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Living organisms are continuously under threat from a vast array of DNA-damaging agents, which impact genome DNA. DNA replication machinery stalls at damaged template DNA. The stalled replication fork is restarted via bypass replication by translesion DNA-synthesis polymerases, including the Y-family polymerases Polη, Polι, and Polκ, which possess the ability to incorporate nucleotides opposite the damaged template. To investigate the division of labor among these polymerasesin vivo, we generatedPOLη−/−,POLι−/−,POLκ−/−, double knockout (KO), and triple knockout (TKO) mutants in all combinations from human TK6 cells. TKO cells exhibited a hypersensitivity to ultraviolet (UV), cisplatin (CDDP), and methyl methanesulfonate (MMS), confirming the pivotal role played by these polymerases in bypass replication of damaged template DNA.POLη−/−cells, but notPOLι−/−orPOLκ−/−cells, showed a strong sensitivity to UV and CDDP, while TKO cells showed a slightly higher sensitivity to UV and CDDP than didPOLη−/−cells. On the other hand, TKO cells, but not all single KO cells, exhibited a significantly higher sensitivity to MMS than didwild-typecells. Consistently, DNA-fiber assay revealed that Polη plays a crucial role in bypassing lesions caused by UV-mimetic agent 4-nitroquinoline-1-oxide and CDDP, while all three polymerases play complementary roles in bypassing MMS-induced damage. Our findings indicate that the three Y-family polymerases play distinctly different roles in bypass replication, according to the type of DNA damage generated on the template strand.

Systemic inflammatory responses in patients undergoing extracorporeal membrane oxygenation (ECMO) contribute significantly to ECMO-associated morbidity and mortality. In recent years, the number of type 2 diabetes mellitus patients has increased, and the number of these patients undergoing ECMO has also increased. Type 2 diabetes mellitus is a high-risk factor for complications during ECMO. We studied the effects of ECMO on inflammatory response in a diabetic rat ECMO model. Twenty-eight rats were divided into 4 groups: normal SHAM group (normal rats: n = 7), diabetic SHAM group (diabetic rats: n = 7), normal ECMO group (normal rats: n = 7), and diabetic ECMO group (diabetic rats: n = 7). We measured the plasma levels of cytokines, tumor necrosis factor-α, and interleukin-6. Aspartate aminotransferase (AST), alanine aminotransferase (ALT), lactate dehydrogenase (LDH), blood urea nitrogen (BUN), creatinine (Cr), and liver-type fatty acid binding protein (L-FABP) were examined in the rat cardiopulmonary bypass model to ascertain organ damage. In addition, the lung wet-to-dry weight (W/D) ratio was measured as an index of pulmonary tissue edema. A pathologic evaluation of kidneys was conducted by hematoxylin-eosin (HE) and periodic-acid-methenamine-silver (PAM) staining. In the diabetic ECMO group, levels of cytokines, AST, ALT, LDH, and L-FABP increased significantly, reaching a maximum at the end of ECMO in comparison with other groups (p &lt, 0.05). In addition, hematoxylin-eosin and periodic acid-methenamine-silver staining of renal tissues showed marked injury in the ECMO group (normal ECMO and diabetic ECMO groups). Furthermore, when the normal ECMO and diabetic ECMO groups were compared, severe organ injury was seen in the diabetic ECMO group. There was remarkable organ injury in the diabetic ECMO group. These data demonstrate that diabetes enhances proinflammatory cytokine release, renal damage, and pulmonary edema during ECMO in an animal model.

