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2.
Environ Sci Technol ; 53(7): 3620-3633, 2019 04 02.
Artigo em Inglês | MEDLINE | ID: mdl-30830765

RESUMO

Little is known about the regional extent and variability of nitrate from atmospheric deposition that is transported to streams without biological processing in forests. We measured water chemistry and isotopic tracers (δ18O and δ15N) of nitrate sources across the Northern Forest Region of the U.S. and Canada and reanalyzed data from other studies to determine when, where, and how unprocessed atmospheric nitrate was transported in catchments. These inputs were more widespread and numerous than commonly recognized, but with high spatial and temporal variability. Only 6 of 32 streams had high fractions (>20%) of unprocessed atmospheric nitrate during baseflow. Seventeen had high fractions during stormflow or snowmelt, which corresponded to large fractions in near-surface soil waters or groundwaters, but not deep groundwater. The remaining 10 streams occasionally had some (<20%) unprocessed atmospheric nitrate during stormflow or baseflow. Large, sporadic events may continue to be cryptic due to atmospheric deposition variation among storms and a near complete lack of monitoring for these events. A general lack of observance may bias perceptions of occurrence; sustained monitoring of chronic nitrogen pollution effects on forests with nitrate source apportionments may offer insights needed to advance the science as well as assess regulatory and management schemes.


Assuntos
Florestas , Nitratos , Canadá , Monitoramento Ambiental , Nitrogênio , Rios
3.
PLoS One ; 14(3): e0212670, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30913212

RESUMO

Immunotherapy has fundamentally changed the landscape of cancer treatment. Despite the encouraging results with the checkpoint modulators, response rates vary widely across tumor types, with a majority of patients exhibiting either primary resistance without a significant initial response to treatment or acquired resistance with subsequent disease progression. Hematopoietic progenitor kinase 1 (HPK1) is predominantly expressed in hematopoietic cell linages and serves as a negative regulator in T cells and dendritic cells (DC). While HPK1 gene knockout (KO) studies suggest its role in anti-tumor immune responses, the involvement of kinase activity and thereof its therapeutic potential remain unknown. To investigate the potential of pharmacological intervention using inhibitors of HPK1, we generated HPK1 kinase dead (KD) mice which carry a single loss-of-function point mutation in the kinase domain and interrogated the role of kinase activity in immune cells in the context of suppressive factors or the tumor microenvironment (TME). Our data provide novel findings that HKP1 kinase activity is critical in conferring suppressive functions of HPK1 in a wide range of immune cells including CD4+, CD8+, DC, NK to Tregs, and inactivation of kinase domain was sufficient to elicit robust anti-tumor immune responses. These data support the concept that an HPK1 small molecule kinase inhibitor could serve as a novel agent to provide additional benefit in combination with existing immunotherapies, particularly to overcome resistance to current treatment regimens.

4.
FEMS Microbiol Ecol ; 95(2)2019 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-30629168

RESUMO

The temporal dynamics of soil bacterial communities are understudied, but such understanding is critical to elucidating the drivers of community variation. The goal of this study was to characterize how soil bacterial communities vary across diurnal, sub-seasonal and seasonal time-scales in a 5.8 m2 plot and test the hypothesis that bacterial diversity varies on each of these scales. We used 16S rDNA gene amplicon sequencing to quantify the alpha and beta diversity of soil bacteria as well as the Net Relatedness Index and Nearest Taxon Indices to assess the degree of phylogenetic clustering, and the extent to which community shifts were driven by stochastic vs. deterministic limitation. We found that species richness was highest in winter, lowest in fall and that communities were compositionally distinct across seasons. There was no evidence of diurnal-scale shifts; the finest temporal scale over which community shifts were detected using our DNA-based analysis was between sampling dates separated by 6 weeks. Phylogenetic analyses suggested that seasonal-scale differences in community composition were the result of environmental filtering and homogeneous selection. Our findings provide insight into temporal variation of soil bacterial communities across the hourly to seasonal scales while minimizing the potential confounding effect of spatial variation.

