Antigens secreted from Mycobacterium tuberculosis: identification by proteomics approach and test for diagnostic marker

Tuberculosis caused by mycobacteria, mainly Mycobacterium tuberculosis, is a major infectious disease of the respiratory system. An early diagnosis followed by chemotherapy is the major control strategy. In an effort to identify the antigens suitable for immunodiagnosis and vaccines, the proteins secreted in a culture medium from the M. tuberculosis K-strain, which is the most prevalent among the clinical isolates in Korea and belongs to the Beijing family, were analyzed by two-dimensional polyacrylamide gel electrophoresis (2-D PAGE) and compared with those from the M. tuberculosis H37Rv and CDC1551 strains. Eight proteins, Rv0652, Rv1636, Rv2818c, Rv3369, Rv3865, Rv0566c, MT3304, and Rv3160, were identified by matrix-assisted laser desorption/ionization-time of flight-mass spectrometry (MALDI-TOF-MS) or liquid chromatography-electrospray ionization-mass spectrometry (LC-ESI-MS) and found to be relatively abundant in the culture medium from the M. tuberculosis K-strain but less so from the CDC1551 or H37Rv strains. In addition, Rv3874 (CFP-10), Rv-0560c and Rv3648c, which were expressed increasingly in the K and CDC1551 strains, were also identified using the same proteomics technology. All proteins were prepared by molecular cloning, expression in Escherichia coli followed by affinity purification. Among them, three proteins, rRv3369, rRv0566c, and rRv3874, were selected by prescreening and examined for their potential as serodiagnostic antigens using an enzyme-linked immunosorbent assay. When 100 sera from tuberculosis patients and 100 sera from the healthy controls were analyzed, rRV3369, rRv3874, and rRv0566c showed a sensitivity of 60%, 74%, and 43%, and a specificity of 96%, 97%, and 84%, respectively. These results suggest that the rRv3369 and rRv3874 proteins, which were expressed more abundantly in the more recently obtained clinical isolates of M. tuberculosis than in the laboratory-adapted H37Rv strain, are promising for use in the serodiagnosis of tuberculosis.     

Nutrient uptake rates by the alien alga Undaria pinnatifida (Phaeophyta) (Nuevo Gulf, Patagonia, Argentina) when exposed to diluted sewage effluent

In the early nineties, Undaria pinnatifida has been accidentally introduced to Nuevo Gulf (Patagonia, Argentina) where the environmental conditions would have favored its expansion. The effect of the secondary treated sewage discharge from Puerto Madryn city into Nueva Bay (located in the western extreme of Nuevo Gulf) is one of the probable factors to be taken into account. Laboratory cultures of this macroalgae were conducted in seawater enriched with the effluent. The nutrients (ammonium, nitrate and phosphate) uptake kinetics was studied at constant temperature and radiation (16?°C and 50 μE m^?2 s^?1 respectively). Uptake kinetics of both inorganic forms of nitrogen were described by the Michaelis–Menten model during the surge phase (ammonium: V _max ^sur: 218.1 μmol h^?1 g^?1, K _s ^sur: 476.5 μM and nitrate V _max ^sur: 10.7 μmol h^?1 g^?1, K _s ^sur: 6.1 μM) and during the assimilation phase (ammonium: V _max ^ass: 135.6 μmol h^?1 g^?1, K _s ^ass: 407.2 μM and nitrate V _max ^ass: 1.9 μmol h^?1 g^?1, K _s ^ass: 2.2 μM), with ammonium rates always higher than those of nitrate. Even though a net phosphate disappearance was observed in all treatments, uptake kinetics of this ion could not be properly estimated by the employed methodology.     

PARP1–TDP1 coupling for the repair of topoisomerase I–induced DNA damage

Poly(ADP-ribose) polymerases (PARP) attach poly(ADP-ribose) (PAR) chains to various proteins including themselves and chromatin. Topoisomerase I (Top1) regulates DNA supercoiling and is the target of camptothecin and indenoisoquinoline anticancer drugs, as it forms Top1 cleavage complexes (Top1cc) that are trapped by the drugs. Endogenous and carcinogenic DNA lesions can also trap Top1cc. Tyrosyl-DNA phosphodiesterase 1 (TDP1), a key repair enzyme for trapped Top1cc, hydrolyzes the phosphodiester bond between the DNA 3′-end and the Top1 tyrosyl moiety. Alternative repair pathways for Top1cc involve endonuclease cleavage. However, it is unknown what determines the choice between TDP1 and the endonuclease repair pathways. Here we show that PARP1 plays a critical role in this process. By generating TDP1 and PARP1 double-knockout lymphoma chicken DT40 cells, we demonstrate that TDP1 and PARP1 are epistatic for the repair of Top1cc. The N-terminal domain of TDP1 directly binds the C-terminal domain of PARP1, and TDP1 is PARylated by PARP1. PARylation stabilizes TDP1 together with SUMOylation of TDP1. TDP1 PARylation enhances its recruitment to DNA damage sites without interfering with TDP1 catalytic activity. TDP1–PARP1 complexes, in turn recruit X-ray repair cross-complementing protein 1 (XRCC1). This work identifies PARP1 as a key component driving the repair of trapped Top1cc by TDP1.     