The replicative polymerase δ (Polδ), consisting of four subunits, plays a pivotal role in chromosomal replication. Pold4, the smallest subunit of Polδ, is believed to contribute to the regulation of replication by facilitating repair in response to DNA damage. However, that contribution has not been fully elucidated. We here show that Pold4 contributes to the suppression of gene conversion in immunoglobulin-variable (IgV) gene diversification in the chicken DT40 lymphocyte cell line, where gene conversion diversifies the IgV gene through intragenic homologous recombination (HR) between diverged pseudo-V segments. IgV gene conversion is initiated by activation-induced cytidine deaminase-mediated uracil formation in the IgV gene, which in turn converts into an abasic site, leading to replication arrest. POLD4−/− cells exhibited an increased rate of IgV gene conversion. Moreover, the gene-conversion tract was lengthened and the usage of pseudo-V segments was altered, showing a preference, to use the diverged sequence as a donor in POLD4−/− cells. These data suggest that Pold4 is involved in the regulation of HR-mediated gene conversion in IgV diversification. By contrast, the rate in HR-mediated, sister-chromatid exchange and gene-targeting induced by an I-SceI endonclease-mediated DNA double-strand break exhibited by POLD4−/− cells was indistinguishable from that by wild-type cells. These findings indicate that the functionality of general HR is preserved in POLD4−/− cells. In conclusion, Pold4 is involved in the suppression of IgV-gene conversion without affecting the general functionality of HR.

Azo dyes, including Sudan I, Orange II and Orange G, are industrial dyes that are assumed to have genotoxic potential. However, neither the type of DNA damage induced nor the structural features responsible for toxicity have been determined. We used a panel of DNA-repair-pathway-deficient mutants generated from chicken DT40 cells to evaluate the ability of these azo dyes to induce DNA damage and to identify the type of DNA damage induced. We compared the structurally related azo dyes Sudan I, Orange II and Orange G to identify the structural features responsible for genotoxicity. Compared with wild type cells, the double-strand break repair defective RAD54-/-/KU70-/- cells were significantly more sensitive to Sudan I, but not to Orange II or Orange G. The quantum-chemical calculations revealed that Sudan I, but not Orange II or Orange G, has a complete planar aromatic ring structure. These suggest that the planar feature of Sudan I is critical to the inducing of double-strand breaks. In summary, we used a DNA-repair mutant panel in combination with quantum-chemical calculations to provide a clue to the chemical structure responsible for genotoxicity.

The high prevalence of house dust mite (HDM) allergy is a growing health problem worldwide, and the characterization of clinically important HDM allergens is a prerequisite for the development of diagnostic and therapeutic strategies. Here, we report a novel HDM allergen that belongs structurally to the highly conserved Rid/YjgF/YER057c/UK114 family (Rid family) with imine deaminase activity. Isolated HDM cDNA, named der f 34, encodes 128 amino acids homologous to Rid-like proteins. This new protein belongs to the Rid family and has seven conserved residues involved in enamine/imine deaminase activity. Indeed, we demonstrated that purified Der f 34 had imine deaminase activity that preferentially acted on leucine and methionine. Native Der f 34 showed a high IgE binding frequency as revealed by two-dimensional immunoblotting (62.5%) or ELISA (68%), which was comparable with those of a major HDM allergen Der f 2 (77.5 and 79%, respectively). We also found that Der f 34 showed cross-reactivity with another prominent indoor allergen source, Aspergillus fumigatus. This is the first report showing that the Rid family imine deaminase represents an additional important pan-allergen that is conserved across organisms.

AND-1/Ctf4 bridges the CMG helicase and DNA polymerase alpha, facilitating replication. Using an inducible degron system in avian cells, we find that AND-1 depletion is incompatible with proliferation, owing to cells accumulating in G2 with activated DNA damage checkpoint. Replication without AND-1 causes fork speed slow-down and accumulation of long single-stranded DNA (ssDNA) gaps at the replication fork junction, with these regions being converted to DNA double strand breaks (DSBs) in G2. Strikingly, resected forks and DNA damage accumulation in G2, but not fork slow-down, are reverted by treatment with mirin, an MRE11 nuclease inhibitor. Domain analysis of AND-1 further revealed that the HMG box is important for fast replication but not for proliferation, whereas conversely, the WD40 domain prevents fork resection and subsequent DSB-associated lethality. Thus, our findings uncover a fork protection function of AND-1/Ctf4 manifested via the WD40 domain that is essential for proliferation and averts genome instability., AND-1, the vertebrate orthologue of Ctf4, is a critical player during DNA replication and for maintenance of genome integrity. Here the authors use a conditional AND-1 depletion system in avian DT40 cells to reveal the consequences of the lack of AND-1 on cell proliferation and DNA replication.

