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Poly (ADP-ribose) polymerase-1 (PARP-1) is a potential target for the discovery of chemosensitizers and anticancer drugs. Amentoflavone (AMF) is reported to be a selective PARP-1 inhibitor. Here, structural modifications and trimming of AMF have led to a series of AMF derivatives (9a-h) and apigenin-piperazine/piperidine hybrids (14a-p, 15a-p, 17a-h, and 19a-f), respectively. Among these compounds, 15l exhibited a potent PARP-1 inhibitory effect (IC50 = 14.7 nM) and possessed high selectivity to PARP-1 over PARP-2 (61.2-fold). Molecular dynamics simulation and the cellular thermal shift assay revealed that 15l directly bound to the PARP-1 structure. In in vitro and in vivo studies, 15l showed a potent chemotherapy sensitizing effect against A549 cells and a selective cytotoxic effect toward SK-OV-3 cells through PARP-1 inhibition. 15l·2HCl also displayed good ADME characteristics, pharmacokinetic parameters, and a desirable safety margin. These findings demonstrated that 15l·2HCl may serve as a lead compound for chemosensitizers and the (BRCA-1)-deficient cancer therapy.

Although overexpression of EZH2, a catalytic subunit of the polycomb repressive complex 2 (PRC2), is an eminent feature of various cancers, the regulation of its abundance and function remains insufficiently understood. We report here that the PRC2 complex is physically associated with ubiquitin-specific protease USP7 in cancer cells where USP7 acts to deubiquitinate and stabilize EZH2. Interestingly, we found that USP7-catalyzed H2BK120ub1 deubiquitination is a prerequisite for chromatin loading of PRC2 thus H3K27 trimethylation, and this process is not affected by H2AK119 ubiquitination catalyzed by PRC1. Genome-wide analysis of the transcriptional targets of the USP7/PRC2 complex identified a cohort of genes including FOXO1 that are involved in cell growth and proliferation. We demonstrated that the USP7/PRC2 complex drives cancer cell proliferation and tumorigenesis in vitro and in vivo. We showed that the expression of both USP7 and EZH2 elevates during tumor progression, corresponding to a diminished FOXO1 expression, and the level of the expression of USP7 and EZH2 strongly correlates with histological grades and prognosis of tumor patients. These results reveal a dual role for USP7 in the regulation of the abundance and function of EZH2, supporting the pursuit of USP7 as a therapeutic target for cancer intervention.

SummaryRegulation of chromatin accessibility determines the transcription activities of genes, which endow the host with function-specific gene expression patterns. It remains unclear how chromatin accessibility is specifically directed, particularly, during host defense against viral infection. We previously reported that the nuclear matrix protein SAFA surveils viral RNA and regulates antiviral immune genes expression. However, how SAFA regulates the expression and what determines the specificity of antiviral immune genes remains unknown. Here, we identified that the depletion of SAFA specifically decreased the chromatin accessibility, activation and expression of virus induced genes in a genome-wide scale after VSV infection. SAFA exclusively bound with antiviral related RNAs, which mediated the specific opening of the according chromatin and robust transcription of these genes. Knockdown of these associated RNAs dampened the accessibility of corresponding genes in an extranuclear signaling pathway dependent manner. Moreover, VSV infection cleaved SAFA protein at the C-terminus which deprived its RNA binding ability for immune evasion. Thus, our results demonstrated that SAFA and the interacting RNA products during viral infection collaborate and remodel chromatin accessibility to facilitate antiviral innate immune response.

The scope and variety of the metabolic intermediates from the mitochondrial tricarboxylic acid (TCA) cycle that are engaged in epigenetic regulation of the chromatin function in the nucleus raise an outstanding question about how timely and precise supply/consumption of these metabolites is achieved in the nucleus. We report here the identification of a nonclassical TCA cycle in the nucleus (nTCA cycle). We found that all the TCA cycle-associated enzymes including citrate synthase (CS), aconitase 2 (ACO2), isocitrate dehydrogenase 3 (IDH3), oxoglutarate dehydrogenase (OGDH), succinyl-CoA synthetase (SCS), fumarate hydratase (FH), and malate dehydrogenase 2 (MDH2), except for succinate dehydrogenase (SDH), a component of electron transport chain for generating ATP, exist in the nucleus. We showed that these nuclear enzymes catalyze an incomplete TCA cycle similar to that found in cyanobacteria. We propose that the nTCA cycle is implemented mainly to generate/consume metabolic intermediates, not for energy production. We demonstrated that the nTCA cycle is intrinsically linked to chromatin dynamics and transcription regulation. Together, our study uncovers the existence of a nonclassical TCA cycle in the nucleus that links the metabolic pathway to epigenetic regulation.