5.
Eur J Protistol ; 67: 15-26, 2018 Oct 26.
Artigo em Inglês | MEDLINE | ID: mdl-30445234

RESUMO

Paleoecological records suggest that growing season length and/or cloudiness may affect peatland carbon accumulation and testate amoeba-based environmental reconstructions, highlighting a need to understand how light intensity affects microbial communities. We shaded plots on two peatlands for two years to examine effects on testate amoeba communities, the relative abundance of mixotrophic and heterotrophic testate amoebae, transfer-function performance, and δ13C values of two species of mixotrophic testate amoebae. Surprisingly, relative abundance of mixotrophic species increased in shade, although compositional changes did not affect transfer-function performance. Shading did not affect δ13C values of Hyalosphenia papilio and Heleopera sphagni, which ranged from -23.5 to -19.6‰ and -23.2 to -19.2‰, respectively. These δ13C values were higher than those of potential food sources and lower than literature-derived values for Chlorella, the zoochlorellae inhabiting mixotrophic testate amoebae. δ13C values thus suggest that these mixotrophic species obtain some carbon from Chlorella, although coupled dietary and isotope studies are needed to quantify this contribution. More research is needed to assess impacts of light variability on peatland microbial communities; however, carbon sources are recorded by δ13C values of testate amoebae, indicating potential for studies of carbon cycling and how mixotrophy varies temporally and spatially.

6.
Nat Ecol Evol ; 2(11): 1735-1744, 2018 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-30349095

RESUMO

Human societies depend on an Earth system that operates within a constrained range of nutrient availability, yet the recent trajectory of terrestrial nitrogen (N) availability is uncertain. Examining patterns of foliar N concentrations and isotope ratios (δ15N) from more than 43,000 samples acquired over 37 years, here we show that foliar N concentration declined by 9% and foliar δ15N declined by 0.6-1.6‰. Examining patterns across different climate spaces, foliar δ15N declined across the entire range of mean annual temperature and mean annual precipitation tested. These results suggest declines in N supply relative to plant demand at the global scale. In all, there are now multiple lines of evidence of declining N availability in many unfertilized terrestrial ecosystems, including declines in δ15N of tree rings and leaves from herbarium samples over the past 75-150 years. These patterns are consistent with the proposed consequences of elevated atmospheric carbon dioxide and longer growing seasons. These declines will limit future terrestrial carbon uptake and increase nutritional stress for herbivores.

7.
Rapid Commun Mass Spectrom ; 32(21): 1859-1866, 2018 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-30076636

RESUMO

RATIONALE: Stable hydrogen isotope (δ2 H) ratios of animal tissues are useful for assessing movement and geographic origin of mobile organisms. However, it is uncertain whether heat and singeing affects feather δ2 H values and thus subsequent geographic assignments. This is relevant for birds of conservation interest that are burned and killed at concentrating solar-energy facilities that reflect sunlight to a receiving tower and generate a solar flux field. METHODS: We used a controlled experiment to test the effect of known heat loads (exposure to 200, 250 or 300°C for 1 min) on the morphology and δ2 H values of feathers from two songbird species. Subsequently, we examined the effects of singeing on δ2 H values of feathers from three other songbird species that were found dead in the field at a concentrating solar-energy facility. RESULTS: Relative to control samples, heating caused visual morphological changes to feathers, including shriveling at 250°C and charring at 300°C. The δ2 H values significantly declined by a mean of 27.8‰ in experimental samples exposed to 300°C. There was no statistically detectable difference between δ2 H values of the singed and unsinged portions of field-collected feathers from the same bird. CONCLUSIONS: Limited singeing that did not dramatically alter the feather morphology did not substantially affect δ2 H values of feathers from these songbirds. However, higher temperatures induced charring and reduced δ2 H values. Therefore, severely charred feathers should be avoided when selecting feathers for δ2 H-based assessment of geographic origin.


