Cleavage of syndecan-4 by ADAMTS1 provokes defects in adhesion

Syndecan-4 is a membrane-bound heparan sulfate proteoglycan that participates in cell-cell and cell-matrix interactions and modulates adhesion and migration of many cell types. Through its extracellular domain, syndecan-4 cooperates with adhesion molecules and binds matrix components relevant for cell migration. Importantly, syndecan-4 is a substrate of extracellular proteases, however the biological significance of this cleavage has not been elucidated. Here, we show that the secreted metalloprotease ADAMTS1, involved in angiogenesis and inflammatory processes, cleaves the ectodomain of syndecan-4. We further showed that this cleavage results in altered distribution of cytoskeleton components, functional loss of adhesion, and gain of migratory capacities. Using syndecan-4 null cells, we observed that ADAMTS1 proteolytic action mimics the outcome of genetic deletion of this proteoglycan with regards to focal adhesion. Our findings suggest that the shedding of syndecan-4 by ADAMTS1 disrupts cell adhesion and promotes cell migration.     

LINE-1 methylation in granulocyte DNA and trihalomethane exposure is associated with bladder cancer risk

DNA methylation changes contribute to bladder carcinogenesis. Trihalomethanes (THM), a class of disinfection by-products, are associated with increased urothelial bladder cancer (UBC) risk. THM exposure in animal models produces DNA hypomethylation. We evaluated the relationship of LINE-1 5-methylcytosine levels (LINE-1%5mC) as outcome of long-term THM exposure among controls and as an effect modifier in the association between THM exposure and UBC risk. We used a case-control study of UBC conducted in Spain. We obtained personal lifetime residential THM levels and measured LINE-1%5mC by pyrosequencing in granulocyte DNA from blood samples in 548 incident cases and 559 hospital controls. Two LINE-1%5mC clusters (above and below 64%) were identified through unsupervised hierarchical cluster analysis. The association between THM levels and LINE-1%5mC was evaluated with β regression analyses and logistic regression was used to estimate odds ratios (OR) adjusting for covariables. LINE-1%5mC change between percentiles 75^th and 25^th of THM levels was 1.8% (95% confidence interval (CI): 0.1, 3.4%) among controls. THM levels above vs. below the median (26 μg/L) were associated with increased UBC risk, OR = 1.86 (95% CI: 1.25, 2.75), overall and among subjects with low levels of LINE-1%5mC (n = 975), OR = 2.14 (95% CI: 1.39, 3.30), but not associated with UBC risk among subjects’ high levels of LINE-1%5mC (n = 162), interaction P = 0.03. Results suggest a positive association between LINE-1%5mC and THM levels among controls, and LINE-1%5mC status may modify the association between UBC risk and THM exposure. Because reverse causation and chance cannot be ruled out, confirmation studies are warranted.     

Traditional Banana Diversity in Oceania: An Endangered Heritage

This study aims to understand the genetic diversity of traditional Oceanian starchy bananas in order to propose an efficient conservation strategy for these endangered varieties. SSR and DArT molecular markers are used to characterize a large sample of Pacific accessions, from New Guinea to Tahiti and Hawaii. All Pacific starchy bananas are shown of New Guinea origin, by interspecific hybridization between Musa acuminata (AA genome), more precisely its local subspecies M. acuminata ssp. banksii, and M. balbisiana (BB genome) generating triploid AAB Pacific starchy bananas. These AAB genotypes do not form a subgroup sensu stricto and genetic markers differentiate two subgroups across the three morphotypes usually identified: Iholena versus Popoulu and Maoli. The Popoulu/Maoli accessions, even if morphologically diverse throughout the Pacific, cluster in the same genetic subgroup. However, the subgroup is not strictly monophyletic and several close, but different genotypes are linked to the dominant genotype. One of the related genotypes is specific to New Caledonia (NC), with morphotypes close to Maoli, but with some primitive characters. It is concluded that the diffusion of Pacific starchy AAB bananas results from a series of introductions of triploids originating in New Guinea area from several sexual recombination events implying different genotypes of M. acuminata ssp. banksii. This scheme of multiple waves from the New Guinea zone is consistent with the archaeological data for peopling of the Pacific. The present geographic distribution suggests that a greater diversity must have existed in the past. Its erosion finds parallels with the erosion of cultural traditions, inexorably declining in most of the Polynesian or Melanesian Islands. Symmetrically, diversity hot spots appear linked to the local persistence of traditions: Maoli in New Caledonian Kanak traditions or Iholena in a few Polynesian islands. These results will contribute to optimizing the conservation strategy for the ex-situ Pacific Banana Collection supported collectively by the Pacific countries.     

