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The compartmentalized anaerobic reactor (CAR) is a novel, promising high-rate bioreactor® for anaerobic digestion process. The feasibility of early-warning on the performance of lab-scale CAR was investigated. CAR showed less stable performance at high organic loading rates (OLRs) than that at regular OLRs. At high OLRs, the system in response to the fluctuation of influent chemical oxygen demand (COD) concentration showed higher variation in terms of effluent COD and volatile fatty acids (VFA) concentration. Two newly defined parameters, VLR saturation ( S VLR ) and VFA saturation ( S VFA ) could be used as indicators for early-warning on the performance of anaerobic reactors. The system performance was relatively stable when CAR was operated at regular OLR, during which S VLR and S VFA were below 0.89 and 0.4 respectively. It tended to be unstable under high OLR conditions when S VLR and S VFA were close to 1, and deteriorated when S VLR and S VFA were above 1. [ABSTRACT FROM AUTHOR]

The compartmentalized anaerobic reactor (CAR) is a patent novel high-rate reactor, which consists of three compartments. The reactor has a great potential for application due to its many advantages. In this work, the microbial consortium, spatial distribution, and their relationship with performance of CAR were investigated by means of polymerase chain reaction, denaturing gradient gel electrophoresis, and fluorescence in situ hybridization. The results showed that the predominant archaea were Methanobacterium, Methanosaeta, and Methanospirillum, and the predominant bacteria were Firmicutes, Deltaproteobacteria, Spirochaetes, Actinobacteria, and Gammaproteobacteria in the microbial consortium. The methanogenic archaea (MA), the hydrogen-producing acetogenic bacteria (HAB), and the hydrolytic fermentative bacteria (HFB) were found to be predominant in the upper, middle, and bottom compartments, respectively. The results revealed that the granular sludge took on a stratified microbial structure. The HFB, HAB, and MA were located in the outer shell, middle layer, and core, respectively. The microbial populations from the bottom compartment were relatively homogeneous in the granular sludge, and from the middle and upper compartments, they were relatively heterogeneous in the granular sludge. The microbial consortia and their spatial distribution were in accordance with the organic loading rate and chemical components in the three compartments. [ABSTRACT FROM AUTHOR]

Abstract: In the present study, a method for prediction of the toxicity of pharmaceutical wastewaters containing antibiotics to microbial communities in anaerobic digestion treatment was developed. Luminescent bacterium assay was carried out with Vibrio fischeri as indicator. The individual and joint toxicities of antibiotics and anaerobic digestion metabolites were investigated by using the 15-min half inhibitory concentration (15min-IC50) at pH 7.00±0.05. The results showed that the 15min-IC50 of Amoxicillin, Kanamycin, Lincomycin and Ciprofloxacin were 3.99, 5.11, 4.32 and 5.63gL−1 respectively, and the toxicity descended in the order of Amoxicillin, Lincomycin, Kanamycin and Ciprofloxacin. Using equitoxic ratio mixing method, the joint toxicities of four anaerobic digestion intermediates, the four intermediates together with Amoxicillin, Ciprofloxacin, Kanamycin or Lincomycin were determined, which displayed that their interactions were additive, additive, synergistic, synergistic and synergistic respectively. Finally the joint effect of all intermediates and antibiotics was synergistic. The method has promising applications in evaluating the joint toxicity of anaerobic digestion intermediates and antibiotics, and has laid the foundations for assessing the feasibility of anaerobic treatment of pharmaceutical wastewater containing antibiotics. [Copyright &y& Elsevier]