Owing to the stable crystal structure and wide range of pH applications, CuFe2O4 particles have been intensively concerned in the field of advanced oxidation, but their serious agglomeration and slow catalytic efficiency are still the stumbling blocks. The composite catalyst (EG-CuFe2O4-U) prepared by urea-assisted hydrothermal method with expanded graphite (EG) as the substrate immobilized CuFe2O4 not only exposed more active sites but also exhibited a higher electron transfer rate. Additionally, EG-CuFe2O4-U showed excellent performance in degrading tetracycline (TC) in model wastewater by activated peroxydisulfate (PDS). The synthesis mechanism of EG-CuFe2O4-U and the principle of urea in the formation of reduction environment were discussed in detail by the experimental results of key preparation parameters and characterization. Meanwhile, several critical influencing factors were examined including PDS concentration, catalyst dosage, initial pH of the solution, and the change of pH in different systems. Furthermore, the removal efficiency and mineralization efficiency of TC (50 ppm) exceed 91% and 34.6%, on the conditions of 0.4 gL−1 EG-CuFe2O4-U, 6 mM PDS, initial solution pH of 4, and room temperature. What’s more, the internal reaction mechanism of free radicals and non-free radicals in the EG-CuFe2O4-U/PDS system was further elaborated via scavenging tests, electron paramagnetic resonance (EPR). Finally, based on twenty-one principal intermediates of TC, four possible degradation pathways were proposed. In general, the catalyst with a rich pore structure and high catalytic activity has great potential in the effective activation of PDS and is prospective to be further applied in the field of antibiotic wastewater degradation.

Large-scale crotonylome reveals CDYL-regulated crotonylation in diverse functions including RPA-mediated HR DNA repair., Previously, we reported that chromodomain Y–like (CDYL) acts as a crotonyl–coenzyme A hydratase and negatively regulates histone crotonylation (Kcr). However, the global CDYL-regulated crotonylome remains unclear. Here, we report a large-scale proteomics analysis for protein Kcr. We identify 14,311 Kcr sites across 3734 proteins in HeLa cells, providing by far the largest crotonylome dataset. We show that depletion of CDYL alters crotonylome landscape affecting diverse cellular pathways. Specifically, CDYL negatively regulated Kcr of RPA1, and mutation of the Kcr sites of RPA1 impaired its interaction with single-stranded DNA and/or with components of resection machinery, supporting a key role of RPA1 Kcr in homologous recombination DNA repair. Together, our study indicates that protein crotonylation has important implication in various pathophysiological processes.

Copy number gain in chromosome 8q21 is frequently detected in breast cancer, yet the oncogenic potential underlying this amplicon in breast carcinogenesis remains to be delineated. We report here that ZNF704, a gene mapped to 8q21, is recurrently amplified in various malignancies including breast cancer. ZNF704 acted as a transcriptional repressor and interacted with the transcriptional corepressor SIN3A complex. Genome-wide interrogation of transcriptional targets revealed that the ZNF704/SIN3A complex represses a panel of genes including PER2 that are critically involved in the function of the circadian clock. Overexpression of ZNF704 prolonged the period and dampened the amplitude of the circadian clock. ZNF704 promoted the proliferation and invasion of breast cancer cells in vitro and accelerated the growth and metastasis of breast cancer in vivo. Consistently, the level of ZNF704 expression inversely correlated with that of PER2 in breast carcinomas, and high level of ZNF704 correlated with advanced histologic grades, lymph node positivity, and poor prognosis of patients with breast cancer, especially those with HER2+ and basal-like subtypes. These results indicate that ZNF704 is an important regulator of the circadian clock and a potential driver for breast carcinogenesis.Significance:This study indicates that ZNF704 could be a potential oncogenic factor, disrupting circadian rhythm of breast cancer cells and contributing to breast carcinogenesis.

The transition of oxidized 5-methylcytosine (5mC) intermediates into the base excision repair (BER) pipeline to complete DNA demethylation remains enigmatic. We report here that UHRF2, the only paralog of UHRF1 in mammals that fails to rescue Uhrf1-/- phenotype, is physically and functionally associated with BER complex. We show that UHRF2 is allosterically activated by 5-hydroxymethylcytosine (5hmC) and acts as a ubiquitin E3 ligase to catalyze K33-linked polyubiquitination of XRCC1. This nonproteolytic action stimulates XRCC1's interaction with the ubiquitin binding domain-bearing RAD23B, leading to the incorporation of TDG into BER complex. Integrative epigenomic analysis in mouse embryonic stem cells reveals that Uhrf2-fostered TDG-RAD23B-BER complex is functionally linked to the completion of DNA demethylation at active promoters and that Uhrf2 ablation impedes DNA demethylation on latent enhancers that undergo poised-to-active transition during neuronal commitment. Together, these observations highlight an essentiality of 5hmC-switched UHRF2 E3 ligase activity in commissioning the accomplishment of active DNA demethylation.