RecQ family DNA helicases function in the maintenance of genome stability. Mice deficient in RecQL5, one of five RecQ helicases, show a cancer predisposition phenotype, suggesting that RecQL5 plays a tumor suppressor role. RecQL5 interacts with Rad51, a key factor in homologous recombination (HR), and displaces Rad51 from Rad51-single stranded DNA (ssDNA) filaments in vitro. However, the precise roles of RecQL5 in the cell remain elusive. Here, we present evidence suggesting that RecQL5 is involved in DNA interstrand crosslink (ICL) repair. Chicken DT40 RECQL5 gene knockout (KO) cells showed sensitivity to ICL-inducing agents such as cisplatin (CDDP) and mitomycin C (MMC) and a higher number of chromosome aberrations in the presence of MMC than wild-type cells. The phenotypes of RECQL5 KO cells resembled those of Fanconi anemia gene KO cells. Genetic analysis using corresponding gene knockout cells showed that RecQL5 is involved in the FANCD1 (BRCA2)-dependent ICL repair pathway in which Rad51-ssDNA filament formation is promoted by BRCA2. The disappearance but not appearance of Rad51-foci was delayed in RECQL5 KO cells after MMC treatment. Deletion of Rad54, which processes the Rad51-ssDNA filament in HR, in RECQL5 KO cells increased sensitivity to CDDP and further delayed the disappearance of Rad51-foci, suggesting that RecQL5 and Rad54 have different effects on the Rad51-ssDNA filament. Furthermore, the frequency and variation of CDDP-induced gene conversion at the immunoglobulin locus were increased in RECQL5 KO cells. These results suggest that RecQL5 plays a role in regulating the incidence and quality of ICL-induced recombination.

Claspin was originally identified as a Check1 (Chk1)-interacting protein. Claspin and Rad17 are reportedly involved in the DNA damage-induced phosphorylation of Chk1, a hallmark of checkpoint activation. To understand the cellular functions of Claspin and the functional relationship between Claspin and Rad17, we generated Claspin(-/-) and Claspin(-/-)/RAD17(-) cells using chicken DT40 cells, which contain an exogenously introduced Claspin that can be suppressed by the addition of doxycycline (Dox). In the presence of Dox, Claspin(-/-) cells ceased growth within 2 days, leading to cell death. In addition, a remarkable reduction in the rate of DNA elongation was observed in Claspin-depleted cells, suggesting that Claspin plays a critical role in DNA replication in the absence of exogenous stress. When cells were exposed to methyl methanesulfonate (MMS), a DNA damaging agent, RAD17(-) cells showed a greater defect in checkpoint activation than Claspin(-/-) cells as monitored by progression of cell cycle and phosphorylation of Chk1. Knocking out RAD17 gene showed almost no additive effects on cell death and DNA elongation rates in Claspin-depleted cells.

DNA double strand breaks (DSBs) induced by etoposide, an inhibitor of DNA topoisomerase II, are repaired mainly by non-homologous end joining (NHEJ). Unexpectedly, it was found that at high doses of etoposide, proteins involved in NHEJ, such as KU70/80, DNA-PKcs and ARTEMIS/SNM1C, trigger apoptosis rather than repair of DSBs. Because ARTEMIS is a member of the SNM1 protein family that includes SNM1A and APOLLO/SNM1B, this study examined whether SNM1A and/or APOLLO are also involved in etoposide-induced apoptosis. Using SNM1A(-/-) and APOLLO(-/-) cells, it was found that both SNM1A and APOLLO participate in etoposide-induced apoptosis. Although cell viability monitored by MTT assay did not differ between SNM1A(-/-)/APOLLO(-/-)/ARTEMIS(-/-), SNM1A(-/-)/APOLLO(-/-), and single gene knockout cells, DNA fragmentation monitored by TUNEL assay differed between these cells, suggesting that the three SNM1 family nucleases function independently, at least during the induction of apoptotic DNA fragmentation.