Abstract: In order to evaluate the effect of anaerobic digestion intermediates and antibiotics in pharmaceutical wastewaters on anaerobic digestion process, their acute toxicities were tested using the 15min median inhibitory concentration (IC50) at pH 7.00±0.05. The results showed that the IC50 of ethanol, acetate, propionate and butyrate were 19.40, 20.71, 10.47 and 12.17gL−1 respectively, which suggested the toxicity descended in the order of propionate, butyrate, ethanol and acetate. The IC50 of aureomycin, polymyxin and chloromycetin were 12.06, 6.24 and 429.90mgL−1 respectively, which indicated the toxicity descended in the order of polymyxin, aureomycin and chloromycetin. Using equitoxic ratio mixing method, the joint toxicities of five groups referred by A (four anaerobic digestion intermediates), B (four anaerobic digestion intermediates and aureomycin), C (four anaerobic digestion intermediates and polymyxin), D (four anaerobic digestion intermediates and chloromycetin) and E (four anaerobic digestion intermediates, aureomycin, polymyxin and chloromycetin) were investigated respectively. Their interactions were additive (A), synergistic (B), additive (C), synergistic (D) and synergistic (E). The investigation would lay a basis for the optimization of anaerobic biotechnology for pharmaceutical wastewater treatment. [Copyright &y& Elsevier]

Abstract: The compartmentalized anaerobic reactor (CAR) is a patent novel high-rate reactor and shows a great potential for its application. The hydraulic characteristics and their effects on the working performance of CAR were investigated. The flow pattern tended to plug flow at normal organic loading rate (OLR) and completely mixed flow at high OLRs. The relation of hydraulic dead space (HDS or V h) with hydraulic loading rate (HLR or L) and biogas production rate (BPR or G) was V h =3.75L+0.19G−9.47. The hydraulic efficiency of CAR was good or near to good. Both HLR and BPR had significant effects on the hydraulic efficiency, but their effect became less at super-high OLR. They also had a slight influence on the effective volume ratios of CAR, but the influence of BPR almost disappeared at super-high OLR. The good working performance of CAR was ascribed to the improved reactor configuration. [Copyright &y& Elsevier]

Summary: Histone-tail modifications play a fundamental role in the processes that establish chromatin structure and determine gene expression . One such modification, histone methylation, was considered irreversible until the recent discovery of histone demethylases. Lsd1 was the first histone demethylase to be identified . Lsd1 is highly conserved in many species, from yeast to humans, but its function has primarily been studied through biochemical approaches. The mammalian ortholog has been shown to demethylate monomethyl- and dimethyl-K4 and -K9 residues of histone H3 . Here we describe the effects of Lsd1 mutation in Drosophila. The inactivation of dLsd1 strongly affects the global level of monomethyl- and dimethyl-H3-K4 methylation and results in elevated expression of a subset of genes. dLsd1 is not an essential gene, but animal viability is strongly reduced in mutant animals in a gender-specific manner. Interestingly, dLsd1 mutants are sterile and possess defects in ovary development, indicating that dLsd1 has tissue-specific functions. Mutant alleles of dLsd1 suppress positional-effect variegation, suggesting a disruption of the balance between euchromatin and heterochromatin. Taken together, these results show that dLsd1-mediated H3-K4 demethylation has a significant and specific role in Drosophila development. [Copyright &y& Elsevier]

