PaCa2 xenografted under the skin of nude mice. When cells were transfected with a vector allowing reg4 expression, they generated tumors almost twice larger in size. In addition, these tumors were more resistant to gemcitabine treatment than control tumors. Interestingly, weekly intraperitoneal administration of a monoclonal antibody to reg4 halved the size of tumors generated by Mia-PaCa2 cells, suggesting that the antibody interfered with a paracrine/autocrine mechanism involving reg4 and stimulating cancer progression. The addition of gemcitabine resulted in further reduction, tumors becoming 5 times smaller than control. Exposure to reg4 antibody resulted in a significant decrease in intra-tumor levels of pAkt, Bcl-xL, Bcl-2, survivin and cyclin D1.

Conclusions/Significance

It was concluded that adjuvant therapies targeting reg4 could improve the standard treatment of pancreatic cancer with gemcitabine.     

Expression of antibodies using single‐open reading frame vector design and polyprotein processing from mammalian cells

We describe a novel polyprotein precursor‐based approach to express antibodies from mammalian cells. Rather than expressing heavy and light chain proteins from separate expression units, the antibody heavy and light chains are contained in one single‐open reading frame (sORF) separated by an intein gene fused in frame. Inside mammalian cells this ORF is transcribed into a single mRNA, and translated into one polypeptide. The antibody heavy and light chains are separated posttranslationally, assembled into the functional antibody molecule, and secreted into culture medium. It is demonstrated that Pol I intein from P. horikoshii mediates protein splicing and cleavage reactions in mammalian cells, in the context of antibody heavy and light chain amino acid sequences. To allow the separation of antibody heavy chain, light chain, and the intein, we investigated a number of intein mutations designed to inhibit intein‐mediated splicing but preserve cleavage reactions. We have also designed constructs in which the signal peptide downstream from intein has altered hydrophobicity. The use of some of these mutant constructs resulted in more efficient antibody secretion, highlighting areas that can be further explored in improving such an expression system. An antibody secreted using one of the sORF constructs was characterized. This antibody has correct N‐terminal sequences for both of its heavy and light chains, correct heavy and light chain MW as well as intact MW as measured by mass spectrometry. Its affinity to antigen, as measured by surface plasmon resonance (SPR), is indistinguishable from that of the same antibody produced using conventional method. © 2009 American Institute of Chemical Engineers Biotechnol. Prog., 2009     

Characterization of Variation in Rumen Methanogenic Communities under Different Dietary and Host Feed Efficiency Conditions,as Determined by PCR-Denaturing Gradient Gel Electrophoresis Analysis

