Does seawater acidification affect survival, growth and shell integrity in bivalve juveniles?

Anthropogenic emissions of carbon dioxide are leading to decreases in pH and changes in the carbonate chemistry of seawater. Ocean acidification may negatively affect the ability of marine organisms to produce calcareous structures while also influencing their physiological responses and growth. The aim of this study was to evaluate the effects of reduced pH on the survival, growth and shell integrity of juveniles of two marine bivalves from the Northern Adriatic sea: the Mediterranean mussel Mytilus galloprovincialis and the striped venus clam Chamelea gallina. An outdoor flow-through plant was set up and two pH levels (natural seawater pH as a control, pH 7.4 as the treatment) were tested in long-term experiments. Mortality was low throughout the first experiment for both mussels and clams, but a significant increase, which was sensibly higher in clams, was observed at the end of the experiment (6 months). Significant decreases in the live weight (-26%) and, surprisingly, in the shell length (-5%) were observed in treated clams, but not in mussels. In the controls of both species, no shell damage was ever recorded; in the treated mussels and clams, damage proceeded via different modes and to different extents. The severity of shell injuries was maximal in the mussels after just 3 months of exposure to a reduced pH, whereas it progressively increased in clams until the end of the experiment. In shells of both species, the damaged area increased throughout the experiment, peaking at 35% in mussels and 11% in clams. The shell thickness of the treated and control animals significantly decreased after 3 months in clams and after 6 months in mussels. In the second experiment (3 months), only juvenile mussels were exposed to a reduced pH. After 3 months, the mussels at a natural pH level or pH 7.4 did not differ in their survival, shell length or live weight. Conversely, shell damage was clearly visible in the treated mussels from the 1st month onward. Monitoring the chemistry of seawater carbonates always showed aragonite undersaturation at 7.4 pH, whereas calcite undersaturation occurred in only 37% of the measurements. The present study highlighted the contrasting effects of acidification in two bivalve species living in the same region, although not exactly in the same habitat.

MEG networks organization is related to white matter integrity

Many studies have assessed the characterization of anatomical or functional connectivity in mild cognitive impairment (MCI), however it is still unknown how they are related in the course of the pathology. Here we integrate the analysis of magnetoencephalographic (MEG) data with white matter (WM) integrity quantification from diffusion weighted imaging (DWI), to asses whether the damage in the WM tracts disrupt the organization of the functional networks.

Polycystin 1 is required for the structural integrity of blood vessels

Autosomal dominant polycystic kidney disease (ADPKD), often caused by mutations in the PKD1 gene, is associated with life-threatening vascular abnormalities that are commonly attributed to the frequent occurrence of hypertension. A previously reported targeted mutation of the mouse homologue of PKD1 was not associated with vascular fragility, leading to the suggestion that the vascular lesion may be of a secondary nature. Here we demonstrate a primary role of PKD1 mutations in vascular fragility. Mouse embryos homozygous for the mutant allele (Pkd1L) exhibit s.c. edema, vascular leaks, and rupture of blood vessels, culminating in embryonic lethality at embryonic day 15.5. Kidney and pancreatic ductal cysts are present. The Pkd1-encoded protein, mouse polycystin 1, was detected in normal endothelium and the surrounding vascular smooth muscle cells. These data reveal a requisite role for polycystin 1 in maintaining the structural integrity of the vasculature as well as epithelium and suggest that the nature of the PKD1 mutation contributes to the phenotypic variance in ADPKD.

Complete Cytolysis and Neonatal Lethality in Keratin 5 Knockout Mice Reveal Its Fundamental Role in Skin Integrity and in Epidermolysis Bullosa Simplex

In human patients, a wide range of mutations in keratin (K) 5 or K14 lead to the blistering skin disorder epidermolysis bullosa simplex. Given that K14 deficiency does not lead to the ablation of a basal cell cytoskeleton because of a compensatory role of K15, we have investigated the requirement for the keratin cytoskeleton in basal cells by inactivating the K5 gene in mice. We report that the K5−/− mice die shortly after birth, lack keratin filaments in the basal epidermis, and are more severely affected than K14−/− mice. In contrast to the K14−/− mice, we detected a strong induction of the wound-healing keratin K6 in the suprabasal epidermis of cytolyzed areas of postnatal K5−/− mice. In addition, K5 and K14 mice differed with respect to tongue lesions. Moreover, we show that in the absence of K5 and other type II keratins, residual K14 and K15 aggregated along hemidesmosomes, demonstrating that individual keratins without a partner are stable in vivo. Our data indicate that K5 may be the natural partner of K15 and K17. We suggest that K5 null mutations may be lethal in human epidermolysis bullosa simplex patients.

A statistical evaluation of the effectiveness of FMVSS 301: Fuel System Integrity. Final report.

National Highway Traffic Safety Administration, Washington, D.C.