The mitigating effects of hymexazole (HMI), choline chloride, indolebutyric acid, ethephon and isoprothiolane on cadmium (Cd) stress and accumulation in Japanese millet were evaluated under aquatic conditions. Among five PGRs, HMI showed the highest mitigating effect; however, in this experiment, almost no effect of calcium (Ca), which has been demonstrated to be a Cd mitigator, on Cd accumulation was observed and the mitigating effect of Ca on Cd stress was less than that of HMI.

KU70(-/-) and DNA-PKcs(-/-/-)chicken DT40 cells are reportedly highly sensitive to the DNA topoisomerase II inhibitor etoposide. Here we report that KU70 and DNA-PKcs unexpectedly function together during the induction of apoptosis after exposure to high levels of etoposide. In the presence of 100 microM etoposide, apoptosis was induced within 1 h in wild type DT40 cells but not in KU70(-/-) and DNA-PKcs(-/-/-) cells. In addition, the DNA-PK inhibitors NU7026 and wortmannin, as well as the caspase inhibitor Z-VAD-FMK, inhibited etoposide-induced apoptosis in wild type cells. Although Artemis(-/-) cells also showed defects in the etoposide-induced apoptosis, the other mutants defective in nonhomologous end-joining (NHEJ), LIG4(-/-), XRCC4(-), and XLF(-/-) cells were capable to induce apoptosis. When cells were treated with high doses of etoposide, the chromatin binding of DNA-PKcs was impaired by deletion of KU70 but not of Artemis, suggesting that KU70 acts upstream of DNA-PKcs and Artemis acts downstream of DNA-PKcs in the apoptotic pathway like the NHEJ pathway. These results suggest that the proteins involved in the early stage of NHEJ pathway including Artemis but not the downstream factors decide the cell fate by selecting apoptosis or DNA repair according to the degree of DNA damage.

RECQL1 and RECQL5 as well as BLM reportedly interact with TOP3α whose defect is lethal for the cell. Therefore in this study, we characterized recql5 / recql1 / blm triple mutants from DT40 cells to determine whether the triple mutants show a top3α disrupted cell-like phenotype. The triple mutants are viable. Moreover, both blm / recql1 and recql5 / blm cells, and recql5 / recql1 / blm cells grew slightly slower than blm cells, that is, triple mutant cells grew almost the same rate as either of the double mutant cells. The blm cells showed sensitivity to methyl methanesulfonate (MMS) and ultraviolet light (UV), about a 10-fold increase in sister chromatid exchange (SCE), and about a 3-fold increase in damage-induced mitotic chiasma compared to wild-type cells. The triple mutants showed the same sensitivity to MMS or UV and the same frequency of damage-induced mitotic chiasma compared to those of blm cells, indicating that unlike BLM, RECQL1 and RECQL5 play a little role in the repair of or tolerance to DNA damages. However, recql5 / blm cells showed higher frequency of SCE than blm cells, whereas the RECQL1 gene disruption had no effect on SCE in blm cells and even in recql5 / blm cells.

Summary 1To investigate whether the role of symbiotic fungi in lignin degradation and food provision differs among fungus-growing termites (four species of Macrotermes, three species of Odontotermes, Hypotermes makhamensis, Ancistrotermes pakistanicus and Pseudacanthotermes militaris), the chemical composition of fungus combs of different ages and the chitinolytic activity in the gut of termites were analysed. In addition, the carbon stable isotope ratios (δ13C) in old combs, worker termite tissues and fungal nodules (aggregated conidia) were compared. 2In Macrotermes spp., the carbohydrate : lignin ratio of the combs increased with increased comb age, but it decreased or remained the same in Odontotermes spp., H. makhamensis, A. pakistanicus and P. militaris. In contrast, the chitin : carbohydrate ratio and ash content increased more over time in the combs of Odontotermes spp., H. makhamensis, A. pakistanicus and P. militaris than in the combs of Macrotermes spp. 3Chitinolytic activity in the gut of workers was higher in O. takensis, H. makhamensis and A. pakistanicus than in M. annandalei. 4In all species examined, the δ13C value increased in the sequence: old combs < termites < fungal nodules. Based on a two-source model of δ13C, the contribution of fungi to termite nutrition was considered higher in Odontotermes spp., H. makhamensis, A. pakistanicus and P. militaris than in Macrotermes spp. 5These results suggest that symbiotic fungi play different roles among fungus-growing termites. In Macrotermes spp., the main role of symbiotic fungi is to degrade lignin, so that the termites can utilize cellulose more efficiently, whereas in Odontotermes spp., H. makhamensis, A. pakistanicus and P. militaris, it is to serve as a food source.

