Solar ultraviolet radiation-induced DNA damage in aquatic organisms: potential environmental impact

Continuing depletion of stratospheric ozone and subsequent increases in deleterious ultraviolet (UV) radiation at the Earth's surface have fueled the interest in its ecological consequences for aquatic ecosystems. The DNA is certainly one of the key targets for UV-induced damage in a variety of aquatic organisms. UV radiation induces two of the most abundant mutagenic and cytotoxic DNA lesions, cyclobutane pyrimidine dimers (CPDs) and pyrimidine pyrimidone photoproducts (6-4PPs) and their Dewar valence isomers. However, aquatic organisms have developed a number of repair and tolerance mechanisms to counteract the damaging effects of UV on DNA. Photoreactivation with the help of the enzyme photolyase is one of the most important and frequently occurring repair mechanisms in a variety of organisms. Excision repair, which can be distinguished into base excision repair (BER) and nucleotide excision repair (NER), also play an important role in DNA repair in several organisms with the help of a number of glycosylases and polymerases, respectively. In addition, mechanisms such as mutagenic repair or dimer bypass, recombinational repair, cell-cycle checkpoints, apoptosis and certain alternative repair pathways are also operative in various organisms. This review deals with the UV-induced DNA damage and repair in a number of aquatic organisms as well as methods of detecting DNA damage.     

Energy Transducing Roles of Antiporter-like Subunits in Escherichia coli NDH-1 with Main Focus on Subunit NuoN (ND2)

The proton-translocating NADH-quinone oxidoreductase (complex I/NDH-1) contains a peripheral and a membrane domain. Three antiporter-like subunits in the membrane domain, NuoL, NuoM, and NuoN (ND5, ND4 and ND2, respectively), are structurally similar. We analyzed the role of NuoN in Escherichia coli NDH-1. The lysine residue at position 395 in NuoN (_NLys³⁹⁵) is conserved in NuoL (_LLys³⁹⁹) but is replaced by glutamic acid (_MGlu⁴⁰⁷) in NuoM. Our mutation study on _NLys³⁹⁵ suggests that this residue participates in the proton translocation. Furthermore, we found that _MGlu⁴⁰⁷ is also essential and most likely interacts with conserved _LArg¹⁷⁵. Glutamic acids, _NGlu¹³³, _MGlu¹⁴⁴, and _LGlu¹⁴⁴, are corresponding residues. Unlike mutants of _MGlu¹⁴⁴ and _LGlu¹⁴⁴, mutation of _NGlu¹³³ scarcely affected the energy-transducing activities. However, a double mutant of _NGlu¹³³ and nearby _KGlu⁷² showed significant inhibition of these activities. This suggests that _NGlu¹³³ bears a functional role similar to _LGlu¹⁴⁴ and _MGlu¹⁴⁴ but its mutation can be partially compensated by the nearby carboxyl residue. Conserved prolines located at loops of discontinuous transmembrane helices of NuoL, NuoM, and NuoN were shown to play a similar role in the energy-transducing activity. It seems likely that NuoL, NuoM, and NuoN pump protons by a similar mechanism. Our data also revealed that _NLys¹⁵⁸ is one of the key interaction points with helix HL in NuoL. A truncation study indicated that the C-terminal amphipathic segments of _NTM14 interacts with the _Mβ sheet located on the opposite side of helix HL. Taken together, the mechanism of H⁺ translocation in NDH-1 is discussed.     

Catharanthus roseus mitogen-activated protein kinase 3 confers UV and heat tolerance to Saccharomyces cerevisiae

Catharanthus roseus is an important source of pharmaceutically important Monoterpenoid Indole Alkaloids (MIAs). Accumulation of many of the MIAs is induced in response to abiotic stresses such as wound, ultra violet (UV) irradiations, etc. Recently, we have demonstrated a possible role of CrMPK3, a C. roseus mitogen-activated protein kinase in stress-induced accumulation of a few MIAs. Here, we extend our findings using Saccharomyces cerevisiae to investigate the role of CrMPK3 in giving tolerance to abiotic stresses. Yeast cells transformed with CrMPK3 was found to show enhanced tolerance to UV and heat stress. Comparison of CrMPK3 and SLT2, a MAPK from yeast shows high-sequence identity particularly at conserved domains. Additionally, heat stress is also shown to activate a 43 kDa MAP kinase, possibly CrMPK3 in C. roseus leaves. These findings indicate the role of CrMPK3 in stress-induced MIA accumulation as well as in stress tolerance.     

Design and synthesis of factor Xa inhibitors and their prodrugs

In addition to our previously reported fluoro acrylamides Xa inhibitors 2 and 3, a series of potent and novel cyclic diimide amidine compounds has been identified. In efforts to improve their oral bioavailability, replacement of the amidine group with methyl amidrazone gives compounds of moderate potency (14, IC(50)=0.028 microM). In the amidoxime prodrug approach, the amidoxime compounds show good oral bioavailability in rats and dogs. High plasma level of prodrug 26 and significant concentration of active drug 26a were obtained upon oral administration of prodrug 26 in rats.     