Copy number gain in chromosome 8q21 is considered as the prototype of genetic abnormalities associated with development of breast cancer, yet the oncogenic potential underlying this amplicon in breast carcinogenesis remains to be delineated. We report here thatZNF704, a gene mapped to 8q21, is recurrently amplified in various malignancies including breast cancer. We found that ZNF704 acts as transcription repressor and interacts with the transcription corepressor SIN3A complex. Genome-wide interrogation of the transcriptional targets identifies that the ZNF704/SIN3A complex represses a panel of genes includingPER2that are critically involved in the function of circadian clock. Indeed, ZNF704 overexpression prolongs the period and dampens the amplitude of circadian clock. We showed that ZNF704 promotes the proliferation and invasion of breast cancer cellsin vitroand accelerates the growth and metastasis of breast cancerin vivo. Consistently, the level of ZNF704 expression is inversely correlated with that of PER2 in breast carcinomas, and high level of ZNF704 correlates with advanced histological grades, lymph node positivity, and poor prognosis of breast cancer patients, especially those with HER2+and basal-like subtypes. These results indicate that ZNF704 is an important regulator of circadian clock and a potential driver for breast carcinogenesis.

The pathophysiological function of the forkhead transcription factor FOXN3 remains to be explored. Here we report that FOXN3 is a transcriptional repressor that is physically associated with the SIN3A repressor complex in estrogen receptor-positive (ER+) cells. RNA immunoprecipitation-coupled high-throughput sequencing identified that NEAT1, an estrogen-inducible long noncoding RNA, is required for FOXN3 interactions with the SIN3A complex. ChIP-Seq and deep sequencing of RNA genomic targets revealed that the FOXN3-NEAT1-SIN3A complex represses genes including GATA3 that are critically involved in epithelial-to-mesenchymal transition (EMT). We demonstrated that the FOXN3-NEAT1-SIN3A complex promotes EMT and invasion of breast cancer cells in vitro as well as dissemination and metastasis of breast cancer in vivo. Interestingly, the FOXN3-NEAT1-SIN3A complex transrepresses ER itself, forming a negative-feedback loop in transcription regulation. Elevation of both FOXN3 and NEAT1 expression during breast cancer progression corresponded to diminished GATA3 expression, and high levels of FOXN3 and NEAT1 strongly correlated with higher histological grades and poor prognosis. Our experiments uncovered that NEAT1 is a facultative component of the SIN3A complex, shedding light on the mechanistic actions of NEAT1 and the SIN3A complex. Further, our study identified the ERα-NEAT1-FOXN3/NEAT1/SIN3A-GATA3 axis that is implicated in breast cancer metastasis, providing a mechanistic insight into the pathophysiological function of FOXN3.

UTX is implicated in embryonic development and lineage specification. However, how this X-linked histone demethylase contributes to the occurrence and progression of breast cancer remains to be clarified. Here we report that UTX is physically associated with estrogen receptor (ER) and functions in ER-regulated transcription. We showed that UTX coordinates with JHDM1D and CBP to direct H3K27 methylation-acetylation transition and to create a permissive chromatin state on ER targets. Genome-wide analysis of the transcriptional targets of UTX by ChIP-seq identified a set of genes such as chemokine receptor CXCR4 that are intimately involved in breast cancer tumorigenesis and metastasis. We demonstrated that UTX promotes the proliferation and migration of ER+ breast cancer cells. Interestingly, UTX itself is transactivated by ER, forming a feed-forward loop in the regulation of hormone response. Indeed, UTX is upregulated during ER+ breast cancer progression, and the expression level of UTX is positively correlated with that of CXCR4 and negatively correlated with the overall survival of ER+ breast cancer patients. Our study identified a feed-forward loop between UTX and ER in the regulation of hormonally responsive breast carcinogenesis, supporting the pursuit of UTX as an emerging therapeutic target for the intervention of certain ER+ breast cancer with specific epigenetic vulnerability.

