RNF168 cooperates with RNF8 to mediate FOXM1 ubiquitination and degradation in breast cancer epirubicin treatment

The forkhead box M1 (FOXM1) transcription factor has a central role in genotoxic agent response in breast cancer. FOXM1 is regulated at the post-translational level upon DNA damage, but the key mechanism involved remained enigmatic. RNF168 is a ubiquitination E3-ligase involved in DNA damage response. Western blot and gene promoter-reporter analyses showed that the expression level and transcriptional activity of FOXM1 reduced upon RNF168 overexpression and increased with RNF168 depletion by siRNA, suggesting that RNF168 negatively regulates FOXM1 expression. Co-immunoprecipitation studies in MCF-7 cells revealed that RNF168 interacted with FOXM1 and that upon epirubicin treatment FOXM1 downregulation was associated with an increase in RNF168 binding and conjugation to the protein degradation-associated K48-linked polyubiquitin chains. Consistently, RNF168 overexpression resulted in an increase in turnover of FOXM1 in MCF-7 cells treated with the protein synthesis inhibitor cycloheximide. Conversely, RNF168, knockdown significantly enhanced the half-life of FOXM1 in both absence and presence of epirubicin. Using a SUMOylation-defective FOXM1-5x(K>R) mutant, we demonstrated that SUMOylation is required for the recruitment of RNF168 to mediate FOXM1 degradation. In addition, clonogenic assays also showed that RNF168 mediates epirubicin action through targeting FOXM1, as RNF168 could synergise with epirubicin to repress clonal formation in wild-type but not in FOXM1-deficient mouse embryo fibroblasts (MEFs). The physiological relevance of RNF168-mediated FOXM1 repression is further emphasized by the significant inverse correlation between FOXM1 and RNF168 expression in breast cancer patient samples. Moreover, we also obtained evidence that RNF8 recruits RNF168 to FOXM1 upon epirubicin treatment and cooperates with RNF168 to catalyse FOXM1 ubiquitination and degradation. Collectively, these data suggest that RNF168 cooperates with RNF8 to mediate the ubiquitination and degradation of SUMOylated FOXM1 in breast cancer genotoxic response

Simulation of Skimming Flow over Moderate Stepped Spillways Using Smoothed Particle Hydrodynamics

Source: ICHE Conference Archive - https://mdi-de.baw.de/icheArchiv

EXTENDED COST BENEFIT ANALYSIS OF A BIOGAS GENERATION PLANT

Solid waste is a growing problem in Sri Lanka in the absence of proper managementmeasures. Development and implementation of a National strategy for municipal solidwaste management is essential in order to reduce environmental, social and the economicproblems associated with the present disposal practices. Such strategies however, need tobe subjected to proper economic analysis in order to arrive at informed decisions. Thispaper presents an extended cost benefit analysis of a biogas generation plant that usesmunicipal solid waste as the raw material.To dispose vegetable market garbage available in Wattala, Kandana and Ja-ela areas a640 Mt capacity biogas/ bio fertilizer project has been housed at Muthurajawela (alongHamilton canal, Elakanda), by the National Engineering Research & DevelopmentCenter of Sri Lanka (NERDC) which uses Dry Batch Anaerobic Digester Technology.Among the other biological treatment options, anaerobic digestion is the most costeffective, due to the high-energy recovery linked to the process and its limitedenvironmental impacts.Economic analysis has been carried out to identify environmental costs and benefitsassociated with the above project. Several environmental valuation methods have beenapplied to value the identified costs and benefits. The main benefit of reduction ofmunicipal solid waste has been estimated by contingent valuation method using samplesfrom Wattala-Mabola, Ja-ela, and Peliyagoda local authority areas. Another benefit is thereduction of green house has emission due to the reduction of open dumping. Greenhouse gas (methane) emission from solid waste was estimated through Sholl Canionmodel and valued using avoided global damage cost approach. Benefits of biogas asenergy and organic fertilizers and employment benefits were estimated by market basedapproachesThe project is viable from economy and environment point of view with net present valueof Rs 142 million for a 20-year period with 10% discount rate. This analysis provides ajustification for undertaking solid waste management strategies in a technologically,environmentally and economically viable manner