RSV test monitoring and data publication - Renault R20TS-to-Calspan RSV 75 degree right side impact at Renault. Final report.

National Highway Traffic Safety Administration, Washington, D.C.

RSV test monitoring and data publication - Renault R20TS-to-Calspan RSV 75 degree left side impact at Peugeot. Final report.

National Highway Traffic Safety Administration, Washington, D.C.

Safety compliance test program: FMVSS 212, 219, 301-75, NCAP, Accident Investigation Division data. Final report.

National Highway Traffic Safety Administration, Office of Research and Development, Washington, D.C.

Data base for light-weight automotive power plants - Volume I: executive summary. Final report.

National Highway Traffic Safety Administration, Office of Passenger Vehicle Research, Washington, D.C.

Data base for light-weight automotive diesel power plants - Volume II: discussion and results. Final report.

National Highway Traffic Safety Administration, Office of Passenger Vehicle Research, Washington, D.C.

Data base for light-weight automotive diesel power plants - Volume III: miscellaneous data. Final report.

National Highway Traffic Safety Administration, Office of Passenger Vehicle Research, Washington, D.C.

The subcellular fractionation properties and function of insulin receptor substrate-1 (IRS-1) are independent of cytoskeletal integrity

Efficient insulin action requires spatial and temporal coordination of signaling cascades. The prototypical insulin receptor substrate, IRS-1 plays a central role in insulin signaling. By subcellular fractionation IRS-1 is enriched in a particulate fraction, termed the high speed pellet (HSP), and its redistribution from this fraction is associated with signal attenuation and insulin resistance. Anecdotal evidence suggests the cytoskeleton may underpin the localization of IRS-1 to the HSP. In the present study we have taken a systematic approach to examine whether the cytoskeleton contributes to the subcellular fractionation properties and function of IRS-1. By standard microscopy or immunoprecipitation we were unable to detect evidence to support a specific interaction between IRS-1 and the major cytoskeletal components actin (microfilaments), vimentin (intermediate filaments), and tubulin (microtubules) in 3T3-L1 adipocytes or in CHO.IR.IRS-1 cells. Pharmacological disruption of microfilaments and microtubules, individually or in combination, was without effect on the subcellular distribution of IRS-1 or insulin-stimulated tyrosine phosphorylation in either cell type. Phosphorylation of Akt was modestly reduced (20-35%) in 3T3-L1 adipocytes but not in CHO.IR.IRS-1 cells. In cells lacking intermediate filaments (Vim(-/-)) IRS-1 expression, distribution and insulin-stimulated phosphorylation appeared normal. Even after depolymerisation of microfilaments and microtubules, insulin-stimulated phosphorylation of IRS-1 and Akt were maintained in Vim-/- cells. Taken together these data indicate that the characteristic subcellular fractionation properties and function of IRS-1 are unlikely to be mediated by cytoskeletal networks and that proximal insulin signaling does not require an intact cytoskeleton. (c) 2006 Elsevier Ltd. All rights reserved.

Integrity-based kernel malware detection

Kernel-level malware is one of the most dangerous threats to the security of users on the Internet, so there is an urgent need for its detection. The most popular detection approach is misuse-based detection. However, it cannot catch up with today's advanced malware that increasingly apply polymorphism and obfuscation. In this thesis, we present our integrity-based detection for kernel-level malware, which does not rely on the specific features of malware. ^ We have developed an integrity analysis system that can derive and monitor integrity properties for commodity operating systems kernels. In our system, we focus on two classes of integrity properties: data invariants and integrity of Kernel Queue (KQ) requests. ^ We adopt static analysis for data invariant detection and overcome several technical challenges: field-sensitivity, array-sensitivity, and pointer analysis. We identify data invariants that are critical to system runtime integrity from Linux kernel 2.4.32 and Windows Research Kernel (WRK) with very low false positive rate and very low false negative rate. We then develop an Invariant Monitor to guard these data invariants against real-world malware. In our experiment, we are able to use Invariant Monitor to detect ten real-world Linux rootkits and nine real-world Windows malware and one synthetic Windows malware. ^ We leverage static and dynamic analysis of kernel and device drivers to learn the legitimate KQ requests. Based on the learned KQ requests, we build KQguard to protect KQs. At runtime, KQguard rejects all the unknown KQ requests that cannot be validated. We apply KQguard on WRK and Linux kernel, and extensive experimental evaluation shows that KQguard is efficient (up to 5.6% overhead) and effective (capable of achieving zero false positives against representative benign workloads after appropriate training and very low false negatives against 125 real-world malware and nine synthetic attacks). ^ In our system, Invariant Monitor and KQguard cooperate together to protect data invariants and KQs in the target kernel. By monitoring these integrity properties, we can detect malware by its violation of these integrity properties during execution.^

Data compilation on the biological response to ocean acidification: environmental and experimental context of data sets and related literature