WAY-100635 antagonist-induced plasticity of 5-HT receptors: regulatory differences between a stable cell line and an in vivo native system

We present evidence that the 5-hydroxytryptamine(1A) (5-HT(1A)) receptor antagonist, N-{2-[4-(2-methoxyphenyl)-1-piperazinyl]-ethyl}-N-(2-pyridinyl)cyclohexanecarboxamide (WAY-100635), can induce receptor internalization in a human (h)5-HT(1A) receptor Chinese hamster ovary (CHO-K1) cell system. Exposure of h5-HT(1A) CHO cells to WAY-100635 decreased the cell-surface h5-HT(1A) receptor density in a way that was both time (24-72 h) and concentration (1-100 nm) dependent.[(3)H]WAY-100635 and [(3)H]8-hydroxy-dipropylaminotetralin ([(3)H]8-OH-DPAT) saturation analyses demonstrated a significant reduction (50-60%) in total h5-HT(1A) receptor number in the WAY-100635-treated (100 nm; 72 h) compared with control cells. In WAY-100635-treated cells, the 8-OH-DPAT-mediated inhibition of forskolin (FSK)-stimulated cAMP accumulation was right-shifted and the maximal inhibitory response of 8-OH-DPAT was impaired compared with control cells. Similar results were obtained for 8-OH-DPAT-mediated Ca(2+) mobilization after WAY-100635 treatment. h5-HT(1A) receptors labeled with [(3)H]WAY-100635, as well as [(3)H]4-(2'-Methoxy)-phenyl-1-[2'-(N-2'-pyridinyl)-p-fluorobenzamido]ethyl-piperazine (MPPF), exhibited a time-dependent rate of cellular internalization that was blocked by endocytotic suppressors and was pertussis-toxin insensitive. In contrast, quantitative autoradiographic studies demonstrated that chronic treatment of rats with WAY-100635 for two weeks produced a region-specific increase in the 5-HT(1A) receptor density. In conclusion, prolonged exposure of an h5-HT(1A) cell-based system to the 5-HT(1A) antagonist, WAY-100635, induced a paradoxical internalization of cell surface receptor resulting in depressed functional activity. This suggests that an antagonist can influence 5-HT(1A) receptor recycling in vitro differently to in vivo regulatory conditions.     

The most frequent short sequences in non-coding DNA

The purpose of this work is to determine the most frequent short sequences in non-coding DNA. They may play a role in maintaining the structure and function of eukaryotic chromosomes. We present a simple method for the detection and analysis of such sequences in several genomes, including Arabidopsis thaliana, Caenorhabditis elegans, Drosophila melanogaster and Homo sapiens. We also study two chromosomes of man and mouse with a length similar to the whole genomes of the other species. We provide a list of the most common sequences of 9–14 bases in each genome. As expected, they are present in human Alu sequences. Our programs may also give a graph and a list of their position in the genome. Detection of clusters is also possible. In most cases, these sequences contain few alternating regions. Their intrinsic structure and their influence on nucleosome formation are not known. In particular, we have found new features of short sequences in C. elegans, which are distributed in heterogeneous clusters. They appear as punctuation marks in the chromosomes. Such clusters are not found in either A. thaliana or D. melanogaster. We discuss the possibility that they play a role in centromere function and homolog recognition in meiosis.     

Interaction study between HCV NS5A-D2 and NS5B using <Superscript>19</Superscript>F NMR

The non structural protein 5A (NS5A) regulates the replication of the hepatitis C viral RNA through a direct molecular interaction of its domain 2 (NS5A-D2) with the RNA dependent RNA polymerase NS5B. Because of conflicting data in the literature, we study here this molecular interaction using fluorinated versions of the NS5A-D2 protein derived from the JFH1 Hepatitis C Virus strain. Two methods to prepare fluorine-labelled NS5A-D2 involving the biosynthetic incorporation of a ¹⁹F-tryptophan using 5-fluoroindole and the posttranslational introduction of fluorine by chemical conjugation of 2-iodo-N-(trifluoromethyl)acetamide with the NS5A-D2 cysteine side chains are presented. The dissociation constants (K_D) between NS5A-D2 and NS5B obtained with these two methods are in good agreement, and yield values comparable to those derived previously from a surface plasmon resonance study. We compare benefits and limitations of both labeling methods to study the interaction between an intrinsically disordered protein and a large molecular target by ¹⁹F NMR.     