Two kinds of PCR-product cDNAs that encode premature lysozyme peptides (Rs-Lys1 and Rs-Lys2) were cloned from workers of a Japanese damp-wood termite, Reticulitermes speratus. The Rs-Lys1 and Rs-Lys2 cDNAs encoded deduced sequences of 170 and 164 amino acids, respectively. Alignment of these sequences with those of other insect lysozymes showed that the cDNAs encode lysozyme homologues with putative signal peptides, insertions eight amino acids long, and a relatively long C-terminus (13-17 amino acids). A maximum likelihood tree, constructed using the cDNA sequences, indicated that the termite lysozymes are related to those of mosquitoes and lepidopterans. Southern-blotting analysis identified single copies of these lysozyme genes in the termite. Reverse transcript (RT)-PCR and in situ hybridization experiments showed that Rs-Lys1 and Rs-Lys2 are expressed in the salivary glands of worker termites. Here, we discuss the possible digestive function of these lysozymes.

1. Nitrogen and carbon isotope ratios and uric acid concentrations in fungus-growing termites (Isoptera: Termitidae: Macrotermitinae), sampled in Cameroon and Thailand, were determined in order to compare castes that are known to differ in behaviour and feeding habits. 2. Nitrogen isotope ratios (δ15N) were either not significantly changed or lower in workers compared with the diet (the fungus combs), whereas carbon isotope ratios (δ13C) were higher in worker termites than in the fungus combs. 3. In old workers, δ15N values were unexpectedly low and correlated negatively with whole-body uric acid concentrations. This indicates that older workers retain uric acid, which has a low δ15N value, to conserve nitrogen within the colony and, furthermore, that older colony members may ultimately be consumed by younger conspecifics.

Tissue N and C stable isotope ratios of two sympatric termite species were investigated in relation to those of their stored food materials at two savanna sites in north-eastern Australia (Woodstock and Oak Valley). At both sites, the δ 15 N values of termite tissues and the stored food materials of the litter-feeding species Amitermes laurensis were significantly greater than their equivalents in the grass-harvesting species Drepanotermes rubriceps . In addition, the carbohydrate/lignin and C/N ratios were higher and chitin/carbohydrate ratios were lower in food materials stored in the mounds of D. rubriceps . Both sets of analyses confirmed that D. rubriceps assimilates less decomposed food materials than A. laurensis . Tissue δ 13 C values indicated that both species assimilated similar proportions of C3 and C4 plant materials at the Woodstock site, but that the workers of A. laurensis assimilated a greater proportion of C3 plant material at the Oak Valley site. The δ 15 N values of termites from the Woodstock site were significantly greater than those from Oak Valley, although the differences in order at each site were consistent between species. The results indicate that N and C stable isotope ratios must be carefully compared across habitats.

Summary Distribution of lysozyme and protease, and amino acid concentration in the guts of a wood-feeding termite, Reticulitermes speratus (Kolbe) (Isoptera, Rhinotermitidae) were studied to examine the possibility that termites digest symbiont bacteria transferred by trophallaxis. Total lysozyme activity was found predominantly in the salivary gland and to a minor extent in the digestive tracts. However, specific lysozyme activity was high in the foregut as well as in the salivary gland. The similarity of the lysozyme pH profile of the salivary gland and of the foregut suggested that the foregut lysozyme came from the salivary gland. Major protease activity having the optimum pH of 7.5 was found in the midgut. Total free amino acid amount and concentration in the midgut was higher than elsewhere in the digestive tract. The possibility that lysozyme secreted from the salivary gland into the foregut digests hindgut bacteria transferred by trophallaxis was discussed.