Dysregulation of CDK8 (Cyclin-Dependent Kinase 8) and its regulatory partner CycC (Cyclin C), two subunits of the conserved Mediator (MED) complex, have been linked to diverse human diseases such as cancer. Thus, it is essential to understand the regulatory network modulating the CDK8-CycC complex in both normal development and tumorigenesis. To identify upstream regulators or downstream effectors of CDK8, we performed a dominant modifier genetic screen in Drosophila based on the defects in vein patterning caused by specific depletion or overexpression of CDK8 or CycC in developing wing imaginal discs. We identified 26 genomic loci whose haploinsufficiency can modify these CDK8- or CycC-specific phenotypes. Further analysis of two overlapping deficiency lines and mutant alleles led us to identify genetic interactions between the CDK8-CycC pair and the components of the Decapentaplegic (Dpp, the Drosophila homolog of TGFβ) signaling pathway. We observed that CDK8-CycC positively regulates transcription activated by Mad (Mothers against dpp), the primary transcription factor downstream of the Dpp/TGFβ signaling pathway. CDK8 can directly interact with Mad in vitro through the linker region between the DNA-binding MH1 (Mad homology 1) domain and the carboxy terminal MH2 (Mad homology 2) transactivation domain. Besides CDK8 and CycC, further analyses of other subunits of the MED complex have revealed six additional subunits that are required for Mad-dependent transcription in the wing discs: Med12, Med13, Med15, Med23, Med24, and Med31. Furthermore, our analyses confirmed the positive roles of CDK9 and Yorkie in regulating Mad-dependent gene expression in vivo. These results suggest that CDK8 and CycC, together with a few other subunits of the MED complex, may coordinate with other transcription cofactors in regulating Mad-dependent transcription during wing development in Drosophila. Author summary: CDK8 and its dedicated partner CycC are conserved subunits of the Mediator complex that bridges transcription factors with RNA Polymerase II in eukaryotes. Here we explore the function and regulation of the CDK8-CycC pair in Drosophila by performing a dominant modifier genetic screen based on wing vein patterning defects caused by specific alteration of CDK8-CycC activities. We have observed that multiple components of the Dpp/TGFβ signaling pathway genetically interact with CDK8-CycC. CDK8 and CycC positively regulate gene expression activated by Mad, the key transcription factor downstream of Dpp/TGFβ signaling, and CDK8 can directly interact with the linker region of the Mad protein. We also identify additional, but not all, subunits of the Mediator complex that play positive roles in regulating Mad-dependent gene expression. Given the fundamental role of Dpp/TGFβ signaling in regulating development and its misregulation in a variety of diseases, understanding how Mad/Smad interacts with the Mediator complex may have broad implications in understanding the pathogenesis of these diseases. [ABSTRACT FROM AUTHOR]

Abstract The complex interplay between genetic and environmental factors, such as diet and lifestyle, defines the initiation and progression of multifactorial diseases, including cancer, cardiovascular and metabolic diseases, and neurological disorders. Given that most of the studies have been performed in controlled experimental settings to ensure the consistency and reproducibility, the impacts of environmental factors, such as dietary perturbation, on the development of animals with different genotypes and the pathogenesis of these diseases remain poorly understood. By analyzing the cdk8 and cyclin C (cycC) mutant larvae in Drosophila , we have previously reported that the CDK8-CycC complex coordinately regulates lipogenesis by repressing dSREBP (sterol regulatory element-binding protein)-activated transcription and developmental timing by activating EcR (ecdysone receptor)-dependent gene expression. Here we report that dietary nutrients, particularly proteins and carbohydrates, modulate the developmental timing through the CDK8/CycC/EcR pathway. We observed that cdk8 and cycC mutants are sensitive to the levels of dietary proteins and seven amino acids (arginine, glutamine, isoleucine, leucine, methionine, threonine, and valine). Those mutants are also sensitive to dietary carbohydrates, and they are more sensitive to monosaccharides than disaccharides. These results suggest that CDK8-CycC mediates the dietary effects on lipid metabolism and developmental timing in Drosophila larvae. Highlights • The cdk8 and cycC mutant Drosophila larvae are hypersensitive to increased levels of dietary proteins. • Of the 20 kinds of amino acids, the cdk8 and cycC mutants are particularly sensitive to the levels of 7 specific amino acids. • Compared to disaccharides, the cdk8 and cycC mutants are more sensitive to increased dietary monosaccharides. • CDK8-CycC may mediate the dietary effects on lipid metabolism and developmental transition in Drosophila larvae. [ABSTRACT FROM AUTHOR]