SUMOylation inhibits FOXM1 activity and delays mitotic transition

The forkhead box transcription factor FOXM1 is an essential effector of G2/M-phase transition, mitosis and the DNA damage response. As such, it is frequently deregulated during tumorigenesis. Here we report that FOXM1 is dynamically modified by SUMO1 but not by SUMO2/3 at multiple sites. We show that FOXM1 SUMOylation is enhanced in MCF-7 breast cancer cells in response to treatment with epirubicin and mitotic inhibitors. Mutation of five consensus conjugation motifs yielded a SUMOylation-deficient mutant FOXM1. Conversely, fusion of the E2 ligase Ubc9 to FOXM1 generated an auto-SUMOylating mutant (FOXM1-Ubc9). Analysis of wild-type FOXM1 and mutants revealed that SUMOylation inhibits FOXM1 activity, promotes translocation to the cytoplasm and enhances APC/Cdh1-mediated ubiquitination and degradation. Further, expression of the SUMOylation-deficient mutant enhanced cell proliferation compared with wild-type FOXM1, whereas the FOXM1-Ubc9 fusion protein resulted in persistent cyclin B1 expression and slowed the time from mitotic entry to exit. In summary, our findings suggest that SUMOylation attenuates FOXM1 activity and causes mitotic delay in cytotoxic drug response

RNF168 cooperates with RNF8 to mediate FOXM1 ubiquitination and degradation in breast cancer epirubicin treatment

The forkhead box M1 (FOXM1) transcription factor has a central role in genotoxic agent response in breast cancer. FOXM1 is regulated at the post-translational level upon DNA damage, but the key mechanism involved remained enigmatic. RNF168 is a ubiquitination E3-ligase involved in DNA damage response. Western blot and gene promoter-reporter analyses showed that the expression level and transcriptional activity of FOXM1 reduced upon RNF168 overexpression and increased with RNF168 depletion by siRNA, suggesting that RNF168 negatively regulates FOXM1 expression. Co-immunoprecipitation studies in MCF-7 cells revealed that RNF168 interacted with FOXM1 and that upon epirubicin treatment FOXM1 downregulation was associated with an increase in RNF168 binding and conjugation to the protein degradation-associated K48-linked polyubiquitin chains. Consistently, RNF168 overexpression resulted in an increase in turnover of FOXM1 in MCF-7 cells treated with the protein synthesis inhibitor cycloheximide. Conversely, RNF168, knockdown significantly enhanced the half-life of FOXM1 in both absence and presence of epirubicin. Using a SUMOylation-defective FOXM1-5x(K>R) mutant, we demonstrated that SUMOylation is required for the recruitment of RNF168 to mediate FOXM1 degradation. In addition, clonogenic assays also showed that RNF168 mediates epirubicin action through targeting FOXM1, as RNF168 could synergise with epirubicin to repress clonal formation in wild-type but not in FOXM1-deficient mouse embryo fibroblasts (MEFs). The physiological relevance of RNF168-mediated FOXM1 repression is further emphasized by the significant inverse correlation between FOXM1 and RNF168 expression in breast cancer patient samples. Moreover, we also obtained evidence that RNF8 recruits RNF168 to FOXM1 upon epirubicin treatment and cooperates with RNF168 to catalyse FOXM1 ubiquitination and degradation. Collectively, these data suggest that RNF168 cooperates with RNF8 to mediate the ubiquitination and degradation of SUMOylated FOXM1 in breast cancer genotoxic response.published_or_final_versio

OTUB1 inhibits the ubiquitination and degradation of FOXM1 in breast cancer and epirubicin resistance

The forkhead transcription factor FOXM1 has a key role in DNA damage response, and its deregulated overexpression is associated with genotoxic drug resistance in breast cancer. However, little is known about the posttranslational mechanisms by which FOXM1 expression is regulated by genotoxic agents and how they are deregulated in resistant cells. Initial co-immunoprecipitation studies verified previous proteomic analysis finding that the OTUB1 is a novel FOXM1-interacting protein. Western blot analysis showed that both OTUB1 and FOXM1 expression reduced upon genotoxic agent treatment in MCF-7 cells, but remained relatively constant in resistant cells. FOXM1 expression reduced upon OTUB1 depletion by siRNA and increased with OTUB1 overexpression in MCF-7 cells, arguing that OTUB1 positively regulates FOXM1 expression. In agreement, co-immunoprecipitation experiments demonstrated that FOXM1 expression is associated with OTUB1 binding but inversely correlates with conjugation to the protein degradation-associated Lys-48-linked ubiquitin-chains. Overexpression of wild-type (WT) OTUB1, but not the OTUB1(C91S) mutant, disrupted the formation of Lys48-linked ubiquitin-conjugates on FOXM1. Importantly, knockdown of OTUB1 by siRNA resulted in an increase in turnover of FOXM1 in MCF-7 cells treated with the protein synthesis inhibitor cycloheximide, whereas overexpression of WT OTUB1, but not the OTUB1(C91S) mutant, significantly enhances the half-life of FOXM1. In addition, proliferative and clonogenic assays also show that OTUB1 can enhance the proliferative rate and epirubicin resistance through targeting FOXM1, as OTUB1 has little effect on FOXM1-deficient cells. The physiological relevance of the regulation of FOXM1 by OTUB1 is further underscored by the significant correlations between FOXM1 and OTUB1 expression in breast cancer patient samples. Cox-regression survival analysis indicates that OTUB1 overexpression is linked to poorer outcome in particular in patients treated with chemotherapy. Collectively, these data suggest that OTUB1 limits the ubiquitination and degradation of FOXM1 in breast cancer and has a key role in genotoxic agent resistance