Background Panax notoginseng (Burk.) F.H. Chen is a traditional medicinal plant widely used to prevent and treat cardiovascular diseases. Ginsenoside Rd (GRd) is a major bioactive component of P. notoginseng, but specific effects on cardiovascular disease-related pathogenic processes are rarely studied, especially vascular endothelial injury. Purpose This study investigated the potential protective efficacy of GRd against nicotine-induced vascular endothelial cell injury, disruption of vascular nitric oxide (NO) signaling, aberrant endothelium–monocyte adhesion, platelet aggregation, and vasoconstriction. Study design/Methods Vascular endothelial injury and functional disruption were investigated in cultured human umbilical vein endothelial cells (HUVECs) by biochemical assays for nitric oxide (NO) and angiotensin II (Ang II), immunofluorescence (IF) and western blotting for expression analyses of apoptosis- related proteins, endothelial nitric oxide synthase (eNOS), inducible nitric oxide synthase (iNOS), Ang II type receptor 1 (AGTR1), toll-like receptor 4 (TLR4), myeloid differentiation factor 88 (MyD88), and nuclear factor-kappa B (NF-κB). In addition, vascular protection by GRd was examined in nicotine-administered Sprague-Dawley (SD) rats by serum NO and Ang II assays, and by hematoxylin-eosin (HE) and immunostaining of aorta. We also examined effects of GRd on monocyte (THP-1 cells) adhesion assays, adenosine diphosphate (ADP)-induced platelet aggregation, and phenylephrine (PE)-induced vasoconstriction of isolated rat aortic rings. Results In HUVECs, nicotine significantly suppressed NO production, enhanced Ang II production, downregulated eNOS expression, and upregulated expression levels of AGTR1, TLR4, MyD88, NF-κB, iNOS, Bax/Bcl-2 ratio, cleaved caspase-3, and cytochrome c (cyt c). All of these changes were significantly reversed by GRd. In rats, oral GRd reversed the reduction NO and enhanced Ang II production in serum induced by nicotine administration, and HE staining revealed protection of aortic endothelial cells. In addition, GRd reversed nicotine-mediated enhancement of HUVECs-monocyte adhesion, inhibited ADP-induced platelet aggregation and PE-induced vasoconstriction. Conclusion GRd may prevent nicotine-induced cardiovascular diseases by preserving normal vascular endothelial NO signaling, suppressing platelet aggregation and vasoconstriction, and by preventing endothelial cell–monocyte adhesion.

Although overexpression of EZH2, a catalytic subunit of the polycomb repressive complex 2 (PRC2), is an eminent feature of various cancers, the regulation of its abundance and function remains insufficiently understood. We report here that the PRC2 complex is physically associated with ubiquitin-specific protease USP7 in melanoma cells where USP7 acts to deubiquitinate and stabilize EZH2. Interestingly, we found that USP7-catalyzed H2BK120 deubiquitination is a prerequisite for chromatin loading of PRC2 thus H3K27 trimethylation. Genome-wide analysis of the transcriptional targets of the USP7/PRC2 complex identified a cohort of genes including FOXO1 that are involved in cell growth and proliferation. We demonstrated that the USP7/PRC2 complex drives melanoma cell proliferation and tumorigenesis in vitro and in vivo. We showed that the expression of both USP7 and EZH2 elevates during melanoma progression, corresponding to a diminished FOXO1 expression, and the level of the expression of USP7 and EZH2 strongly correlates with histological grades and prognosis of melanoma patients. These results reveal a dual role for USP7 in the regulation of the abundance and function of EZH2, supporting the pursuit of USP7 as a therapeutic target for melanoma.

The interconversion of guanosine triphosphate (GTP) and guanosine diphosphate (GDP) is integral to a wide variety of biological cellular activities. However, analytical methods which directly detect the ratio of intracellular GTP and GDP concentrations have not been available. Herein, we report GNEPS, a genetically encoded fluorescent sensor that enables real-time monitoring of the GTP:GDP ratio, which is a fusion protein comprising a eukaryotic G-protein and a circularly permuted yellow fluorescent protein. GNEPS has distinct fluorescence spectra between its GTP-bound and GDP-bound states. Its apparent fluorescence signal therefore depends upon the competitive binding of GTP and GDP. Live cell imaging experiments demonstrated that GNEPS can be used to monitor spatiotemporal changes in the intracellular GTP:GDP ratio in various cell types and organelles in response to metabolic perturbations. We anticipate that GNEPS will become a valuable tool for understanding the metabolic and regulatory contributions of guanosine nucleotides.

Amentoflavone, a kind of biflavonoid existing in several medicinal plants such as Selaginella moellendorfi and Gingko biloba, possesses anti-inflammatory, antioxidant, anti-virus, anti-tumor activities. In the present study, a new reliable and sensitive UHPLC-ESI-MS/MS method was developed to determine the permeability of amentoflavone under different conditions, and its metabolites in Caco-2 cells were identified by means of UHPLC-Q-TOF-MS/MS method. The results showed that amentoflavone could be considered as a compound with moderate intestinal absorption in Caco-2 cell model and its absorption characteristics might be involved in paracellular passive penetration and clathrin-mediated endocytosis with no participation of efflux transporters. Eight metabolites of amentoflavone were identified in Caco-2 cell model, indicating that the main metabolic pathways were oxidation, reduction, methylation and glucuronide conjugation. This study can provide valuable evidence for an in-depth understanding of absorption mechanism and transformation of amentoflavone in the intestine.

