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Andrew S Belmont - One of the best experts on this subject based on the ideXlab platform.
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tyramide signal amplification mass spectrometry tsa ms ratio identifies nuclear Speckle proteins
Journal of Cell Biology, 2020Co-Authors: Joseph Dopie, Michael J Sweredoski, Annie Moradian, Andrew S BelmontAbstract:We present a simple ratio method to infer protein composition within cellular structures using proximity labeling approaches but compensating for the diffusion of free radicals. We used tyramide signal amplification (TSA) and label-free mass spectrometry (MS) to compare proteins in nuclear Speckles versus centromeres. Our "TSA-MS ratio" approach successfully identified known nuclear Speckle proteins. For example, 96% and 67% of proteins in the top 30 and 100 sorted proteins, respectively, are known nuclear Speckle proteins, including proteins that we validated here as enriched in nuclear Speckles. We show that MFAP1, among the top 20 in our list, forms droplets under certain circumstances and that MFAP1 expression levels modulate the size, stability, and dynamics of nuclear Speckles. Localization of MFAP1 and its binding partner, PRPF38A, in droplet-like nuclear bodies precedes formation of nuclear Speckles during telophase. Our results update older proteomic studies of nuclear Speckles and should provide a useful reference dataset to guide future experimental dissection of nuclear Speckle structure and function.
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gene expression amplification by nuclear Speckle association
Journal of Cell Biology, 2019Co-Authors: Jiah Kim, Nimish Khanna, Neha Chivukula Venkata, Gabriela Andrea Hernandez Gonzalez, Andrew S BelmontAbstract:Many active genes reproducibly position near nuclear Speckles, but the functional significance of this positioning is unknown. Here we show that HSPA1B BAC transgenes and endogenous Hsp70 genes turn on 2-4 min after heat shock (HS), irrespective of their distance to Speckles. However, both total HSPA1B mRNA counts and nascent transcript levels measured adjacent to the transgene are approximately twofold higher for Speckle-associated alleles 15 min after HS. Nascent transcript level fold-increases for Speckle-associated alleles are 12-56-fold and 3-7-fold higher 1-2 h after HS for HSPA1B transgenes and endogenous genes, respectively. Severalfold higher nascent transcript levels for several Hsp70 flanking genes also correlate with Speckle association at 37°C. Live-cell imaging reveals that HSPA1B nascent transcript levels increase/decrease with Speckle association/disassociation. Initial investigation reveals that increased nascent transcript levels accompanying Speckle association correlate with reduced exosome RNA degradation and larger Ser2p CTD-modified RNA polymerase II foci. Our results demonstrate stochastic gene expression dependent on positioning relative to a liquid-droplet nuclear compartment through "gene expression amplification."
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nuclear Speckle fusion via long range directional motion regulates Speckle morphology after transcriptional inhibition
Journal of Cell Science, 2019Co-Authors: Jiah Kim, Kyu Young Han, Nimish Khanna, Andrew S BelmontAbstract:ABSTRACT Although the formation of RNA-protein bodies has been studied intensively, their mobility and how their number and size are regulated are still poorly understood. Here, we show significantly increased mobility of nuclear Speckles after transcriptional inhibition, including long-range directed motion of one Speckle towards another Speckle, terminated by Speckle fusion, over distances up to 4 µm and with velocities between 0.2 µm/min and 1.5 µm/min. Frequently, three or even four Speckles follow very similar paths, with new Speckles appearing along the path followed by a preceding Speckle. Speckle movements and fusion events contribute to fewer, but larger, Speckles after transcriptional inhibition. These Speckle movements are not actin dependent, but occur within chromatin-depleted channels enriched with small granules containing the Speckle marker protein SON. Similar long-range Speckle movements and fusion events were observed after heat shock or heavy metal stress, and during late G2 and early prophase. Our observations suggest a mechanism for long-range, directional nuclear Speckle movements, contributing to overall regulation of nuclear Speckle number and size as well as overall nuclear organization. This article has an associated First Person interview with the first author of the paper.