In order to investigate the role of the mutualistic fungus, Termitomyces sp., in the fungus-growing termite, Macrotermes gilvus , we applied CP/MAS 13 C NMR and selected proximate analyses to fungus comb of different ages and degrees of maturation. We found evidence that lignin degradation took place progressively in the fungus comb. In vitro digestibility of cellulose in old fungus comb, on which the termites feed, was approximately 3-fold higher than that in the fresh part. These results confirm the ‘lignin degradation hypothesis’ that the role of the mutualistic fungi is to degrade lignin and enhance the digestibility of cellulose for the termites, suggesting the ability of the termite–fungus association to make extremely efficient use of plant material.

Emission rates and carbon isotope ratios of CH4, emitted by workers of termites, and of CH4, emitted from their mounds, were observed in a dry evergreen forest in Thailand to estimate the proportion of CH4 oxidized during emission through the mound. The δ13C of CH4 emitted from a termite mound (−70.9 to −82.4‰) was higher than that of CH4 emitted by workers in the mound (−85.4 to −97. l‰). Using a fractionation factor (a = 0.987) for oxidation of CH4 which was obtained in the incubation experiment, an emission factor defined as (CH4 emitted from a termite mound/CH4 produced by termites) was calculated. The emission factor obtained in each termite mound was nearly zero for Macrotermes (fungus-growing termites), of which the nest has a thick soil wall and subterrannean termites, and 0.17 to 0.47 for Termitinae (small-mound-making termites). Global CH4 emission by termites was estimated on the basis of the CH4 emission rates by workers and termite biomass with the emission factors. The calculated result was 1.5 to 7.4 Tg/y (0.3 to 1.3% of total source), which is considerably smaller than the estimate by the IPCC [1994].

Methane and hydrogen emission rates and the δ13C of CH4 were observed for various termites in Australia, Thailand and Japan. Combined with the already reported emission rates of CH4 in the literature, the phylogenetic trend was examined. Emission rates of the observed termites were categorized into five groups: group I with high CH4 and low H2 emission rates with a CH4/H2 ratio of typically 10/1; group II with high CH4 and high H2 emissions with a CH4/H2 ratio of 4/1–1/2; group III with low emission rates of CH4 and H2; group IV with high H2 and insignificant CH4 emissions; and group V with insignificant emissions for both CH4 and H2. In lower termites, there are both colonies infected and uninfected with methanogens even in the same species, and no specific trend in CH4 and H2 emissions was observed within a genus. Whether protozoa in the hindgut of termites are infected with methanogens or not and the differences in species compositions of protozoa are possibly responsible for the inter-colonial variations. The proportions of infected colonies were possibly small for the family Kalotermitidae (dry wood feeders), and relatively large for families of wet or damp wood feeders. The hydrogen emission rate possibly depends on the locality of methanogens: namely, whether they are intracellular symbionts of protozoa or whether they are attached to the hindgut wall. Emission rates of higher termites were classified into groups according to genera and the diet. Most species of soil or wood/soil interface feeders classified into group I, while the soil feeders Dicuspiditermes in Thailand and Amitermes in Australia were classified into groups with high H2 emission rates. Typical wood-feeding termites and fungus-growing termites were classified into group III. The results indicate that higher termites tend to increase the CH4 emission rate during dietary evolution from wood- to soil-feeding, and two types of the system with different efficiencies of interspecies transfer of H2 have been formed. The δ13C of CH4 was discernible with a difference in the decomposition process in the termite–symbiont system among lower termites, fungus-growing termites and other higher termites.