Hyperactivation of EGFR/PI3K/AKT is a prominent feature of various human cancers. Thus, understanding how this molecular cascade is balanced is of great importance. We report here that the ubiquitin-specific protease USP43 is physically associated with the chromatin remodeling NuRD complex and catalyzes H2BK120 deubiquitination. Functionally this coordinates the NuRD complex to repress a cohort of genes, including EGFR, which are critically involved in cell proliferation and carcinogenesis. We show that USP43 strongly suppresses the growth and metastasis of breast cancer in vivo. Interestingly, USP43 also exists in the cytoplasm, where it is phosphorylated by AKT, enabling its binding to the 14-3-3β/e heterodimer and sequestration in the cytoplasm. Significantly, hyperactivation of EGFR/PI3K/AKT in breast cancer is associated with the cytoplasmic retention of USP43 and thus, the inhibition of its transcriptional regulatory function. Moreover, cancer-associated mutations of USP43 affect its subcellular localization and/or epigenetic regulatory functions. Nuclear USP43 is significantly reduced in breast carcinomas and is associated with EGFR accumulation and AKT hyperactivation. A low level of nuclear USP43 correlates with higher histologic grades and poor prognosis. Our study identifies USP43 to be an H2BK120 deubiquitinase and a potential tumor suppressor and reveals a reciprocally inhibitory loop between USP43 and EGFR/PI3K/AKT, whose imbalance drives breast carcinogenesis.

An ideal separator is essential for efficient power production from air-cathode single-chamber microbial fuel cells (MFCs). In this study, we use different kinds of membranes as separators, including Nafion 117 proton exchange membrane, polyethersulfone and poly(vinylidene fluoride) microfiltration membranes. Temporal variations of cathode performance are monitored during the experiment. Results show that MFCs with microfiltration membranes present higher power output but deterioration is still observed after about 600-h operation. With the utilization of appropriate separators (e.g., polyethersulfone membrane), biofouling, cation fouling and chemical scale fouling of the cathodes are alleviated while reaction fouling seems inevitable. Moreover, it is found that Coulombic efficiency (CE) and energy efficiency (EE) are also related to the cathode performance. Despite relatively high oxygen diffusivity (1.49 × 10 −5 cm 2 s −1 ), CE and EE of the MFC with 0.1 μm pore-size polyethersulfone membrane can reach 92.8% and 13.7%, respectively, when its average power density registers 403.5 mW m −2 . This phenomenon might be attributed to the finding that the overall substrate consumption rate due to oxygen reduction and respiration is almost constant in the air-cathode MFCs. Oxygen leakage into the electrolyte can be inhibited due to the efficient oxygen reduction reaction on the surface of the cathode.

Background Melanoma, originated from melanocytes located on the basal membrane of the epithelial tissue, is the most aggressive form of skin cancer that accounts for 75% of skin cancer-related death. Although it is believed that BRAF mutation and the mitogen-activated protein kinase (MAPK) pathway play critical roles in the pathogenesis of melanoma, how the MAPK signaling is regulated in melanoma carcinogenesis is still not fully understood. Methods We characterized JMJD6 expression in melanoma tissue array by immunohistochemistry analysis. We used human melanoma A375, 451Lu and SK-MEL-1 cell lines for in vitro proliferation and invasion experiments, and xenograft transplanted mice using murine melanoma B16F10 cells by bioluminescence imaging for in vivo tumor growth and pulmonary metastasis assessments. Endothelial tube formation assay, chicken yolk sac membrane assay and matrigel plug assay were performed to test the effect of JMJD6 on the angiogenic potential in vitro and in vivo. Results Here we report that the jumonji C domain-containing demethylase/hydroxylase JMJD6 is markedly up-regulated in melanoma. We found that high expression of JMJD6 is closely correlated with advanced clinicopathologic stage, aggressiveness, and poor prognosis of melanoma. RNA-seq showed that knockdown of JMJD6 affects the alternative splicing of a panel of transcripts including that encoding for PAK1, a key component in MAPK signaling pathway. We demonstrated that JMJD6 enhances the MAPK signaling and promotes multiple cellular processes including melanogenesis, proliferation, invasion, and angiogenesis in melanoma cells. Interestingly, JMJD6 is transcriptionally activated by c-Jun, generating a feedforward loop to drive the development and progression of melanoma. Conclusions Our results indicate that JMJD6 is critically involved in melanoma carcinogenesis, supporting the pursuit of JMJD6 as a potential biomarker for melanoma aggressiveness and a target for melanoma intervention. Electronic supplementary material The online version of this article (10.1186/s12943-017-0744-2) contains supplementary material, which is available to authorized users.

