1. |
- Klionsky, Daniel J., et al.
(author)
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Guidelines for the use and interpretation of assays for monitoring autophagy
- 2012
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In: Autophagy. - : Informa UK Limited. - 1554-8635 .- 1554-8627. ; 8:4, s. 445-544
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Research review (peer-reviewed)abstract
- In 2008 we published the first set of guidelines for standardizing research in autophagy. Since then, research on this topic has continued to accelerate, and many new scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Accordingly, it is important to update these guidelines for monitoring autophagy in different organisms. Various reviews have described the range of assays that have been used for this purpose. Nevertheless, there continues to be confusion regarding acceptable methods to measure autophagy, especially in multicellular eukaryotes. A key point that needs to be emphasized is that there is a difference between measurements that monitor the numbers or volume of autophagic elements (e.g., autophagosomes or autolysosomes) at any stage of the autophagic process vs. those that measure flux through the autophagy pathway (i.e., the complete process); thus, a block in macroautophagy that results in autophagosome accumulation needs to be differentiated from stimuli that result in increased autophagic activity, defined as increased autophagy induction coupled with increased delivery to, and degradation within, lysosomes (in most higher eukaryotes and some protists such as Dictyostelium) or the vacuole (in plants and fungi). In other words, it is especially important that investigators new to the field understand that the appearance of more autophagosomes does not necessarily equate with more autophagy. In fact, in many cases, autophagosomes accumulate because of a block in trafficking to lysosomes without a concomitant change in autophagosome biogenesis, whereas an increase in autolysosomes may reflect a reduction in degradative activity. Here, we present a set of guidelines for the selection and interpretation of methods for use by investigators who aim to examine macroautophagy and related processes, as well as for reviewers who need to provide realistic and reasonable critiques of papers that are focused on these processes. These guidelines are not meant to be a formulaic set of rules, because the appropriate assays depend in part on the question being asked and the system being used. In addition, we emphasize that no individual assay is guaranteed to be the most appropriate one in every situation, and we strongly recommend the use of multiple assays to monitor autophagy. In these guidelines, we consider these various methods of assessing autophagy and what information can, or cannot, be obtained from them. Finally, by discussing the merits and limits of particular autophagy assays, we hope to encourage technical innovation in the field.
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2. |
- Schael, S, et al.
(author)
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Precision electroweak measurements on the Z resonance
- 2006
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In: Physics Reports. - : Elsevier BV. - 0370-1573 .- 1873-6270. ; 427:5-6, s. 257-454
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Research review (peer-reviewed)abstract
- We report on the final electroweak measurements performed with data taken at the Z resonance by the experiments operating at the electron-positron colliders SLC and LEP. The data consist of 17 million Z decays accumulated by the ALEPH, DELPHI, L3 and OPAL experiments at LEP, and 600 thousand Z decays by the SLID experiment using a polarised beam at SLC. The measurements include cross-sections, forward-backward asymmetries and polarised asymmetries. The mass and width of the Z boson, m(Z) and Gamma(Z), and its couplings to fermions, for example the p parameter and the effective electroweak mixing angle for leptons, are precisely measured: m(Z) = 91.1875 +/- 0.0021 GeV, Gamma(Z) = 2.4952 +/- 0.0023 GeV, rho(l) = 1.0050 +/- 0.0010, sin(2)theta(eff)(lept) = 0.23153 +/- 0.00016. The number of light neutrino species is determined to be 2.9840 +/- 0.0082, in agreement with the three observed generations of fundamental fermions. The results are compared to the predictions of the Standard Model (SM). At the Z-pole, electroweak radiative corrections beyond the running of the QED and QCD coupling constants are observed with a significance of five standard deviations, and in agreement with the Standard Model. Of the many Z-pole measurements, the forward-backward asymmetry in b-quark production shows the largest difference with respect to its SM expectation, at the level of 2.8 standard deviations. Through radiative corrections evaluated in the framework of the Standard Model, the Z-pole data are also used to predict the mass of the top quark, m(t) = 173(+10)(+13) GeV, and the mass of the W boson, m(W) = 80.363 +/- 0.032 GeV. These indirect constraints are compared to the direct measurements, providing a stringent test of the SM. Using in addition the direct measurements of m(t) and m(W), the mass of the as yet unobserved SM Higgs boson is predicted with a relative uncertainty of about 50% and found to be less than 285 GeV at 95% confidence level. (c) 2006 Elsevier B.V. All rights reserved.
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3. |
- Abreu, P., et al.