SLC10A4 belongs to the sodium bile acid cotransporter family, but has no transport activity for bile acids. We performed multiple amino acid alignments and examined the relationships between the SLC10 proteins. The extracellular N-terminus of SLC10A4 was predicted to be relatively longer at the amino acid level than those of SLC10A1, SLC10A2 and SLC10A6. We examined the relationship between the N-terminus and transport activity of SLC10A4. Rat Slc10a4 is predominantly expressed in rat cholinergic neurons; therefore, TE671 cells expressing the acetylcholine receptor and acetylcholinesterase were used. After thrombin treatment, western blotting and immunofluorescence staining demonstrated that the N-terminus of SLC10A4 might be cleaved. Substrates were added to the cells, and their uptake was quantified by liquid chromatography tandem mass spectrometry. Lithocholic acid (LCA) and taurocholic acid (TCA) uptake and cell death effects of LCA were increased by thrombin treatment. After RNA interference treatment for SLC10A4, bile acid uptake was also quantified. In consequence, increases in the LCA and TCA uptake did not occur. Therefore, SLC10A4 may have low activity but becomes activated by proteases, including thrombin, following cleavage. We have demonstrated that SLC10A4 appears to be a protease-activated transporter and transports bile acids.

SUMO conjugation is a reversible posttranslational modification that regulates protein function. SENP1 is one of the six SUMO-specific proteases present in vertebrate cells and its altered expression is observed in several carcinomas. To characterize SENP1 role in genome integrity, we generated Senp1 knockout chicken DT40 cells. SENP1(-/-) cells show normal proliferation, but are sensitive to spindle poisons. This hypersensitivity correlates with increased sister chromatid separation, mitotic slippage, and apoptosis. To test whether the cohesion defect had a causal relationship with the observed mitotic events, we restored the cohesive status of sister chromatids by introducing the TOP2α(+/-) mutation, which leads to increased catenation, or by inhibiting Plk1 and Aurora B kinases that promote cohesin release from chromosomes during prolonged mitotic arrest. Although TOP2α is SUMOylated during mitosis, the TOP2α(+/-) mutation had no obvious effect. By contrast, inhibition of Plk1 or Aurora B rescued the hypersensitivity of SENP1(-/-) cells to colcemid. In conclusion, we identify SENP1 as a novel factor required for mitotic arrest and cohesion maintenance during prolonged mitotic arrest induced by spindle poisons.

The distributions of lysozyme and protease activities and of amino acids was measured in the guts of five species of higher termites, Macrotermes annandalei, Odontotermes formosanus, Pericaproitermes nitobei, Termes comis and Nasutitermes takasagoensis. Lysozyme activity was found only in M. annandalei, P. nitobei and N. takasagoensis. Protease activity was high in the midgut of all species but negligible elsewhere in the gut. Amino acid concentration was highest in the midgut of all species of workers.

Currently, various formulas with different fatty acid compositions are used for enteral nutrition (EN). All formulas contain various concentrations of essential fatty acids: linoleic acid (LA) and alpha-linolenic acid (ALA); LA is biotransformed into arachidonic acid (AA) and ALA into eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) in vivo. Some formulas contain preformed EPA and DHA. However, the effects of the differences in the fatty acid composition on the fatty acid status of patients receiving long-term EN is not clear. We measured serum fatty acid concentrations in 50 patients with neurological diseases receiving long-term EN. The data were then compared retrospectively with reference to the fatty acid compositions of the formulas used. All of the patients received almost their entire nutritional intake via EN for at least 1 year. Blood samples were obtained just before injecting the EN solution. Among the formulas that did not include EPA or DHA, formulas with low ALA concentrations were associated with low serum EPA and DHA. Conversely, the ALA-enriched formulas with reduced LA concentrations significantly increased EPA and DHA levels, although the levels remained lower than the control values. With the formula containing EPA and DHA, the EPA and DHA levels reached control values. Therefore, the fatty acid composition of the EN formulas affected the fatty acid status of patients receiving long-term EN. Formulas containing preformed EPA and DHA with suitable amounts of essential fatty acids may benefit these patients.