Nitrate-dependent anaerobic ferrous oxidation (NAFO) is a new and valuable bio-process for the treatment of wastewaters with low C/N ratio, and the NAFO process is in state of the art. The heterotrophic denitrifying sludge (HDS), possessing NAFO activity, was used as bioaugmentation to enhance NAFO efficiency. At a dosage of 6% (V/V), the removal of nitrate and ferrous was 2.4 times and 2.3 times of as primary, and the volumetric removal rate (VRR) of nitrate and ferrous was 2.4 times and 2.2 times of as primary. Tracing experiments of HDS indicated that the bioaugmentation on NAFO reactor was resulted from the NAFO activity by HDS itself. The predominant bacteria in HDS were identified as Thauera (52.5%) and Hyphomicrobium (20.0%) which were typical denitrifying bacteria and had potential ability to oxidize ferrous. In conclusion, HDS could serve as bioaugmentation or a new seeding sludge for operating high-efficiency NAFO reactors. [ABSTRACT FROM AUTHOR]

The steroid hormone ecdysone and its receptor (EcR) play critical roles in orchestrating developmental transitions in arthropods. However, the mechanism by which EcR integrates nutritional and developmental cues to correctly activate transcription remains poorly understood. Here, we show that EcR-dependent transcription, and thus, developmental timing in Drosophila, is regulated by CDK8 and its regulatory partner Cyclin C (CycC), and the level of CDK8 is affected by nutrient availability. We observed that cdk8 and cycC mutants resemble EcR mutants and EcR-target genes are systematically down-regulated in both mutants. Indeed, the ability of the EcR-Ultraspiracle (USP) heterodimer to bind to polytene chromosomes and the promoters of EcR target genes is also diminished. Mass spectrometry analysis of proteins that co-immunoprecipitate with EcR and USP identified multiple Mediator subunits, including CDK8 and CycC. Consistently, CDK8-CycC interacts with EcR-USP in vivo; in particular, CDK8 and Med14 can directly interact with the AF1 domain of EcR. These results suggest that CDK8-CycC may serve as transcriptional cofactors for EcR-dependent transcription. During the larval–pupal transition, the levels of CDK8 protein positively correlate with EcR and USP levels, but inversely correlate with the activity of sterol regulatory element binding protein (SREBP), the master regulator of intracellular lipid homeostasis. Likewise, starvation of early third instar larvae precociously increases the levels of CDK8, EcR and USP, yet down-regulates SREBP activity. Conversely, refeeding the starved larvae strongly reduces CDK8 levels but increases SREBP activity. Importantly, these changes correlate with the timing for the larval–pupal transition. Taken together, these results suggest that CDK8-CycC links nutrient intake to developmental transitions (EcR activity) and fat metabolism (SREBP activity) during the larval–pupal transition. [ABSTRACT FROM AUTHOR]

Post-translational modification of histones plays essential roles in the transcriptional regulation of genes in eukaryotes. Methylation on basic residues of histones is regulated by histone methyltransferases and histone demethylases, and misregulation of these enzymes has been linked to a range of diseases such as cancer. Histone lysine demethylase 2 (KDM2) family proteins have been shown to either promote or suppress tumorigenesis in different human malignancies. However, the roles and regulation of KDM2 in development are poorly understood, and the exact roles of KDM2 in regulating demethylation remain controversial. Since KDM2 proteins are highly conserved in multicellular animals, we analyzed the KDM2 ortholog in Drosophila . We have observed that dKDM2 is a nuclear protein and its level fluctuates during fly development. We generated three deficiency lines that disrupt the dKdm2 locus, and together with 10 transposon insertion lines within the dKdm2 locus, we characterized the developmental defects of these alleles. The alleles of dKdm2 define three phenotypic classes, and the intragenic complementation observed among these alleles and our subsequent analyses suggest that dKDM2 is not required for viability. In addition, loss of dKDM2 appears to have rather weak effects on histone H3 lysine 36 and 4 methylation (H3K36me and H3K4me) in the third instar wandering larvae, and we observed no effect on methylation of H3K9me2, H3K27me2 and H3K27me3 in dKdm2 mutants. Taken together, these genetic, molecular and biochemical analyses suggest that dKDM2 is not required for viability of flies, indicating that dKdm2 is likely redundant with other histone lysine demethylases in regulating normal development in Drosophila . [ABSTRACT FROM AUTHOR]