Assuntos
Deutério/análise , Plumas/química , Plumas/efeitos da radiação , Aves Canoras/fisiologia , Migração Animal/fisiologia , Animais , Temperatura Alta , Espectrometria de Massas/métodos
8.
Drug Metab Dispos ; 46(2): 178-188, 2018 02.
Artigo em Inglês | MEDLINE | ID: mdl-29162614

RESUMO

Perturbation of organic anion transporter (OAT) 1- and OAT3-mediated transport can alter the exposure, efficacy, and safety of drugs. Although there have been reports of the endogenous biomarkers for OAT1/3, none of these have all of the characteristics required for a clinical useful biomarker. Cynomolgus monkeys were treated with intravenous probenecid (PROB) at a dose of 40 mg/kg in this study. As expected, PROB increased the area under the plasma concentration-time curve (AUC) of coadministered furosemide, a known substrate of OAT1 and OAT3, by 4.1-fold, consistent with the values reported in humans (3.1- to 3.7-fold). Of the 233 plasma metabolites analyzed using a liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based metabolomics method, 29 metabolites, including pyridoxic acid (PDA) and homovanillic acid (HVA), were significantly increased after either 1 or 3 hours in plasma from the monkeys pretreated with PROB compared with the treated animals. The plasma of animals was then subjected to targeted LC-MS/MS analysis, which confirmed that the PDA and HVA AUCs increased by approximately 2- to 3-fold by PROB pretreatments. PROB also increased the plasma concentrations of hexadecanedioic acid (HDA) and tetradecanedioic acid (TDA), although the increases were not statistically significant. Moreover, transporter profiling assessed using stable cell lines constitutively expressing transporters demonstrated that PDA and HVA are substrates for human OAT1, OAT3, OAT2 (HVA), and OAT4 (PDA), but not OCT2, MATE1, MATE2K, OATP1B1, OATP1B3, and sodium taurocholate cotransporting polypeptide. Collectively, these findings suggest that PDA and HVA might serve as blood-based endogenous probes of cynomolgus monkey OAT1 and OAT3, and investigation of PDA and HVA as circulating endogenous biomarkers of human OAT1 and OAT3 function is warranted.


Assuntos
Biomarcadores/sangue , Ácido Homovanílico/sangue , Proteína 1 Transportadora de Ânions Orgânicos/metabolismo , Transportadores de Ânions Orgânicos Sódio-Independentes/metabolismo , Ácido Piridóxico/sangue , Animais , Transporte Biológico/fisiologia , Linhagem Celular , Células HEK293 , Humanos , Macaca fascicularis , Metabolômica/métodos , Probenecid/metabolismo
9.
Sci Rep ; 7(1): 15932, 2017 Nov 21.
Artigo em Inglês | MEDLINE | ID: mdl-29162842

RESUMO

Bats perform important ecosystem services, but it remains difficult to quantify their dietary strategies and trophic position (TP) in situ. We conducted measurements of nitrogen isotopes of individual amino acids (δ 15NAA) and bulk-tissue carbon (δ 13Cbulk) and nitrogen (δ 15Nbulk) isotopes for nine bat species from different feeding guilds (nectarivory, frugivory, sanguivory, piscivory, carnivory, and insectivory). Our objective was to assess the precision of δ 15NAA-based estimates of TP relative to other approaches. TPs calculated from δ 15N values of glutamic acid and phenylalanine, which range from 8.3-33.1‰ and 0.7-15.4‰ respectively, varied between 1.8 and 3.8 for individuals of each species and were generally within the ranges of those anticipated based on qualitative dietary information. The δ 15NAA approach reveals variation in TP within and among species that is not apparent from δ 15Nbulk data, and δ 15NAA data suggest that two insectivorous species (Lasiurus noctivagans and Lasiurus cinereus) are more omnivorous than previously thought. These results indicate that bats exhibit a trophic discrimination factor (TDF) similar to other terrestrial organisms and that δ 15NAA provides a reliable approach for addressing questions about variation in the TP of bats that have heretofore proven elusive.