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nuclear Speckle fusion via long range directional motion regulates the number and size of Speckles
bioRxiv, 2018Co-Authors: Jiah Kim, Kyu Young Han, Nimish Khanna, Andrew S BelmontAbstract:Although the formation of RNA-protein bodies has been studied intensively, their mobility and how their number and size are regulated are still poorly understood. Here, we show significant increased mobility of nuclear Speckles after transcriptional inhibition, including long-range directed motion of one Speckle towards another Speckle, terminated by Speckle fusion, over distances up to 4 um and with velocities between 0.2-1.5 micron/min. Frequently, 3 or even 4 Speckles follow very similar paths, with new Speckles appearing along the path followed by a preceding Speckle. Speckle movements and fusion events contribute to fewer but larger Speckles after transcriptional inhibition. These Speckle movements are not actin-dependent, but occur within chromatin-depleted channels enriched with small granules containing the Speckle-marker protein SON. Our observations suggest a mechanism for long-range, directed nuclear Speckle movements, contributing to overall regulation of nuclear Speckle number and size as well as overall nuclear organization.
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hsp70 transgene directed motion to nuclear Speckles facilitates heat shock activation
Current Biology, 2014Co-Authors: Nimish Khanna, Andrew S BelmontAbstract:Summary Association and disassociation of gene loci with respect to specific nuclear compartments accompany changes in gene expression, yet little is known concerning the mechanisms by which this occurs or its functional consequences. Previously, we showed that tethering acidic activators to a peripheral chromosome site led to movement of the chromosome site away from the nuclear periphery, but the physiological relevance of this movement was unclear [1]. Nuclear Speckles, or interchromatin granule clusters, are enriched in factors involved in RNA processing [2], and the association of a subset of active genes at their periphery suggests Speckles may play a role in gene expression [3, 4]. Here, we show an actin-dependent association of HSP70 transgenes with nuclear Speckles after heat shock. We visualized HSP70 transgenes moving curvilinearly toward nuclear Speckles over ∼0.5–6 μm distances at velocities of 1–2 μm min −1 . Chromatin stretching in the direction of movement demonstrates a force-generating mechanism. Transcription in nearly all cases increased noticeably only after initial contact with a nuclear Speckle. Moreover, blocking new HSP70 transgene/Speckle association by actin depolymerization prevented significant heat shock-induced transcriptional activation in transgenes not associated with Speckles, although robust transcriptional activation was observed for HSP70 transgenes associated with nuclear Speckles. Our results demonstrate the existence of a still-to-be-revealed machinery for moving chromatin in a direct path over long distances toward nuclear Speckles in response to transcriptional activation; moreover, this Speckle association enhances the heat shock activation of these HSP70 transgenes.
J L Beuzit - One of the best experts on this subject based on the ideXlab platform.
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Speckle temporal stability in xao coronagraphic images ii refine model for quasi static Speckle temporal evolution for vlt sphere
Astronomy and Astrophysics, 2013Co-Authors: P Martinez, M Kasper, A Costille, J F Sauvage, K Dohlen, P Puget, J L BeuzitAbstract:Context. Observing sequences have shown that the major noise source limitation in high-contrast imaging is due to the presence of quasi-static Speckles. The timescale on which quasi-stati c Speckles evolve, is determined by various factors, among others mechanical or thermal deformations. Aims. Understanding of these time-variable instrumental Speckles, and especially their interaction with other aberration s, referred to as the pinning effect, is paramount for the search of faint stellar companions. The temporal evolution of quasi-static Speckles is for instance required for a quantification of the gain expect ed when using angular differential imaging (ADI), and to determine the interval on which Speckle nulling techniques must be carried out. Methods. Following an early analysis of a time series of adaptively corrected, coronagraphic images obtained in a laboratory condition with the High-Order Test bench (HOT) at ESO Headquarters, we confirm our results with new measurements carried out with th e SPHERE instrument during its final test phase in Europe. The a nalysis of the residual Speckle pattern in both direct and differential coronagraphic images enables the characterization of the temporal stability of quasi-static Speckles. Data were obta ined in a thermally actively controlled environment reproducing realistic conditions encountered at the telescope. Results. The temporal evolution of the quasi-static wavefront error exhibits linear power law, which can be used to model quasi-static Speckle evolution in the context of forthcoming high-contrast imaging instruments, with implications for instrumentation (design, observing strategies, data reduction). Such a model can be used for instance to derive the timescale on which non-common path aberrations must be sensed and corrected. We found in our data that quasi-static wavefront error increases with∼0.7A per minute.