Inactivation of the Rb tumor suppressor can lead to increased cell proliferation or cell death depending on specific cellular context. Therefore, identification of the interacting pathways that modulate the effect of Rb loss will provide novel insights into the roles of Rb in cancer development and promote new therapeutic strategies. Here, we identify a novel synthetic lethal interaction between Rb inactivation and deregulated Wg/Wnt signaling through unbiased genetic screens. We show that a weak allele of axin, which deregulates Wg signaling and increases cell proliferation without obvious effects on cell fate specification, significantly alters metabolic gene expression, causes hypersensitivity to metabolic stress induced by fasting, and induces synergistic apoptosis with mutation of fly Rb ortholog, rbf. Furthermore, hyperactivation of Wg signaling by other components of the Wg pathway also induces synergistic apoptosis with rbf. We show that hyperactivated Wg signaling significantly increases TORC1 activity and induces excessive energy stress with rbf mutation. Inhibition of TORC1 activity significantly suppressed synergistic cell death induced by hyperactivated Wg signaling and rbf inactivation, which is correlated with decreased energy stress and decreased induction of apoptotic regulator expression. Finally the synthetic lethality between Rb and deregulated Wnt signaling is conserved in mammalian cells and that inactivation of Rb and APC induces synergistic cell death through a similar mechanism. These results suggest that elevated TORC1 activity and metabolic stress underpin the evolutionarily conserved synthetic lethal interaction between hyperactivated Wnt signaling and inactivated Rb tumor suppressor. [ABSTRACT FROM AUTHOR]

Abstract: Microscopic observation and settling test were carried out to investigate the structure, density and settlability of anammox granules taken from a high-rate upflow anaerobic sludge blanket (UASB) reactor. The results showed that the anammox granules were irregular in shape and uneven on surface, and their structure included granule, subunit, microbial cell cluster and single cell. The gas pockets were often observed in the anammox granules, and they originated from the obstruction of gas tunnel by extracellular polymer substances (EPSs) and the inflation of produced dinitrogen gas. The volume of gas pockets became larger with the increasing diameter of anammox granules, which led to the decreasing density and the floatation of anammox granules. The diameter of anammox granules should be controlled at less than 2.20mm to avoid the granule floatation. A hypothesized mechanism for the granulation and floatation of anammox biomass was proposed. [Copyright &y& Elsevier]

Altered lipid metabolism underlies several major human diseases, including obesity and type 2 diabetes. However, lipid metabolism pathophysiology remains poorly understood at the molecular level. Insulin is the primary stimulator of hepatic lipogenesis through activation of the SREBP-1c transcription factor. Here we identified cyclin-dependent kinase 8 (CDK8) and its regulatory partner cyclin C (CycC) as negative regulators of the lipogenic pathway in Drosophila, mammalian hepatocytes, and mouse liver. The inhibitory effect of CDK8 and CycC on de novo lipogenesis was mediated through CDK8 phosphorylation of nuclear SREBP-1c at a conserved threonine residue. Phosphorylation by CDK8 enhanced SREBP-1c ubiquitination and protein degradation. Importantly, consistent with the physiologic regulation of lipid biosynthesis, CDK8 and CycC proteins were rapidly downregulated by feeding and insulin, resulting in decreased SREBP-1c phosphorylation. Moreover, overexpression of CycC efficiently suppressed insulin and feeding-induced lipogenic gene expression. Taken together, these results demonstrate that CDK8 and CycC function as evolutionarily conserved components of the insulin signaling pathway in regulating lipid homeostasis. [ABSTRACT FROM AUTHOR]