EGFR is required for animal development, and dysregulation of EGFR is critically implicated in malignant transformation. However, the molecular mechanism underlying the regulation of EGFR expression remains poorly explored. Here we report that the zinc-finger protein ZNF516 is a transcription repressor. ZNF516 is physically associated with the CtBP/LSD1/CoREST complex and transcriptionally represses a cohort of genes including EGFR that are critically involved in cell proliferation and motility. We demonstrate that the ZNF516–CtBP/LSD1/CoREST complex inhibits the proliferation and invasion of breast cancer cells in vitro and suppresses breast cancer growth and metastasis in vivo. Significantly, low expression of ZNF516 is positively associated with advanced pathological staging and poor survival of breast carcinomas. Our data indicate that ZNF516 is a transcription repressor and a potential suppressor of EGFR, adding to the understanding of EGFR-related breast carcinogenesis and supporting the pursuit of ZNF516 as a potential therapeutic target for breast cancer., EGFR is a well-known oncogene; however, the mechanisms regulating its expression are still unclear. Here, analysing genome-wide chromatin associations, the authors show that in breast cancer cells ZNF516 represses EGFR transcription through the interaction with the CtBP/LSD1/CoREST complex.

The de novo assembly and post-splicing reassembly of the U4/U6.U5 tri-snRNP remain to be investigated. We report here that ZIP, a protein containing a CCCH-type zinc finger and a G-patch domain, as characterized by us previously, regulates pre-mRNA splicing independent of RNA binding. We found that ZIP physically associates with the U4/U6.U5 tri-small nuclear ribonucleoprotein (tri-snRNP). Remarkably, the ZIP-containing tri-snRNP, which has a sedimentation coefficient of ∼35S, is a tri-snRNP that has not been described previously. We also found that the 35S tri-snRNP contains hPrp24, indicative of a state in which the U4/U6 di-snRNP is integrating with the U5 snRNP. We found that the 35S tri-snRNP is enriched in the Cajal body, indicating that it is an assembly intermediate during 25S tri-snRNP maturation. We showed that the 35S tri-snRNP also contains hPrp43, in which ATPase/RNA helicase activities are stimulated by ZIP. Our study identified, for the first time, a tri-snRNP intermediate, shedding new light on the de novo assembly and recycling of the U4/U6.U5 tri-snRNP.

Alternative splicing events in MAPK pathway. (DOCX 19Â kb)

Correlation between JMJD6 expression and clinicopathologic characteristics of 88 melanoma patients by chi-square test. (DOCX 503Â kb)

Faithful transmission or restoration of epigenetic information such as repressive histone modifications through generations is critical for the maintenance of cell identity. We report here that chromodomain Y-like protein (CDYL), a chromodomain-containing transcription corepressor, is physically associated with chromatin assembly factor 1 (CAF-1) and the replicative helicase MCM complex. We showed that CDYL bridges CAF-1 and MCM, facilitating histone transfer and deposition during DNA replication. We demonstrated that CDYL recruits histone-modifying enzymes G9a, SETDB1, and EZH2 to replication forks, leading to the addition of H3K9me2/3 and H3K27me2/3 on newly deposited histone H3. Significantly, depletion of CDYL impedes early S phase progression and sensitizes cells to DNA damage. Our data indicate that CDYL plays an important role in the transmission/restoration of repressive histone marks, thereby preserving the epigenetic landscape for the maintenance of cell identity.

FOXA1 functions in epigenetic reprogramming and is described as a 'pioneer factor'. However, exactly how FOXA1 achieves these remarkable biological functions is not fully understood. Here we report that FOXA1 associates with DNA repair complexes and is required for genomic targeting of DNA polymerase β (POLB) in human cells. Genome-wide DNA methylomes demonstrate that the FOXA1 DNA repair complex is functionally linked to DNA demethylation in a lineage-specific fashion. Depletion of FOXA1 results in localized reestablishment of methylation in a large portion of FOXA1-bound regions, and the regions with the most consistent hypermethylation exhibit the greatest loss of POLB and are represented by active promoters and enhancers. Consistently, overexpression of FOXA1 commits its binding sites to active DNA demethylation in a POLB-dependent manner. Finally, FOXA1-associated DNA demethylation is tightly coupled with estrogen receptor genomic targeting and estrogen responsiveness. Together, these results link FOXA1-associated DNA demethylation to transcriptional pioneering by FOXA1.