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Speckle temporal stability in xao coronagraphic images ii refine model for quasi static Speckle temporal evolution for vlt sphere
arXiv: Instrumentation and Methods for Astrophysics, 2013Co-Authors: P Martinez, M Kasper, A Costille, J F Sauvage, K Dohlen, P Puget, J L BeuzitAbstract:Observing sequences have shown that the major noise source limitation in high-contrast imaging is due to the presence of quasi-static Speckles. The timescale on which quasi-static Speckles evolve, is determined by various factors, among others mechanical or thermal deformations. Understanding of these time-variable instrumental Speckles, and especially their interaction with other aberrations, referred to as the pinning effect, is paramount for the search of faint stellar companions. The temporal evolution of quasi-static Speckles is for instance required for a quantification of the gain expected when using angular differential imaging (ADI), and to determine the interval on which Speckle nulling techniques must be carried out. Following an early analysis of a time series of adaptively corrected, coronagraphic images obtained in a laboratory condition with the High-Order Test bench (HOT) at ESO Headquarters, we confirm our results with new measurements carried out with the SPHERE instrument during its final test phase in Europe. The analysis of the residual Speckle pattern in both direct and differential coronagraphic images enables the characterization of the temporal stability of quasi-static Speckles. Data were obtained in a thermally actively controlled environment reproducing realistic conditions encountered at the telescope. The temporal evolution of the quasi-static wavefront error exhibits linear power law, which can be used to model quasi-static Speckle evolution in the context of forthcoming high-contrast imaging instruments, with implications for instrumentation (design, observing strategies, data reduction). Such a model can be used for instance to derive the timescale on which non-common path aberrations must be sensed and corrected. We found in our data that quasi-static wavefront error increases with ~0.7 angstrom per minute.
P Martinez - One of the best experts on this subject based on the ideXlab platform.
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Speckle temporal stability in xao coronagraphic images ii refine model for quasi static Speckle temporal evolution for vlt sphere
Astronomy and Astrophysics, 2013Co-Authors: P Martinez, M Kasper, A Costille, J F Sauvage, K Dohlen, P Puget, J L BeuzitAbstract:Context. Observing sequences have shown that the major noise source limitation in high-contrast imaging is due to the presence of quasi-static Speckles. The timescale on which quasi-stati c Speckles evolve, is determined by various factors, among others mechanical or thermal deformations. Aims. Understanding of these time-variable instrumental Speckles, and especially their interaction with other aberration s, referred to as the pinning effect, is paramount for the search of faint stellar companions. The temporal evolution of quasi-static Speckles is for instance required for a quantification of the gain expect ed when using angular differential imaging (ADI), and to determine the interval on which Speckle nulling techniques must be carried out. Methods. Following an early analysis of a time series of adaptively corrected, coronagraphic images obtained in a laboratory condition with the High-Order Test bench (HOT) at ESO Headquarters, we confirm our results with new measurements carried out with th e SPHERE instrument during its final test phase in Europe. The a nalysis of the residual Speckle pattern in both direct and differential coronagraphic images enables the characterization of the temporal stability of quasi-static Speckles. Data were obta ined in a thermally actively controlled environment reproducing realistic conditions encountered at the telescope. Results. The temporal evolution of the quasi-static wavefront error exhibits linear power law, which can be used to model quasi-static Speckle evolution in the context of forthcoming high-contrast imaging instruments, with implications for instrumentation (design, observing strategies, data reduction). Such a model can be used for instance to derive the timescale on which non-common path aberrations must be sensed and corrected. We found in our data that quasi-static wavefront error increases with∼0.7A per minute.
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Speckle temporal stability in xao coronagraphic images ii refine model for quasi static Speckle temporal evolution for vlt sphere
arXiv: Instrumentation and Methods for Astrophysics, 2013Co-Authors: P Martinez, M Kasper, A Costille, J F Sauvage, K Dohlen, P Puget, J L BeuzitAbstract:Observing sequences have shown that the major noise source limitation in high-contrast imaging is due to the presence of quasi-static Speckles. The timescale on which quasi-static Speckles evolve, is determined by various factors, among others mechanical or thermal deformations. Understanding of these time-variable instrumental Speckles, and especially their interaction with other aberrations, referred to as the pinning effect, is paramount for the search of faint stellar companions. The temporal evolution of quasi-static Speckles is for instance required for a quantification of the gain expected when using angular differential imaging (ADI), and to determine the interval on which Speckle nulling techniques must be carried out. Following an early analysis of a time series of adaptively corrected, coronagraphic images obtained in a laboratory condition with the High-Order Test bench (HOT) at ESO Headquarters, we confirm our results with new measurements carried out with the SPHERE instrument during its final test phase in Europe. The analysis of the residual Speckle pattern in both direct and differential coronagraphic images enables the characterization of the temporal stability of quasi-static Speckles. Data were obtained in a thermally actively controlled environment reproducing realistic conditions encountered at the telescope. The temporal evolution of the quasi-static wavefront error exhibits linear power law, which can be used to model quasi-static Speckle evolution in the context of forthcoming high-contrast imaging instruments, with implications for instrumentation (design, observing strategies, data reduction). Such a model can be used for instance to derive the timescale on which non-common path aberrations must be sensed and corrected. We found in our data that quasi-static wavefront error increases with ~0.7 angstrom per minute.