10.
Rapid Commun Mass Spectrom ; 31(21): 1825-1834, 2017 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-28833668

RESUMO

RATIONALE: Carbon isotope (δ13 C ) data from arthropod cuticles provide invaluable information on past and present biogeochemical processes. However, such analyses typically require large sample sizes that may mask important variation in δ13 C values within or among species. METHODS: We have evaluated a spooling-wire microcombustion (SWiM) device and isotope ratio mass spectrometry (IRMS) to measure the δ13 C values of carbon dissolved from the cuticle of chitinous aquatic zooplankton. The effects of temperature, pH, and reaction time on the δ13 C values of acid-dissolved bulk cuticle and purified chitin fractions obtained from a single species of chironomid from four commercial suppliers were assessed. These results were compared with baseline δ13 C values obtained on solid cuticle using conventional EA (elemental analyzer)/IRMS. RESULTS: The results indicate differential, time-dependent dissolution of chitin, lipid and protein fractions of cuticle concomitant with slow depolymerization and deacetylation of chitin. Isotopic offsets between dissolved bulk head capsules and a purified chitin fraction suggest the contributions of other isotopically lighter components of the bulk head capsules to bulk chitin extracts. The SWiM/IRMS δ13 C results obtained on dissolved cuticle using a treatment of 4 N HCl at 25 °C for 24 h produced generally stable δ13 C values, large sample/blank CO2 yields and a positive correlation with conventional EA/IRMS results on unprocessed cuticle. CONCLUSIONS: The SWiM/IRMS system offers a reliable method to determine δ13 C values on nanogram quantities of carbon from dissolved insect cuticle, thus reducing sample size requirements and providing new opportunities to use δ13 C variation among/within species for reconstructing paleo-biogeochemical processes.


Assuntos
Isótopos de Carbono/análise , Insetos/química , Paleontologia/métodos , Animais , Quitina/química , Fósseis , Proteínas de Insetos/química , Espectrometria de Massas/métodos , Reprodutibilidade dos Testes
11.
Sci Rep ; 7(1): 7759, 2017 Aug 10.
Artigo em Inglês | MEDLINE | ID: mdl-28798483

RESUMO

Variation across climate gradients in the isotopic composition of nitrogen (N) and carbon (C) in foliar tissues has the potential to reveal ecological processes related to N and water availability. However, it has been a challenge to separate spatial patterns related to direct effects of climate from effects that manifest indirectly through species turnover across climate gradients. Here we compare variation along environmental gradients in foliar N isotope (δ15N) and C isotopic discrimination (Δ13C) measured in 755 specimens of a single widely distributed tree species, Populus balsamifera, with variation represented in global databases of foliar isotopes. After accounting for mycorrhizal association, sample size, and climatic range, foliar δ15N in P. balsamifera was more weakly related to mean annual precipitation and foliar N concentration than when measured across species, yet exhibited a stronger negative effect of mean annual temperature. Similarly, the effect of precipitation and elevation on Δ13C were stronger in a global data base of foliar Δ13C samples than observed in P. balsamifera. These results suggest that processes influencing foliar δ15N and Δ13C in P. balsamifera are partially normalized across its climatic range by the habitat it occupies or by the physiology of the species itself.