Summary: Epigenetic regulation of gene expression by histone-modifying corepressor complexes is central to normal animal development. The NAD+-dependent deacetylase and gene repressor SIRT1 removes histone H4K16 acetylation marks and facilitates heterochromatin formation. However, the mechanistic contribution of SIRT1 to epigenetic regulation at euchromatic loci and whether it acts in concert with other chromatin-modifying activities to control developmental gene expression programs remain unclear. We describe here a SIRT1 corepressor complex containing the histone H3K4 demethylase LSD1/KDM1A and several other LSD1-associated proteins. SIRT1 and LSD1 interact directly and play conserved and concerted roles in H4K16 deacetylation and H3K4 demethylation to repress genes regulated by the Notch signaling pathway. Mutations in Drosophila SIRT1 and LSD1 orthologs result in similar developmental phenotypes and genetically interact with the Notch pathway in Drosophila. These findings offer new insights into conserved mechanisms of epigenetic gene repression and regulation of development by SIRT1 in metazoans. [Copyright &y& Elsevier]

Cyclin-dependent kinase 8 (CDK8) and its regulatory partner Cyclin C (CycC) play conserved roles in modulating RNA polymerase II (Pol II)-dependent gene expression. To understand the structure and function relations of CDK8, we analyzed the structures of human and Drosophila CDK8 proteins using molecular dynamics simulations, combined with functional analyses in Drosophila. Specifically, we evaluated the structural differences between hCDK8 and dCDK8 to predict the effects of the LXXLL motif mutation (AQKAA), the P154L mutations, and drug binding on local structures of the CDK8 proteins. First, we have observed that both the LXXLL motif and the kinase activity of CDK8 are required for the normal larval-to-pupal transition in Drosophila. Second, our molecular dynamic analyses have revealed that hCDK8 has higher hydrogen bond occupation of His149-Asp151 and Asp151-Asn156 than dCDK8. Third, the substructure of Asp282, Phe283, Arg285, Thr287 and Cys291 can distinguish human and Drosophila CDK8 structures. In addition, there are two hydrogen bonds in the LXXLL motif: a lower occupation between L312 and L315, and a relatively higher occupation between L312 and L316. Human CDK8 has higher hydrogen bond occupation between L312 and L316 than dCDK8. Moreover, L312, L315 and L316 in the LXXLL motif of CDK8 have the specific pattern of hydrogen bonds and geometries, which could be crucial for the binding to nuclear receptors. Furthermore, the P154L mutation dramatically decreases the hydrogen bond between L312 and L315 in hCDK8, but not in dCDK8. The mutations of P154L and AQKAA modestly alter the local structures around residues 154. Finally, we identified the inhibitor-induced conformational changes of hCDK8, and our results suggest a structural difference in the drug-binding site between hCDK8 and dCDK8. Taken together, these results provide the structural insights into the roles of the LXXLL motif and the kinase activity of CDK8 in vivo. [ABSTRACT FROM AUTHOR]

The sterol regulatory element-binding protein (SREBP) transcription factor family is a critical regulator of lipid and sterol homeostasis in eukaryotes. In mammals, SREBPs are highly active in the fed state to promote the expression of lipogenic and cholesterogenic genes and facilitate fat storage. During fasting, SREBP-dependent lipid/cholesterol synthesis is rapidly diminished in the mouse liver; however, the mechanism has remained incompletely understood. Moreover, the evolutionary conservation of fasting regulation of SREBP-dependent programs of gene expression and control of lipid homeostasis has been unclear. We demonstrate here a conserved role for orthologs of the NAD+-dependent deacetylase SIRT1 in metazoans in down-regulation of SREBP orthologs during fasting, resulting in inhibition of lipid synthesis and fat storage. Our data reveal that SIRT1 can directly deacetylate SREBP, and modulation of SIRT1 activity results in changes in SREBP ubiquitination, protein stability, and target gene expression. In addition, chemical activators of SIRT1 inhibit SREBP target gene expression in vitro and in vivo, correlating with decreased hepatic lipid and cholesterol levels and attenuated liver steatosis in diet-induced and genetically obese mice. We conclude that SIRT1 orthologs play a critical role in controlling SREBP-dependent gene regulation governing lipid/cholesterol homeostasis in metazoans in response to fasting cues. These findings may have important biomedical implications for the treatment of metabolic disorders associated with aberrant lipid/cholesterol homeostasis, including metabolic syndrome and atherosclerosis. [ABSTRACT FROM AUTHOR]