(author)
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Measurement of the gluon fragmentation function and a comparison of the scaling violation in gluon and quark jets
- 2000
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In: European Physical Journal C. - : Springer Science and Business Media LLC. - 1434-6044 .- 1434-6052. ; 13:4, s. 573-589
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Journal article (peer-reviewed)abstract
- The fragmentation functions of quarks and gluons are measured in various three-jet topologies in Z decays from the full data set collected with the DELPHI detector at the Z resonance between 1992 and 995. The results at different values of transverse momentum-like scales are compared. A parameterization of the quark and gluon fragmentation functions at a fixed reference scale is given. The quark and gluon fragmentation functions show the predicted pattern of scaling violations. The scaling violation for quark jets as a function of a transverse momentum-like scale is in a good agreement with that observed in lower energy e+e- annihilation experiments. For gluon jets it appears to be significantly stronger. The scale dependences of the gluon and quark fragmentation functions agree with the prediction of the DGLAP evolution equations from which the colour factor ratio CA/CF is measured to be: CA/CF = 2.26 ± 0.09stat. ± 0.06sys. ± 0.12clus.,scale..
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4. |
- Abreu, P., et al.
(author)
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Study of dimuon production in photon-photon collisions and measurement of QED photon structure functions at LEP
- 2001
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In: European Physical Journal C. - : Springer Science and Business Media LLC. - 1434-6044 .- 1434-6052. ; 19:1, s. 15-28
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Journal article (peer-reviewed)abstract
- Muon pair production in the process e+e- → e+e- μ+μ- is studied using the data taken at LEP1 (√s ≃ mz) with the DELPHI detector during the years 1992-1995. The corresponding integrated luminosity is 138.5 pb-1. The QED predictions have been tested over the whole Q2 range accessible at LEP1 (from several GeV2/c4 to several hundred GeV2/c4) by comparing experimental distributions with distributions resulting from Monte Carlo simulations using various generators. Selected events are used to extract the leptonic photon structure function Fγ 2. Azimuthal correlations are used to obtain information on additional structure functions, Fγ A and Fγ B, which originate from interference terms of the scattering amplitudes. The measured ratios Fγ A/Fγ 2 and FγB/Fγ 2 are significantly different from zero and consistent with QED predictions.
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5. |
- Gerin, M., et al.
(author)
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Interstellar OH+, H2O+ and H3O+ along the sight-line to G10.6-0.4
- 2010
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In: Astronomy and Astrophysics. - : EDP Sciences. - 0004-6361 .- 1432-0746. ; 518:4, s. 110-
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Journal article (peer-reviewed)abstract
- We report the detection of absorption lines by the reactive ions OH+, H2O+ and H3O+ along the line of sight to the submillimeter continuum source G10.6$-$0.4 (W31C). We used the Herschel HIFI instrument in dual beam switch mode to observe the ground state rotational transitions of OH+ at 971 GHz, H2O+ at 1115 and 607 GHz, and H3O+ at 984 GHz. The resultant spectra show deep absorption over a broad velocity range that originates in the interstellar matter along the line of sight to G10.6$-$0.4 as well as in the molecular gas directly associated with that source. The OH+ spectrum reaches saturation over most velocities corresponding to the foreground gas, while the opacity of the H2O+ lines remains lower than 1 in the same velocity range, and the H3O+ line shows only weak absorption. For LSR velocities between 7 and 50 kms$^{-1}$ we estimate total column densities of $N$(OH+) $> 2.5 \times 10^{14}$ cm$^{-2}$, $N$(H2O+) $\sim 6 \times 10^{13}$ cm$^{-2}$ and $N$(H3O+) $\sim 4.0 \times 10^{13}$ cm$^{-2}$. These detections confirm the role of O$^+$ and OH$^+$ in initiating the oxygen chemistry in diffuse molecular gas and strengthen our understanding of the gas phase production of water. The high ratio of the OH+ by the H2O+ column density implies that these species predominantly trace low-density gas with a small fraction of hydrogen in molecular form.
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6. |
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7. |
- Klionsky, Daniel J., et al.
(author)
-
Guidelines for the use and interpretation of assays for monitoring autophagy in higher eukaryotes
- 2008
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In: Autophagy. - : Landes Bioscience. - 1554-8627 .- 1554-8635. ; 4:2, s. 151-175
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Research review (peer-reviewed)abstract
- Research in autophagy continues to accelerate,1 and as a result many new scientists are entering the field. Accordingly, it is important to establish a standard set of criteria for monitoring macroautophagy in different organisms. Recent reviews have described the range of assays that have been used for this purpose.2,3 There are many useful and convenient methods that can be used to monitor macroautophagy in yeast, but relatively few in other model systems, and there is much confusion regarding acceptable methods to measure macroautophagy in higher eukaryotes. A key point that needs to be emphasized is that there is a difference between measurements that monitor the numbers of autophagosomes versus those that measure flux through the autophagy pathway; thus, a block in macroautophagy that results in autophagosome accumulation needs to be differentiated from fully functional autophagy that includes delivery to, and degradation within, lysosomes (in most higher eukaryotes) or the vacuole (in plants and fungi). Here, we present a set of guidelines for the selection and interpretation of the methods that can be used by investigators who are attempting to examine macroautophagy and related processes, as well as by reviewers who need to provide realistic and reasonable critiques of papers that investigate these processes. This set of guidelines is not meant to be a formulaic set of rules, because the appropriate assays depend in part on the question being asked and the system being used. In addition, we emphasize that no individual assay is guaranteed to be the most appropriate one in every situation, and we strongly recommend the use of multiple assays to verify an autophagic response.