12.
J Pharmacol Exp Ther ; 362(3): 385-394, 2017 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-28645914

RESUMO

The interference of bile acid secretion through bile salt export pump (BSEP) inhibition is one of the mechanisms for troglitazone (TGZ)-induced hepatotoxicity. Here, we investigated the impact of single or repeated oral doses of TGZ (200 mg/kg/day, 7 days) on bile acid homoeostasis in wild-type (WT) and Bsep knockout (KO) rats. Following oral doses, plasma exposures of TGZ were not different between WT and KO rats, and were similar on day 1 and day 7. However, plasma exposures of the major metabolite, troglitazone sulfate (TS), in KO rats were 7.6- and 9.3-fold lower than in WT on day 1 and day 7, respectively, due to increased TS biliary excretion. With Bsep KO, the mRNA levels of multidrug resistance-associated protein 2 (Mrp2), Mrp3, Mrp4, Mdr1, breast cancer resistance protein (Bcrp), sodium taurocholate cotransporting polypeptide, small heterodimer partner, and Sult2A1 were significantly altered in KO rats. Following seven daily TGZ treatments, Cyp7A1 was significantly increased in both WT and KO rats. In the vehicle groups, plasma exposures of individual bile acids demonstrated variable changes in KO rats as compared with WT. WT rats dosed with TGZ showed an increase of many bile acid species in plasma on day 1, suggesting the inhibition of Bsep. Conversely, these changes returned to base levels on day 7. In KO rats, alterations of most bile acids were observed after seven doses of TGZ. Collectively, bile acid homeostasis in rats was regulated through bile acid synthesis and transport in response to Bsep deficiency and TGZ inhibition. Additionally, our study is the first to demonstrate that repeated TGZ doses can upregulate Cyp7A1 in rats.


Assuntos
Transportadores de Cassetes de Ligação de ATP/genética , Ácidos e Sais Biliares/metabolismo , Cromanos/farmacologia , Homeostase/efeitos dos fármacos , Homeostase/genética , Hipoglicemiantes/farmacologia , Tiazolidinedionas/farmacologia , Membro 11 da Subfamília B de Transportadores de Cassetes de Ligação de ATP , Animais , Bile/metabolismo , Colesterol 7-alfa-Hidroxilase/biossíntese , Colesterol 7-alfa-Hidroxilase/genética , Técnicas de Inativação de Genes , Fígado/efeitos dos fármacos , Fígado/metabolismo , Masculino , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , Ratos , Ratos Sprague-Dawley , Troglitazona , Regulação para Cima/efeitos dos fármacos
13.
J Invest Dermatol ; 137(10): 2197-2207, 2017 10.
Artigo em Inglês | MEDLINE | ID: mdl-28647344

RESUMO

On acquisition of an oncogenic mutation, primary human and mouse cells can enter oncogene-induced senescence (OIS). OIS is characterized by a stable proliferation arrest and secretion of proinflammatory cytokines and chemokines, the senescence-associated secretory phenotype. Proliferation arrest and the senescence-associated secretory phenotype collaborate to enact tumor suppression, the former by blocking cell proliferation and the latter by recruiting immune cells to clear damaged cells. However, the interactions of OIS cells with the immune system are still poorly defined. Here, we show that engagement of OIS in primary human melanocytes, specifically by melanoma driver mutations NRASQ61K and BRAFV600E, causes expression of the major histocompatibility class II antigen presentation apparatus, via secreted IL-1ß signaling and expression of CIITA, a master regulator of major histocompatibility class II gene transcription. In vitro, OIS melanocytes activate T-cell proliferation. In vivo, nonproliferating oncogene-expressing melanocytes localize to skin-draining lymph nodes, where they induce T-cell proliferation and an antigen presentation gene expression signature. In patients, expression of major histocompatibility class II in melanoma is linked to favorable disease outcome. We propose that OIS in melanocytes is accompanied by an antigen presentation phenotype, likely to promote tumor suppression via activation of the adaptive immune system.