Manganese-dependent superoxide dismutase (SOD2) serves as the primary defense against mitochondrial superoxide, and decreased SOD2 activity results in a range of pathologies. To investigate the events occurring soon after depletion of SOD2, we generated SOD2 gene knockout chicken DT40 cells complemented with a human SOD2 (hSOD2) cDNA, whose expression can be switched off by doxycycline (Dox). When SOD2 was depleted by the addition of Dox, the cells grew slightly slower and formed fewer colonies than cells expressing hSOD2. In addition, these cells showed a high sensitivity to paraquat, which produces superoxide, and died through apoptosis. In contrast to results obtained with mouse and DrosophilaSod2 mutants, we found no indication of an increase in DNA lesions due to depletion of SOD2.

Nitrogen and carbon stable isotope ratios and uric acid concentrations in termites sampled from a dry evergreen forest in Thailand, were determined across three kinds of feeding habits. Feeding habits of Microcerotermes crassus, which is an abundant wood-feeder, and Dicuspiditermes makhamensis, a common soil-feeding termite, were confirmed by isotopic signatures. Lichen feeding termites (Hospitalitermes birmanicus, H. bicolor and H. ataramensis) were characterized by low delta15N values, suggesting that they assimilated nitrogen deposited from the atmosphere. There was also a significant difference in uric acid concentrations between termites representing different feeding habits. No significant relationships were found between uric acid concentrations and delta15N or delta13C in Hospitalitermes. However, delta15N values were correlated with C/N ratios in H. birmanicus, except in one colony of H. ataramensis. delta13C values in both species were negatively correlated with C/N ratios.

We have developed a top-gate type of field-effect transistor with a single-crystal SrTiO3 channel and a DyScO3 gate insulator stack consisting of an epitaxial interface layer and an amorphous breakdown barrier layer. We show that the zero-bias conductivity of the transistor channel is strongly affected by the presence of charged traps in the amorphous gate insulator. Low off-state current could only be achieved in devices that were fabricated at an oxygen ambient pressure of 10 mTorr. At lower pressures, metallic channel interfaces were obtained, even after post-annealing in air. When both epitaxial and amorphous DyScO3 films were grown at 10 mTorr of oxygen, the on/off ratio of the field-effect transistors (FETs) reached 106. We argue that when designing oxide FETs, it is necessary to consider not only breakdown characteristics, but also the charged trap density in wide-gap oxide insulators.

It has recently been claimed that our β-bungarotoxin preparation contained three contaminants, including a postsynaptic toxin. We have extended our purification procedure and found no evidence of such contaminants.

The neurotoxin, β-bungarotoxin (β-BuTX) purified from Bungarus multicinctus snake venom exerts various effects on frog and rat motor nerve terminals. Exposure to β-BuTX triggers, in the terminals, impulses which are rapidly distributed to the whole motor unit and elicit contractions of the muscle. β-BuTX raises the frequency of spontaneous release of acetylcholine quanta and inhibits the release evoked by nerve impulses, all of which leads to complete cessation of transmitter release. Subsequently, the motor axon disappears from the endplate and the Schwann cell establishes extensive synaptic contact with the muscle fibre. After only 30 min in β-BuTX, some endplates attain a stage of denervation similar to that seen several days after transecting the motor nerve. The neuromuscular effects of β-RuTX may result from its phospholipase activity and from an increase in the level of ionized calcium inside the terminals.

β-Bungarotoxin (β-BuTX) induces twitching of innervated frog muscle and subsequently blocks transmitter release from motor nerve endings. These actions of β-BuTX are prevented if the nerve-muscle junctions are pretreated with a low concentration of phospholipase A 2 from bee venom. When the Ca 2+ in the external fluid is replaced by Sr 2+ , the phospho­lipase activity of β-BuTX is negligible and, in these conditions, β-BuTX causes a decrease in the amplitude of endplate potentials but does not go on to block transmitter release completely.

Methane emissions from different castes of four species of termites were observed in Narathiwat, Thailand. The results show that reproductive castes (queen and king), soldier and immature, which cannot feed themselves, emit less methane than dose worker. Observed results suggest that the quality of their diet causes the difference in methane emission among the castes.