Preliminary study on herpetofaunal diversity of Nilgala forest area in Monaragala district, Sri Lanka

Nilgala ForestArea (NFA) is one of the largest and important forest area in Monaragala District, UvaProvince. It contain 12,432 hectares and lies within 7° 08' - 7° 14' NL and 81 D 16' - 81020' EL. Itselevation range between 200m to 700m within the Irindahela, Hangale, Yakun hela (highest point 700rn), Hamapola, Badangarnuwa, Keenagoda, Makada, Karadugala, Kukulagoda, Ewalahela, Gorikkadahills. The mean annual rainfall varies between where the average annual rainfall 1500mm - 2000mm(rain during northeast monsoon). while the mean annual temperature of the area is 28 DC - 31°C.The vegetation comprised with lowland tropical moist semi evergreen forest and savannah forest,home gardens and small patch paddy cultivations. The dominant tree species are Aralu (Terminaliachebulav, Bulu iTerminalia bellirica) and Nelli iPhyllanthus emblicay. Other than biodiversity,Nilgala is rich of archaeological monuments, such as prehistoric, proto-historic and historical Buddhistmonasteries.During the two-year study period, total number of 70 reptile species were and 19 amphibian speciesrecorded. Reptiles include 44 genera of 17 families and 20 (28.5%) endemic species. Amphibianfauna contain 13 genera including 4 families and 6 (31.5%) endemic species. 41.4% (29) of reptilesand 26.3% (5) of Amphibians listed as 'Nationally Threatened' in the 1999 IUCN National threatenedlist. Out of70 species 38 (54.2%) are Serpentoid reptiles (11 endemics) and 32 (45.7%) species areof Tetrapod reptiles (9 endemics). Among the recorded species, 11 Serpentoid, 3 Tetrapod, and 2amphibians have not been recorded by previous workers. Furthermore seven unidentified specieswere also recorded during the survey, which probably include new amphibian species belonging togenus Nannophrys. Human activities such as man-made fire, ilIegallogging, extensive use of chemicalsfor agriculture, forest clearing for chena cultivation and road kills were identified as a main threat forthe natural habitats as well as faunal species.

Finding needles in haystacks: linking scientific names, reference specimens and molecular data for Fungi

DNA phylogenetic comparisons have shown that morphology-based species recognition often underestimates fungal diversity. Therefore, the need for accurate DNA sequence data, tied to both correct taxonomic names and clearly annotated specimen data, has never been greater. Furthermore, the growing number of molecular ecology and microbiome projects using high-throughput sequencing require fast and effective methods for en masse species assignments. In this article, we focus on selecting and re-annotating a set of marker reference sequences that represent each currently accepted order of Fungi. The particular focus is on sequences from the internal transcribed spacer region in the nuclear ribosomal cistron, derived from type specimens and/or ex-type cultures. Re-annotated and verified sequences were deposited in a curated public database at the National Center for Biotechnology Information (NCBI), namely the RefSeq Targeted Loci (RTL) database, and will be visible during routine sequence similarity searches with NR_prefixed accession numbers. A set of standards and protocols is proposed to improve the data quality of new sequences, and we suggest how type and other reference sequences can be used to improve identification of Fungi

Finding needles in haystacks: Linking scientific names, reference specimens and molecular data for Fungi

DNA phylogenetic comparisons have shown that morphology-based species recognition often underestimates fungal diversity. Therefore, the need for accurate DNA sequence data, tied to both correct taxonomic names and clearly annotated specimen data, has never been greater. Furthermore, the growing number of molecular ecology and microbiome projects using high-throughput sequencing require fast and effective methods for en masse species assignments. In this article, we focus on selecting and re-annotating a set of marker reference sequences that represent each currently accepted order of Fungi. The particular focus is on sequences from the internal transcribed spacer region in the nuclear ribosomal cistron, derived from type specimens and/or ex-type cultures. Reannotated and verified sequences were deposited in a curated public database at the National Center for Biotechnology Information (NCBI), namely the RefSeq Targeted Loci (RTL) database, and will be visible during routine sequence similarity searches with NR_prefixed accession numbers. A set of standards and protocols is proposed to improve the data quality of new sequences, and we suggest how type and other reference sequences can be used to improve identification of Fungi.B.R. and C.L.S. acknowledge support from the Intramural Research Program of the National Institutes of Health, National Library of MedicinePeer Reviewe