Assuntos
Genes MHC da Classe II/genética , Melanócitos/metabolismo , Melanoma/genética , Oncogenes/genética , Regulação para Cima , Animais , Linhagem Celular Tumoral , Proliferação de Células , Senescência Celular , Humanos , Melanócitos/patologia , Melanoma/metabolismo , Melanoma/patologia , Camundongos , Transdução de Sinais
14.
Conserv Biol ; 31(2): 406-415, 2017 04.
Artigo em Inglês | MEDLINE | ID: mdl-27677518

RESUMO

Renewable energy production is expanding rapidly despite mostly unknown environmental effects on wildlife and habitats. We used genetic and stable isotope data collected from Golden Eagles (Aquila chrysaetos) killed at the Altamont Pass Wind Resource Area (APWRA) in California in demographic models to test hypotheses about the geographic extent and demographic consequences of fatalities caused by renewable energy facilities. Geospatial analyses of δ2 H values obtained from feathers showed that ≥25% of these APWRA-killed eagles were recent immigrants to the population, most from long distances away (>100 km). Data from nuclear genes indicated this subset of immigrant eagles was genetically similar to birds identified as locals from the δ2 H data. Demographic models implied that in the face of this mortality, the apparent stability of the local Golden Eagle population was maintained by continental-scale immigration. These analyses demonstrate that ecosystem management decisions concerning the effects of local-scale renewable energy can have continental-scale consequences.


Assuntos
Conservação dos Recursos Naturais , Águias , Vento , Animais , California , Plumas , Dinâmica Populacional , Energia Renovável
15.
Ecol Appl ; 26(5): 1381-1395, 2016 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-27755755

RESUMO

An unanticipated impact of wind-energy development has been large-scale mortality of insectivorous bats. In eastern North America, where mortality rates are among the highest in the world, the hoary bat (Lasiurus cinereus) and the eastern red bat (L. borealis) comprise the majority of turbine-associated bat mortality. Both species are migratory tree bats with widespread distributions; however, little is known regarding the geographic origins of bats killed at wind-energy facilities or the diversity and population structure of affected species. We addressed these unknowns by measuring stable hydrogen isotope ratios (δ2 H) and conducting population genetic analyses of bats killed at wind-energy facilities in the central Appalachian Mountains (USA) to determine the summering origins, effective size, structure, and temporal stability of populations. Our results indicate that ~1% of hoary bat mortalities and ~57% of red bat mortalities derive from non-local sources, with no relationship between the proportion of non-local bats and sex, location of mortality, or month of mortality. Additionally, our data indicate that hoary bats in our sample consist of an unstructured population with a small effective size (Ne ) and either a stable or declining history. Red bats also showed no evidence of population genetic structure, but in contrast to hoary bats, the diversity contained in our red bat samples is consistent with a much larger Ne that reflects a demographic expansion after a bottleneck. These results suggest that the impacts of mortality associated with intensive wind-energy development may affect bat species dissimilarly, with red bats potentially better able to absorb sustained mortality than hoary bats because of their larger Ne . Our results provide important baseline data and also illustrate the utility of stable isotopes and population genetics for monitoring bat populations affected by wind-energy development.


Assuntos
Distribuição Animal/fisiologia , Quirópteros/genética , Quirópteros/fisiologia , Monitoramento Ambiental , Energia Renovável , Vento , Migração Animal , Animais , Dinâmica Populacional , Fatores de Tempo
16.
Nat Plants ; 2: 16133, 2016 09 12.
Artigo em Inglês | MEDLINE | ID: mdl-27618399

RESUMO

There is wide agreement that anthropogenic climate warming has influenced the phenology of forests during the late twentieth and early twenty-first centuries(1,2). Longer growing seasons can lead to increased photosynthesis and productivity(3), which would represent a negative feedback to rising CO2 and consequently warming(4,5). Alternatively, increased demand for soil resources because of a longer photosynthetically active period in conjunction with other global change factors might exacerbate resource limitation(6,7), restricting forest productivity response to a longer growing season(8,9). In this case, increased springtime productivity has the potential to increase plant nitrogen limitation by increasing plant demand for nitrogen more than nitrogen supplies, or increasing early-season ecosystem nitrogen losses(10,11). Here we show that for 222 trees representing three species in eastern North America earlier spring phenology during the past 30 years has caused declines in nitrogen availability to trees by increasing demand for nitrogen relative to supply. The observed decline in nitrogen availability is not associated with reduced wood production, suggesting that other environmental changes such as increased atmospheric CO2 and water availability are likely to have overwhelmed reduced nitrogen availability. Given current trajectories of environmental changes, nitrogen limitation is likely to continue to increase for these forests, possibly further limiting carbon sequestration potential.