Chromium is not an essential trace element for mammals: effects of a “low-chromium�?diet

Chromium was proposed to be an essential trace element over 50?years ago and has been accepted as an essential element for over 30?years. However, the studies on which chromium’s status are based are methodologically flawed. Whether chromium is an essential element has been examined for the first time in carefully controlled metal-free conditions using a series of purified diets containing various chromium contents. Male Zucker lean rats were housed in specially designed metal-free cages for 6?months and fed the AIN-93G diet with no added chromium in the mineral mix component of the diet, the standard AIN-93G diet, the standard AIN-93G diet supplemented with 200?μg?Cr/kg, or the standard AIN-93G diet supplemented with 1,000?μg?Cr/kg. The chromium content of the diet had no effect on body mass or food intake. Similarly, the chromium content of the diet had no effect on glucose levels in glucose tolerance or insulin tolerance tests. However, a distinct trend toward lower insulin levels under the curve after a glucose challenge was observed with increasing chromium content in the diet; rats on the supplemented AIN-93G diets had significantly lower areas (P?<?0.05) than rats on the low-chromium diet. The studies reveal that a diet with as little chromium as reasonably possible had no effect on body composition, glucose metabolism, or insulin sensitivity compared with a chromium-“sufficient??diet. Together with the results of other recent studies, these results clearly indicate that chromium can no longer be considered an essential element.     

Targeted deletion of integrin-linked kinase reveals a role in T-cell chemotaxis and survival

Integrin-linked kinase (ILK) is a serine/threonine kinase that is important in cell-matrix interactions and cell signaling. To examine the role of ILK in leukocyte trafficking and survival, we generated T cell-specific ILK knockouts by breeding ILK(flox/flox) mice to transgenic mice expressing Cre recombinase under control of the Lck proximal promoter. Thymic T cells from Lck-Cre(+)/ILK(flox/flox) mice had a marked reduction (>95%) in ILK protein levels. Thymic cellularity was comparable in 3- to 4-week-old mice, but a threefold diminution of thymic T cells became evident by 6 to 8 weeks of age in the T cell-specific ILK knockout mice due to increased cell death of double-positive (DP) T cells. Analysis of peripheral T cells by quantitative PCR and by breeding Lck-Cre(+)/ILK(flox/flox) mice to a YFP-transgenic reporter strain demonstrated an approximate 20-fold enrichment of ILK-competent cells, suggesting these cells have a competitive advantage in trafficking to and/or survival in peripheral lymphatic organs. We explored mechanisms related to altered cell trafficking and survival that might explain the decreases in thymic cellularity and enrichment for ILK-competent cells in the spleen and lymph nodes. We observed a >50% reduction in chemotaxis of ILK-deficient T cells to the chemokines CXCL12 (stromal cell-derived factor [SDF]-1alpha) and CCL19 (macrophage inflammatory protein [MIP]-3beta), as well as enhanced apoptosis of ILK-deficient cells upon stress. Signaling studies in ILK-deficient T cells demonstrated diminished phosphorylation of Akt on the activating phosphorylation site, Ser 473, and a concordant decrease in Akt kinase activity following stimulation with the chemokine SDF-1. Rac1 activation was also markedly diminished in ILK-deficient T cells following chemokine stimulation. These data extend the role of ILK to immune-cell trafficking and survival via modulation of Akt- and Rac-dependent substrates, and have implications for cell recruitment in both homeostatic and pathological processes.     

Effect of a fall of blood pressure on the release of catecholamines in the hypothalamus

The posterior hypothalamus of anaesthetized cats was superfused through a push-pull cannula and the release of endogenous catecholamines was determined in the superfusate which was continuously collected in 15 min periods. Fall in blood pressure elicited by nitroprusside or bleeding led to an increased rate of release of noradrenaline, adrenaline and dopamine in the hypothalamus. Transection of the brain causal to hypothalamus greatly reduced the rate of resting release of the catecholamines and abolished the enhancing effects of bleeding and nitroprusside. Determination of the catecholamines in samples which were collected in 90 s periods suggested a different pattern of release of the three catecholamines. Further shortening of the collection period (10 s) showed that the fall in blood pressure immediately increased the release of dopamine, while the rates of release of noradrenaline and adrenaline were increased gradually. Hypotension did not influence the rates of release of the catecholamines in the anterior hypothalamus. It is concluded that dopamine, adrenaline and noradrenaline systems of the hypothalamus are involved in the regulation of the arterial blood pressure. The different patterns of release might indicate that dopamine exerts a different function from those of noradrenaline and adrenaline in the normalization of the blood pressure after acute hypotension.     

Effect of 3-phenylindole on lipophilic ion and carrier-mediated ion transport across bilayer lipid membranes

The physical effects of 3-phenylindole, an antimicrobial compound which interacts with phospholipids, on ion transport across phosphatidylcholine-cholesterol bilayers have been investigated using three lipophilic ions and one ion-carrier complex. It was found that 3-phenylindole increased membrane electrical conductance of positively charged membrane probes and decreased electrical conductance of negatively charged probes. The enhancement of conductance detected by nonactin-K+ complex and tetraphenylarsonium+ was several orders of magnitude, whereas the suppression of conductance due to tetraphenylborate- and dipicrylamine- was less than a factor of ten. Presence of 3-phenylindole in aqueous phase slightly decreased adsorption of tetraphenylborate- and dipicrylamine- at the membrane surface. From the voltage dependence of the steady-state conductance it was shown that 3-phenylindole induced kinetic limitation of membrane transport of potassium mediated by nonactin. No such limitation was found in the case of tetraphenylarsonium+ transport. These results are shown to be consistent with the present concept of ion diffusion in membranes and the assumption that 3-phenylindole decreases the electric potential in the membrane interior. The asymmetry of the effect of 3-phenylindole on the magnitude of conductance changes for positively and negatively charged membrane permeable ions is also discussed as a reflection of the discreteness of both the absorbed 3-phenylindole and lipid dipoles.