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8. |
- Knævelsrud, Helene, et al.
(author)
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Membrane remodeling by the PX-BAR protein SNX18 promotes autophagosome formation
- 2013
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In: Journal of Cell Biology. - : Rockefeller University Press. - 0021-9525 .- 1540-8140. ; 202:2, s. 331-349
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Journal article (peer-reviewed)abstract
- The membrane remodeling events required for autophagosome biogenesis are still poorly understood. Because PX domain proteins mediate membrane remodeling and trafficking, we conducted an imaging-based siRNA screen for autophagosome formation targeting human PX proteins. The PX-BAR protein SNX18 was identified as a positive regulator of autophagosome formation, and its Drosophila melanogaster homologue SH3PX1 was found to be required for efficient autophagosome formation in the larval fat body. We show that SNX18 is required for recruitment of Atg16L1-positive recycling endosomes to a perinuclear area and for delivery of Atg16L1- and LC3-positive membranes to autophagosome precursors. We identify a direct interaction of SNX18 with LC3 and show that the pro-autophagic activity of SNX18 depends on its membrane binding and tubulation capacity. We also show that the function of SNX18 in membrane tubulation and autophagy is negatively regulated by phosphorylation of S233. We conclude that SNX18 promotes autophagosome formation by virtue of its ability to remodel membranes and provide membrane to forming autophagosomes.
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9. |
- Neufeld, David A., et al.
(author)
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HERSCHEL OBSERVATIONS OF INTERSTELLAR CHLORONIUM
- 2012
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In: Astrophysical Journal. - 0004-637X .- 1538-4357. ; 748:1, s. 37-
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Journal article (peer-reviewed)abstract
- Using the Herschel Space Observatory's Heterodyne Instrument for the Far-Infrared, we have observed parachloronium (H2Cl+) toward six sources in the Galaxy. We detected interstellar chloronium absorption in foreground molecular clouds along the sight lines to the bright submillimeter continuum sources Sgr A (+50 km s(-1) cloud) and W31C. Both the para-(H2Cl+)-Cl-35 and para-(H2Cl+)-Cl-37 isotopologues were detected, through observations of their 1(11)-0(00) transitions at rest frequencies of 485.42 and 484.23 GHz, respectively. For an assumed ortho-to-para ratio (OPR) of 3, the observed optical depths imply that chloronium accounts for similar to 4%-12% of chlorine nuclei in the gas phase. We detected interstellar chloronium emission from two sources in the Orion Molecular Cloud 1: the Orion Bar photodissociation region and the Orion South condensation. For an assumed OPR of 3 for chloronium, the observed emission line fluxes imply total beam-averaged column densities of similar to 2 x 10(13) cm(-2) and similar to 1.2 x 10(13) cm(-2), respectively, for chloronium in these two sources. We obtained upper limits on the para-(H2Cl+)-Cl-35 line strengths toward H-2 Peak 1 in the Orion Molecular cloud and toward the massive young star AFGL 2591. The chloronium abundances inferred in this study are typically at least a factor similar to 10 larger than the predictions of steady-state theoretical models for the chemistry of interstellar molecules containing chlorine. Several explanations for this discrepancy were investigated, but none has proven satisfactory, and thus the large observed abundances of chloronium remain puzzling.
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10. |
- Neufeld, D.A., et al.
(author)
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Strong absorption by interstellar hydrogen fluoride: Herschel/HIFI observations of the sight-line to G10.6-0.4 (W31C)
- 2010
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In: Astronomy and Astrophysics. - : EDP Sciences. - 0004-6361 .- 1432-0746. ; 518:4, s. 108-
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Journal article (peer-reviewed)abstract
- We report the detection of strong absorption by interstellar hydrogen fluoride along the sight-line to the submillimeter continuum source G10.6-0.4 (W31C). We have used Herschel's HIFI instrument, in dual beam switch mode, to observe the 1232.4763 GHz J=1-0 HF transition in the upper sideband of the Band 5a receiver. The resultant spectrum shows weak HF emission from G10.6-0.4 at LSR velocities in the range -10 to -3 km/s, accompanied by strong absorption by foreground material at LSR velocities in the range 15 to 50 km/s. The spectrum is similar to that of the 1113.3430 GHz 1(11)-0(00) transition of para-water, although at some frequencies the HF (hydrogen fluoride) optical depth clearly exceeds that of para-H2O. The optically-thick HF absorption that we have observed places a conservative lower limit of 1.6E+14 cm-2 on the HF column density along the sight-line to G10.6-0.4. Our lower limit on the HF abundance, 6E-9 relative to hydrogen nuclei, implies that hydrogen fluoride accounts for between ~ 30 and 100% of the fluorine nuclei in the gas phase along this sight-line. This observation corroborates theoretical predictions that - because the unique thermochemistry of fluorine permits the exothermic reaction of F atoms with molecular hydrogen - HF will be the dominant reservoir of interstellar fluorine under a wide range of conditions.
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