Assuntos
Mudança Climática , Florestas , Nitrogênio/metabolismo , Árvores/metabolismo , Sequestro de Carbono , Maryland , Estações do Ano , Sudeste dos Estados Unidos , Estados Unidos
17.
Genome Biol ; 17(1): 158, 2016 07 25.
Artigo em Inglês | MEDLINE | ID: mdl-27457071

RESUMO

BACKGROUND: Histone modification H4K20me3 and its methyltransferase SUV420H2 have been implicated in suppression of tumorigenesis. The underlying mechanism is unclear, although H4K20me3 abundance increases during cellular senescence, a stable proliferation arrest and tumor suppressor process, triggered by diverse molecular cues, including activated oncogenes. Here, we investigate the function of H4K20me3 in senescence and tumor suppression. RESULTS: Using immunofluorescence and ChIP-seq we determine the distribution of H4K20me3 in proliferating and senescent human cells. Altered H4K20me3 in senescence is coupled to H4K16ac and DNA methylation changes in senescence. In senescent cells, H4K20me3 is especially enriched at DNA sequences contained within specialized domains of senescence-associated heterochromatin foci (SAHF), as well as specific families of non-genic and genic repeats. Altered H4K20me3 does not correlate strongly with changes in gene expression between proliferating and senescent cells; however, in senescent cells, but not proliferating cells, H4K20me3 enrichment at gene bodies correlates inversely with gene expression, reflecting de novo accumulation of H4K20me3 at repressed genes in senescent cells, including at genes also repressed in proliferating cells. Although elevated SUV420H2 upregulates H4K20me3, this does not accelerate senescence of primary human cells. However, elevated SUV420H2/H4K20me3 reinforces oncogene-induced senescence-associated proliferation arrest and slows tumorigenesis in vivo. CONCLUSIONS: These results corroborate a role for chromatin in underpinning the senescence phenotype but do not support a major role for H4K20me3 in initiation of senescence. Rather, we speculate that H4K20me3 plays a role in heterochromatinization and stabilization of the epigenome and genome of pre-malignant, oncogene-expressing senescent cells, thereby suppressing epigenetic and genetic instability and contributing to long-term senescence-mediated tumor suppression.


Assuntos
Carcinogênese/genética , Senescência Celular/genética , Cromatina/genética , Histona-Lisina N-Metiltransferase/genética , Linhagem Celular Tumoral , Proliferação de Células/genética , Metilação de DNA/genética , Epigênese Genética/genética , Regulação Neoplásica da Expressão Gênica/genética , Heterocromatina/genética , Histonas/genética , Humanos , Nevo/metabolismo , Nevo/patologia
18.
Anal Biochem ; 503: 71-8, 2016 06 15.
Artigo em Inglês | MEDLINE | ID: mdl-27033006

RESUMO

The growing field of biomarker bioanalysis by liquid chromatography mass spectrometry (LC-MS) is challenged with the selection of suitable matrices to construct relevant and valid calibration curves resulting in not only precise but also accurate data. Because surrogate matrices are often employed with the associated concerns about the accuracy of the obtained data, here we present an assay using surrogate analytes in naive biological matrices. This approach is illustrated with the analysis of endogenous bile acids (e-BAs) in serum and plasma using stable isotope-labeled (SIL) analogues as calibration standards to address the matrix concerns. Several deuterated BAs (d-BAs) were used as standards representing respectively grouped e-BAs with structural similarity allowing for the simultaneous bioanalysis of 16 e-BA. The utility of this LC-MS assay employing d-BAs is demonstrated with the analysis of samples resultant of a controlled metabolomics study where a cohort of rats was fed/fasted to investigate the change of e-BAs dependent on food consumption and fasting time.