Understanding ruminal methanogens is essential for greenhouse gas mitigation, as well as for improving animal performance in the livestock industry. It has been speculated that ruminal methanogenic diversity affects host feed efficiency and results in differences in methane production. This study examined methanogenic profiles in the rumen using culture-independent PCR-denaturing gradient gel electrophoresis (PCR-DGGE) analysis for 56 beef cattle which differed in feed efficiency, as well as diet (the cattle were fed a low-energy diet or a high-energy diet). The methanogenic PCR-DGGE profiles detected were greatly affected by diet, and the major pattern changed from a community containing predominantly Methanobrevibacter ruminantium NT7 with the low-energy diet to a community containing predominantly Methanobrevibacter smithii, Methanobrevibacter sp. AbM4, and/or M. ruminantium NT7 with the high-energy diet. For each diet, the methanogenic PCR-DGGE pattern was strongly associated with the feed efficiency of the host. Diet-associated bands for Methanobrevibacter sp. AbM4 and M. smithii SM9 and a feed efficiency-related band for M. smithii PS were identified. The abundance of total methanogens was estimated by determining the numbers of copies of the 16S rRNA genes of methanogens. However, the size of the methanogen population did not correlate with differences in feed efficiency, diet, or metabolic measurements. Thus, the structure of the methanogenic community at the species or strain level may be more important for determining host feed efficiency under different dietary conditions.Ruminal methanogens use methanogenesis pathways to maintain low hydrogen partial pressure and to facilitate fiber digestion in the rumen by converting hydrogen into methane gas (24, 37). However, although it is necessary, this process also has adverse effects because the released methane represents a significant loss of dietary energy for the host animal (14) and it constitutes a large proportion of the agricultural greenhouse gas emitted (4, 10). Many studies to obtain a better understanding of rumen methanogens have been conducted in order to improve the efficiency of ruminal function and to mitigate methane release. Assessments by both cultivation-dependent and cultivation-independent methods have found that members of the genus Methanobrevibacter account for the majority of the methanogens in the rumens of sheep and cattle (11, 18, 21-23, 28, 31, 33, 34). In addition, Methanosphaera stadtmanae, Methanobacterium species, and Methanosarcina barkeri have also been found in some studies (13, 32). Although the phylogenetic positions of the methanogens in the rumen are diverse, these organisms utilize only three major pathways for methanogenesis: the CO₂ reduction pathway, the C₁ compound (e.g., methanol and methylamine) conversion pathway, and the acetate fermentation pathway. Each methanogen species has a substrate preference, and most methanogens can use only one or two substrates (37).Previous studies of rumen methanogens focused primarily on determining the methanogen species composition in different samples and developing strategies to reduce the methane yield from ruminants. Recently, there has been a strong desire to understand the impact of methanogens on host biology. Two primary studies found that feedlot beef cattle with higher feed efficiency (designated “efficient” animals) produced about 20% less methane gas than animals with lower feed efficiency (designated “inefficient” animals) (8, 19). The methanogenic communities of efficient and inefficient animals fed a low-energy diet have been compared, and divergence between the two communities has been reported (36). However, it is not clear how the methanogens in the rumen of cattle change when the animals are fed a different diet.The aims of this study were to describe the methanogenic communities in 56 steers with different feed efficiencies that were fed two distinct diets (a low-energy diet and a high-energy diet) and to understand how methanogenic communities change in response to diet modification using PCR-denaturing gradient gel electrophoresis (PCR-DGGE) and sequence analysis. Multivariate analysis was used to analyze the association of PCR-DGGE bands with the daily dry matter intake (DMI), average daily gain (ADG), feed conversion ratio (FCR), and residual feed intake (RFI). Methanogens that were associated with diet and with host feed efficiency were identified. In addition, the methanogen population of each rumen sample was examined by quantitative real-time PCR (qRT-PCR), and the results for different RFI groups and both diets were compared.     

Disentangling controls on animal abundance: Prey availability,thermal habitat,and microhabitat structure

The question of what controls animal abundance has always been fundamental to ecology, but given rapid environmental change, understanding the drivers and mechanisms governing abundance is more important than ever. Here, we determine how multidimensional environments and niches interact to determine population abundance along a tropical habitat gradient. Focusing on the endemic lizard Anolis bicaorum on the island of Utila (Honduras), we evaluate direct and indirect effects of three interacting niche axes on abundance: thermal habitat quality, structural habitat quality, and prey availability. We measured A. bicaorum abundance across a series of thirteen plots and used N‐mixture models and path analysis to disentangle direct and indirect effects of these factors. Results showed that thermal habitat quality and prey biomass both had positive direct effects on anole abundance. However, thermal habitat quality also influenced prey biomass, leading to a strong indirect effect on abundance. Thermal habitat quality was primarily a function of canopy density, measured as leaf area index (LAI). Despite having little direct effect on abundance, LAI had a strong overall effect mediated by thermal quality and prey biomass. Our results demonstrate the role of multidimensional environments and niche interactions in determining animal abundance and highlight the need to consider interactions between thermal niches and trophic interactions to understand variation in abundance, rather than focusing solely on changes in the physical environment.     

Kinetics of Regulated Actin Transitions Measured by Probes on Tropomyosin

Changes in the muscle regulatory protein complex, troponin, are important for modulation of activity and may occur as a result of disease-causing mutations. Both increases and decreases in the rate of ATP hydrolysis by myosin may occur as dictated by changes in the distribution of actin-tropomyosin-troponin among its different states. It is important to measure the rates of transition among these states to study physiological adaptation and disease processes. We show here that acrylodan or pyrene probes on tropomyosin can be used to monitor the transition from active to intermediate and inactive states of actin-tropomyosin-troponin. Transitions measured in the absence of calcium had two phases, as previously reported for some other probes on troponin and actin. The first step was a rapid equilibrium that favored the formation of the intermediate state and had an apparent rate constant less than that of S1-ATP dissociation. The second fluorescence transition was slower, with an apparent constant that increased from ∼5 to 80/s over a range of 1-37°C. Only the initial rapid transition was seen in the presence of saturating calcium. The acrylodan probe had the advantage of yielding a larger signal than the pyrene probe. Furthermore, the acrylodan signal decreased in going from the active state to the intermediate state, and then increased upon going to the inactive state.