Summary Produced water from polymer flooding (polymer-produced water) is difficult to treat by the conventional gravity settling process. A new type of double-cone air-sparged hydrocyclone (DcASH) has been designed, and its fundamental structure and operating principles are introduced in this paper. Experimental research on treating produced water from polymer flooding has been carried out, showing that the DcASH has a high treatment capacity. The oil concentration of treated water was less than 100 mg/L, which satisfied the requirements of the next deep-bed filtration process stage. Compared to gravity settling, conventional hydrocyclones, and flotation, the DcASH has a higher separation efficiency, which indicates that DcASH will have good application prospects in oilfield produced-water treatment.

Lysine crotonylation (Kcr) is a newly identified histone modification that is associated with active transcription in mammalian cells. Here we report that the chromodomain Y-like transcription corepressor CDYL negatively regulates histone Kcr by acting as a crotonyl-CoA hydratase to convert crotonyl-CoA to β-hydroxybutyryl-CoA. We showed that the negative regulation of histone Kcr by CDYL is intrinsically linked to its transcription repression activity and functionally implemented in the reactivation of sex chromosome-linked genes in round spermatids and genome-wide histone replacement in elongating spermatids. Significantly, Cdyl transgenic mice manifest dysregulation of histone Kcr and reduction of male fertility with a decreased epididymal sperm count and sperm cell motility. Our study uncovers a biochemical pathway in the regulation of histone Kcr and implicates CDYL-regulated histone Kcr in spermatogenesis, adding to the understanding of the physiology of male reproduction and the mechanism of the spermatogenic failure in AZFc (Azoospermia Factor c)-deleted infertile men.

Dendritic arborization is one of the key determinants of precise circuits for information processing in neurons. Unraveling the molecular mechanisms underlying dendrite morphogenesis is critical to understanding the establishment of neuronal connections. Here, using gain- and loss-of-function approaches, we defined the chromodomain protein and transcription corepressor chromodomain Y-like (CDYL) protein as a negative regulator of dendrite morphogenesis in rat/mouse hippocampal neurons bothin vitroandin vivo. Overexpressing CDYL decreased, whereas knocking it down increased, the dendritic complexity of the primary cultured rat neurons. High-throughput DNA microarray screening identified a number of CDYL downstream target genes, including the brain-derived neurotrophic factor (BDNF). Knock-down of CDYL in neuronal cells led to increased expression of BDNF, which is primarily responsible for CDYL's effects on dendrite patterns. Mechanistically, CDYL interacts with EZH2, the catalytic subunit of Polycomb Repressive Complex 2 (PRC2), directly and recruits the H3K27 methyltransferase activity to the promoter region of the BDNF gene. In doing so, CDYL and EZH2 coordinately restrict dendrite morphogenesis in an interdependent manner. Finally, we found that neural activity increased dendritic complexity through degradation of CDYL protein to unleash its inhibition on BDNF. These results link, for the first time, the epigenetic regulators CDYL and EZH2 to dendrite morphogenesis and might shed new light on our understanding of the regulation of the neurodevelopment.

p53 hydroxylation by JMJD6 represents a novel post-translational modification for p53. JMJD6-mediated hydroxylation regulates p53's transcriptional activity and the p53-dependent control of colon cancer., Jumonji domain-containing 6 (JMJD6) is a member of the Jumonji C domain-containing family of proteins. Compared to other members of the family, the cellular activity of JMJD6 is still not clearly defined and its biological function is still largely unexplored. Here we report that JMJD6 is physically associated with the tumor suppressor p53. We demonstrated that JMJD6 acts as an α-ketoglutarate– and Fe(II)-dependent lysyl hydroxylase to catalyze p53 hydroxylation. We found that p53 indeed exists as a hydroxylated protein in vivo and that the hydroxylation occurs mainly on lysine 382 of p53. We showed that JMJD6 antagonizes p53 acetylation, promotes the association of p53 with its negative regulator MDMX, and represses transcriptional activity of p53. Depletion of JMJD6 enhances p53 transcriptional activity, arrests cells in the G1 phase, promotes cell apoptosis, and sensitizes cells to DNA damaging agent-induced cell death. Importantly, knockdown of JMJD6 represses p53-dependent colon cell proliferation and tumorigenesis in vivo, and significantly, the expression of JMJD6 is markedly up-regulated in various types of human cancer especially in colon cancer, and high nuclear JMJD6 protein is strongly correlated with aggressive clinical behaviors of colon adenocarcinomas. Our results reveal a novel posttranslational modification for p53 and support the pursuit of JMJD6 as a potential biomarker for colon cancer aggressiveness and a potential target for colon cancer intervention., Author Summary JMJD6 belongs to the Jumonji C domain-containing family of proteins. The majority of this family are histone demethylases implicated in chromatin-associated events, but there have also been some reports of lysyl hydroxylase activity for JMJD6. Here we report a new posttranslational modification for the tumor suppressor protein p53 that is mediated by JMJD6. Via a physical associations with p53, JMJD6 catalyzes the hydroxylation of p53, thereby repressing its transcriptional activity. Depletion of JMJD6 promotes cell apoptosis, arrests cells in the G1 phase, sensitizes cells to DNA damaging agent-induced cell death, and represses p53-dependent colon cell proliferation and tumorigenesis. Significantly, the expression of JMJD6 is markedly up-regulated in various types of human cancer especially in colon cancer, and high nuclear JMJD6 protein is strongly correlated with aggressive clinical behaviors of colon adenocarcinomas. Our results support the pursuit of JMJD6 as a potential biomarker for colon cancer aggressiveness and a potential target for colon cancer intervention.