The exponential growth of studies on the biological response to ocean acidification over the last few decades has generated a large amount of data. To facilitate data comparison, a data compilation hosted at the data publisher PANGAEA was initiated in 2008 and is updated on a regular basis (doi:10.1594/PANGAEA.149999). By January 2015, a total of 581 data sets (over 4 000 000 data points) from 539 papers had been archived. Here we present the developments of this data compilation five years since its first description by Nisumaa et al. (2010). Most of study sites from which data archived are still in the Northern Hemisphere and the number of archived data from studies from the Southern Hemisphere and polar oceans are still relatively low. Data from 60 studies that investigated the response of a mix of organisms or natural communities were all added after 2010, indicating a welcomed shift from the study of individual organisms to communities and ecosystems. The initial imbalance of considerably more data archived on calcification and primary production than on other processes has improved. There is also a clear tendency towards more data archived from multifactorial studies after 2010. For easier and more effective access to ocean acidification data, the ocean acidification community is strongly encouraged to contribute to the data archiving effort, and help develop standard vocabularies describing the variables and define best practices for archiving ocean acidification data.

Defining the Role of the Histone Methyltransferase, PR-Set7, in Maintaining the Genome Integrity of Drosophila Melanogaster

The complete and faithful duplication of the genome is essential to ensure normal cell division and organismal development. Eukaryotic DNA replication is initiated at multiple sites termed origins of replication that are activated at different time through S phase. The replication timing program is regulated by the S-phase checkpoint, which signals and repairs replicative stress. Eukaryotic DNA is packaged with histones into chromatin, thus DNA-templated processes including replication are modulated by the local chromatin environment such as post-translational modifications (PTMs) of histones.

One such epigenetic mark, methylation of lysine 20 on histone H4 (H4K20), has been linked to chromatin compaction, transcription, DNA repair and DNA replication. H4K20 can be mono-, di- and tri-methylated. Monomethylation of H4K20 (H4K20me1) is mediated by the cell cycle-regulated histone methyltransferase PR-Set7 and subsequent di-/tri- methylation is catalyzed by Suv4-20. Prior studies have shown that PR-Set7 depletion in mammalian cells results in defective S phase progression and the accumulation of DNA damage, which may be partially attributed to defects in origin selection and activation. Meanwhile, overexpression of mammalian PR-Set7 recruits components of pre-Replication Complex (pre-RC) onto chromatin and licenses replication origins for re-replication. However, these studies were limited to only a handful of mammalian origins, and it remains unclear how PR-Set7 impacts the replication program on a genomic scale. Finally, the methylation substrates of PR-Set7 include both histone (H4K20) and non-histone targets, therefore it is necessary to directly test the role of H4K20 methylation in PR-Set7 regulated phenotypes.

I employed genetic, cytological, and genomic approaches to better understand the role of H4K20 methylation in regulating DNA replication and genome stability in Drosophila melanogaster cells. Depletion of Drosophila PR-Set7 by RNAi in cultured Kc167 cells led to an ATR-dependent cell cycle arrest with near 4N DNA content and the accumulation of DNA damage, indicating a defect in completing S phase. The cells were arrested at the second S phase following PR-Set7 downregulation, suggesting that it was an epigenetic effect that coupled to the dilution of histone modification over multiple cell cycles. To directly test the role of H4K20 methylation in regulating genome integrity, I collaborated with the Duronio Lab and observed spontaneous DNA damage on the imaginal wing discs of third instar mutant larvae that had an alanine substitution on H4K20 (H4K20A) thus unable to be methylated, confirming that H4K20 is a bona fide target of PR-Set7 in maintaining genome integrity.

One possible source of DNA damage due to loss of PR-Set7 is reduced origin activity. I used BrdU-seq to profile the genome-wide origin activation pattern. However, I found that deregulation of H4K20 methylation states by manipulating the H4K20 methyltransferases PR-Set7 and Suv4-20 had no impact on origin activation throughout the genome. I then mapped the genomic distribution of DNA damage upon PR-Set7 depletion. Surprisingly, ChIP-seq of the DNA damage marker γ-H2A.v located the DNA damage to late replicating euchromatic regions of the Drosophila genome, and the strength of γ-H2A.v signal was uniformly distributed and spanned the entire late replication domain, implying stochastic replication fork collapse within late replicating regions. Together these data suggest that PR-Set7-mediated monomethylation of H4K20 is critical for maintaining the genomic integrity of late replicating domains, presumably via stabilization of late replicating forks.

In addition to investigating the function of H4K20me, I also used immunofluorescence to characterize the cell cycle regulated chromatin loading of Mcm2-7 complex, the DNA helicase that licenses replication origins, using H4K20me1 level as a proxy for cell cycle stages. In parallel with chromatin spindown data by Powell et al. (Powell et al. 2015), we showed a continuous loading of Mcm2-7 during G1 and a progressive removal from chromatin through S phase.