SEASONAL VARIATIONS OF PHOTOSYNTHETIC PIGMENTS, TOTAL C, N, AND P CONTENT, AND PHOTOSYNTHESIS IN PHYLLARIOPSIS PURPURASCENS: (PHAEOPHYTA) FROM THE STRAIT OF GIBRALTAR

Photosynthetic pigments, C, N, and P tissue composition, and photosynthetic rate were measured from April to October in the brown alga Phyllariopsis purpurascens (C. Agardh) Henry et South (Laminariales, Phaeophyta) growing at a 30-m depth in the Strait of Gibraltar. Ir-radiance reaching the population ranged from 13.5 to 27.5 mol.m^-2.mo^-1. The available light for this species, expressed as a percentage of the irradiance above the water, was 1.8%. Dissolved inorganic nitrogen forms, NO3-and NH₄+, were constant from April to October, whereas phosphate was depleted in August. Chlorophyll a decreased from 520.0 ± 165.0 to 199.6 ± 159.9 μg.g^-1 dry weight; in contrast, chlorophyll c and carotenoids did not change until September but increased threefold in October. C:N and N:P ratios changed in the same way and in the same range. They were constant until July but increased from 15–17 up to 42 (C:N) and from 14 to 40 (N:P) in October, suggesting a severe P limitation of growth of this species. The dark respiration rate and the light compensation point were constant from April to October (0.5 ± 0.1 μmol O₂. m^-2.s^-1 and 6.5 ± 0.2 μmol.m^-2. s^-1, respectively), whereas the maximum rate of apparent photosynthesis, light onset saturation parameter, and half saturation constant for light were maximum in April to May (3.7 μmol O₂. m^-2.s^-1and 40 and 41.5 μmol.m^-2. s^-1, respectively) and October (3.6 μmol O₂. m^-2.s^-1 and 50 and 53.7 μmol.m^-2. s^-1, respectively). They were minimum in August (1.2 μmol O₂.m^-2.s^-1 and 11.3 and 12 μmol.m^-2.s^-1, respectively). These minimum figures yielded a negative carbon budget in August and 0 in September, whereas it was positive the rest of the year. Photosynthetic efficiency, estimated by the ratio between maximum apparent photosynthesis and light half saturation constant, showed a strong agreement with productivity measured by means of an independent method. These results indicate that lamina expansion in this species is controlled by photosynthetic efficiency.     

Meningococcal disease: A paradigm of type‐IV pilus dependent pathogenesis

Neisseria meningitidis (meningococcus) is a Gram‐negative bacterium responsible for two devastating forms of invasive diseases: purpura fulminans and meningitis. Interaction with both peripheral and cerebral microvascular endothelial cells is at the heart of meningococcal pathogenesis. During the last two decades, an essential role for meningococcal type IV pili in vascular colonisation and disease progression has been unravelled. This review summarises 20 years of research on meningococcal type IV pilus‐dependent virulence mechanisms, up to the identification of promising anti‐virulence compounds that target type IV pili.     

Hypothalamic S-Nitrosylation Contributes to the Counter-Regulatory Response Impairment following Recurrent Hypoglycemia

Aims

Hypoglycemia is a severe side effect of intensive insulin therapy. Recurrent hypoglycemia (RH) impairs the counter-regulatory response (CRR) which restores euglycemia. During hypoglycemia, ventromedial hypothalamus (VMH) production of nitric oxide (NO) and activation of its receptor soluble guanylyl cyclase (sGC) are critical for the CRR. Hypoglycemia also increases brain reactive oxygen species (ROS) production. NO production in the presence of ROS causes protein S-nitrosylation. S-nitrosylation of sGC impairs its function and induces desensitization to NO. We hypothesized that during hypoglycemia, the interaction between NO and ROS increases VMH sGC S-nitrosylation levels and impairs the CRR to subsequent episodes of hypoglycemia. VMH ROS production and S-nitrosylation were quantified following three consecutive daily episodes of insulin-hypoglycemia (RH model). The CRR was evaluated in rats in response to acute insulin-induced hypoglycemia or via hypoglycemic-hyperinsulinemic clamps. Pretreatment with the anti-oxidant N-acetyl-cysteine (NAC) was used to prevent increased VMH S-nitrosylation.

Results

Acute insulin-hypoglycemia increased VMH ROS levels by 49±6.3%. RH increased VMH sGC S-nitrosylation. Increasing VMH S-nitrosylation with intracerebroventricular injection of the nitrosylating agent S-nitroso-L-cysteine (CSNO) was associated with decreased glucagon secretion during hypoglycemic clamp. Finally, in RH rats pre-treated with NAC (0.5% in drinking water for 9 days) hypoglycemia-induced VMH ROS production was prevented and glucagon and epinephrine production was not blunted in response to subsequent insulin-hypoglycemia.

Conclusion

These data suggest that NAC may be clinically useful in preventing impaired CRR in patients undergoing intensive-insulin therapy.