Polymer flooding technology has been widely used in Daqing oilfield of China in recent years. To maintain the viscosity of polymer solution, fresh water was used to confect polymer solution instead of produced water. Then, a problem was generated that the produced water could not be injected into the stratum totally. The superfluous produced water must be treated to meet the requirement of discharge, which increased the cost of production. In this paper, we try to desalt the produced water using electrodialysis principle. A set of experimental equipments that could generate desalted produced water 300 m3/d, was operated at a produced water treatment station of Daqing Oilfield. The experimental results indicated that the total salinity of desalted produced water less than 1000 mg/L. Comparing with fresh water, the polymer solution confected with desalted produced water has a sound character, its viscosity increases 63.5%, and the recovery ratio increases 4.5 percentage points. This experimental study has an important significance for the balance of oily water of production and injection in Daqing Oilfield. This produced water desalination technology would have a better application prospect in polymer flooding production.

The structure, separation principle, varieties, and application fields of hydrocyclones are introduced first. Based on Computational Fluid Dynamics (CFD) method, by using FLUENT pack, the flow field inside hydrocyclones were studied and analyzed. Velocity distribution and flow path line characteristics of the hydrocyclone were mainly studied, especially when the operating parameters changed. By using a Laser Doppler Anemometer (LDA), the simulation results were verified to some extent, which is beneficial to further understanding of hydrocyclonic separation mechanism and its research. In addition, the application aspects and main results of CFD analysis on hydrocyclonic technology study are discussed, such as Locus of Zero Vertical Velocity (LZVV), recirculation flow, and short circuit flow (SCF) inside hydrocyclones. The CFD methods can be applied on the velocity field and density field distribution characteristics research, separation mechanism analysis, separation efficiency prediction, and so forth. The CFD analysis of flow field simulation, efficiency prediction, and part erosion study on hydrocyclonic optimization design are introduced. The CFD application outlook and future work are also addressed.Copyright © 2006 by ASME

A new type hydrocyclone, air-injected de-oil hydrocyclone (AIDOH), was developed. Basic separation principle of the AIDOH, prototype structure, experimental technical process and facilities are introduced. The effect of different geometric parameters, including vortex finder diameter and length, micro-pore diameter, and different operating conditions, such as flowrate, split ratio, gas-liquid ratio, and air-injecting position, were studied. Laboratory experiments were carried out first to provide basis for field tests. The field research indicates that, for around 1000 mg/l polymer-flooding oily produced-water, the optimum flowrate is 4.20 m3/h for the testing prototype, the split ratio should be 30%; gas-liquid ratio 0.45. It also shows that 20–40 µm micro-pore materials are optimum diameter for hydrocyclonic separation enhancement Research indicates that the AIDOH has satisfied separation effect, which is feasible for emulsified oil treatment. The AIDOH will have better application prospect in petrochemical, environmental field, especially for offshore application, to reduce space occupied by normal water treatment facilities.

Produced water from polymer flooding(polymer produced water) was difficult to be treated by conventional two-stage settling process. A new type of double-cone air sparged hydrocyclone (DcASH) was designed to treat produced water from polymer flooding. The experimental results showed that DcASH had a high capacity, and it was a very promising device for treatment of produced water from polymer flooding.

In this study, an approach using influent COD/N ratio reduction was employed to improve process performance and nitrification efficiency in a membrane bioreactor (MBR). Besides sludge reduction, membrane fouling alleviation was observed during 330 d operation, which was attributed to the decreased production of soluble microbial products (SMP) and efficient carbon metabolism in the autotrophic nitrifying community. 454 high-throughput 16S rRNA gene pyrosequencing revealed that the diversity of microbial sequences was mainly determined by the feed characteristics, and that microbes could derive energy by switching to a more autotrophic metabolism to resist the environmental stress. The enrichment of nitrifiers in an MBR with a low COD/N-ratio demonstrated that this condition stimulated nitrification, and that the community distribution of ammonia oxidizing bacteria (AOB) and nitrite oxidizing bacteria (NOB) resulted in faster nitrite uptake rates. Further, ammonia oxidation was the rate-limiting step during the full nitrification.