An <Emphasis Type="Italic">in vivo</Emphasis> RNA interference screen identifies gene networks controlling <Emphasis Type="Italic">Drosophila melanogaster</Emphasis>blood cell homeostasis

Background  

In metazoans, the hematopoietic system plays a key role both in normal development and in defense of the organism. In Drosophila, the cellular immune response involves three types of blood cells: plasmatocytes, crystal cells and lamellocytes. This last cell type is barely present in healthy larvae, but its production is strongly induced upon wasp parasitization or in mutant contexts affecting larval blood cell homeostasis. Notably, several zygotic mutations leading to melanotic mass (or "tumor") formation in larvae have been associated to the deregulated differentiation of lamellocytes. To gain further insights into the gene regulatory network and the mechanisms controlling larval blood cell homeostasis, we conducted a tissue-specific loss of function screen using hemocyte-specific Gal4 drivers and UAS-dsRNA transgenic lines.     

Increasing Crop Diversity Mitigates Weather Variations and Improves Yield Stability

Cropping sequence diversification provides a systems approach to reduce yield variations and improve resilience to multiple environmental stresses. Yield advantages of more diverse crop rotations and their synergistic effects with reduced tillage are well documented, but few studies have quantified the impact of these management practices on yields and their stability when soil moisture is limiting or in excess. Using yield and weather data obtained from a 31-year long term rotation and tillage trial in Ontario, we tested whether crop rotation diversity is associated with greater yield stability when abnormal weather conditions occur. We used parametric and non-parametric approaches to quantify the impact of rotation diversity (monocrop, 2-crops, 3-crops without or with one or two legume cover crops) and tillage (conventional or reduced tillage) on yield probabilities and the benefits of crop diversity under different soil moisture and temperature scenarios. Although the magnitude of rotation benefits varied with crops, weather patterns and tillage, yield stability significantly increased when corn and soybean were integrated into more diverse rotations. Introducing small grains into short corn-soybean rotation was enough to provide substantial benefits on long-term soybean yields and their stability while the effects on corn were mostly associated with the temporal niche provided by small grains for underseeded red clover or alfalfa. Crop diversification strategies increased the probability of harnessing favorable growing conditions while decreasing the risk of crop failure. In hot and dry years, diversification of corn-soybean rotations and reduced tillage increased yield by 7% and 22% for corn and soybean respectively. Given the additional advantages associated with cropping system diversification, such a strategy provides a more comprehensive approach to lowering yield variability and improving the resilience of cropping systems to multiple environmental stresses. This could help to sustain future yield levels in challenging production environments.     

The expression of FTO in human adipose tissue is influenced by fat depot,adiposity, and insulin sensitivity

Objective: The fat mass and obesity associated (FTO) gene is related to obesity, but the regulation of FTO expression in adipose tissue is not fully understood. We investigated FTO expression in paired subcutaneous and omental adipose tissues (SAT and OAT) from healthy women undergoing gynecological surgeries, and its relation with adiposity and insulin sensitivity. Design and Methods: FTO expression in SAT of type 2 diabetic patients treated or not with Rosiglitazone was also compared. Results: Both the mRNA and protein levels of FTO were higher in OAT from women than in SAT. Only OAT FTO protein levels negatively correlated with BMI and body fat mass, whereas SAT FTO mRNA levels were negatively correlated with subcutaneous fat deposition. In addition, SAT FTO mRNA and protein levels were increased in insulin resistant women (high HOMA) compared to insulin sensitive women (low HOMA), whereas OAT FTO expression was not different between these two subgroups. Interestingly, FTO mRNA levels were increased in SAT of type 2 diabetic patients, and treatment of diabetics with Rosiglitazone improved insulin sensitivity and reduced SAT FTO mRNA levels. Lastly, FTO expression was transiently increased in the early phase of 3T3‐L1 cell differentiation, which coincides with the induction of PPARγ2 expression. However, partial reduction of FTO did not impact PPARγ2 expression and adipocyte differentiation. Conclusion: Therefore, FTO gene expression is higher in OAT than in SAT in lean to moderately obese women. OAT FTO expression is associated with adiposity, whereas SAT FTO expression is associated with insulin sensitivity. These associations are independent of an effect of FTO on adipocyte differentiation.