Assuntos
Ácidos e Sais Biliares/sangue , Ácidos e Sais Biliares/metabolismo , Marcação por Isótopo , Metabolômica , Animais , Ácidos e Sais Biliares/química , Cromatografia Líquida , Humanos , Espectrometria de Massas , Estrutura Molecular , Ratos
19.
PLoS One ; 11(1): e0148023, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-26821228

RESUMO

Solubilization of cross-linked cells followed by chromatin shearing is essential for successful chromatin immunoprecipitation (ChIP). However, this task, typically accomplished by ultrasound treatment, may often become a pitfall of the process, due to inconsistent results obtained between different experiments under seemingly identical conditions. To address this issue we systematically studied ultrasound-mediated cell lysis and chromatin shearing, identified critical parameters of the process and formulated a generic strategy for rational optimization of ultrasound treatment. We also demonstrated that whereas ultrasound treatment required to shear chromatin to within a range of 100-400 bp typically degrades large proteins, a combination of brief sonication and benzonase digestion allows for the generation of similarly sized chromatin fragments while preserving the integrity of associated proteins. This approach should drastically improve ChIP efficiency for this class of proteins.


Assuntos
Imunoprecipitação da Cromatina/métodos , Cromatina/química , Sonicação/métodos , Cromatina/metabolismo , Endodesoxirribonucleases/metabolismo , Endorribonucleases/metabolismo , Células HeLa , Humanos , Peso Molecular
20.
Ecology ; 96(3): 642-53, 2015 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-26236861

RESUMO

Various environmental factors, including atmospheric CO2 (pCO2), regional climate, and fire, have been invoked as primary drivers of long-term variation in C4 grass abundance. Evaluating these hypotheses has been difficult because available paleorecords often lack information on past C4 grass abundance or potential environmental drivers. We analyzed carbon isotope ratios (delta13C) of individual grains of grass pollen in the sediments of two East African lakes to infer changes in the relative abundance of C3 vs. C4 grasses during the past 25 000 years. Results were compared with concurrent changes in pCO2, temperature, moisture balance, and fire activity. Our grass-pollen delta13C analysis reveals a dynamic history of grass-dominated vegetation in equatorial East Africa: C4 grasses have not consistently dominated lowland areas, and high-elevation grasses have not always been predominantly C3. On millennial timescales, C4 grass abundance does not correlate with charcoal influx at either site, suggesting that fire was not a major proximate control of the competitive balance between C3 and C4 grasses. Above the present-day treeline on Mt. Kenya, C4 grass abundance declined from an average of approximately 90% during the glacial period to less than approximately 60% throughout the Holocene, coincident with increases in pCO2 and temperature, and shifts in moisture balance. In the lowland savanna southeast of Mt. Kilimanjaro, C4 grass abundance showed no such directional trend, but fluctuated markedly in association with variation in rainfall amount and seasonal-drought severity. These results underscore spatiotemporal variability in the relative influence of pCO2 and climate on the interplay of C3 and C4 grasses and shed light on an emerging conceptual model regarding the expansion of C4-dominated grasslands in Earth's history. They also suggest that future changes in the C3/C4 composition of grass-dominated ecosystems will likely exhibit striking spatiotemporal variability as a result of varying combinations of environmental controls.


Assuntos
Dióxido de Carbono/análise , Clima , Fogo , Poaceae/fisiologia , Atmosfera , Carbono/química , Ciclo do Carbono , Isótopos de Carbono/metabolismo , Pradaria , Quênia , Paleontologia , Poaceae/química , Densidade Demográfica
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