A Costille - One of the best experts on this subject based on the ideXlab platform.
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Speckle temporal stability in xao coronagraphic images ii refine model for quasi static Speckle temporal evolution for vlt sphere
Astronomy and Astrophysics, 2013Co-Authors: P Martinez, M Kasper, A Costille, J F Sauvage, K Dohlen, P Puget, J L BeuzitAbstract:Context. Observing sequences have shown that the major noise source limitation in high-contrast imaging is due to the presence of quasi-static Speckles. The timescale on which quasi-stati c Speckles evolve, is determined by various factors, among others mechanical or thermal deformations. Aims. Understanding of these time-variable instrumental Speckles, and especially their interaction with other aberration s, referred to as the pinning effect, is paramount for the search of faint stellar companions. The temporal evolution of quasi-static Speckles is for instance required for a quantification of the gain expect ed when using angular differential imaging (ADI), and to determine the interval on which Speckle nulling techniques must be carried out. Methods. Following an early analysis of a time series of adaptively corrected, coronagraphic images obtained in a laboratory condition with the High-Order Test bench (HOT) at ESO Headquarters, we confirm our results with new measurements carried out with th e SPHERE instrument during its final test phase in Europe. The a nalysis of the residual Speckle pattern in both direct and differential coronagraphic images enables the characterization of the temporal stability of quasi-static Speckles. Data were obta ined in a thermally actively controlled environment reproducing realistic conditions encountered at the telescope. Results. The temporal evolution of the quasi-static wavefront error exhibits linear power law, which can be used to model quasi-static Speckle evolution in the context of forthcoming high-contrast imaging instruments, with implications for instrumentation (design, observing strategies, data reduction). Such a model can be used for instance to derive the timescale on which non-common path aberrations must be sensed and corrected. We found in our data that quasi-static wavefront error increases with∼0.7A per minute.
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Speckle temporal stability in xao coronagraphic images ii refine model for quasi static Speckle temporal evolution for vlt sphere
arXiv: Instrumentation and Methods for Astrophysics, 2013Co-Authors: P Martinez, M Kasper, A Costille, J F Sauvage, K Dohlen, P Puget, J L BeuzitAbstract:Observing sequences have shown that the major noise source limitation in high-contrast imaging is due to the presence of quasi-static Speckles. The timescale on which quasi-static Speckles evolve, is determined by various factors, among others mechanical or thermal deformations. Understanding of these time-variable instrumental Speckles, and especially their interaction with other aberrations, referred to as the pinning effect, is paramount for the search of faint stellar companions. The temporal evolution of quasi-static Speckles is for instance required for a quantification of the gain expected when using angular differential imaging (ADI), and to determine the interval on which Speckle nulling techniques must be carried out. Following an early analysis of a time series of adaptively corrected, coronagraphic images obtained in a laboratory condition with the High-Order Test bench (HOT) at ESO Headquarters, we confirm our results with new measurements carried out with the SPHERE instrument during its final test phase in Europe. The analysis of the residual Speckle pattern in both direct and differential coronagraphic images enables the characterization of the temporal stability of quasi-static Speckles. Data were obtained in a thermally actively controlled environment reproducing realistic conditions encountered at the telescope. The temporal evolution of the quasi-static wavefront error exhibits linear power law, which can be used to model quasi-static Speckle evolution in the context of forthcoming high-contrast imaging instruments, with implications for instrumentation (design, observing strategies, data reduction). Such a model can be used for instance to derive the timescale on which non-common path aberrations must be sensed and corrected. We found in our data that quasi-static wavefront error increases with ~0.7 angstrom per minute.
P Puget - One of the best experts on this subject based on the ideXlab platform.
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Speckle temporal stability in xao coronagraphic images ii refine model for quasi static Speckle temporal evolution for vlt sphere
Astronomy and Astrophysics, 2013Co-Authors: P Martinez, M Kasper, A Costille, J F Sauvage, K Dohlen, P Puget, J L BeuzitAbstract:Context. Observing sequences have shown that the major noise source limitation in high-contrast imaging is due to the presence of quasi-static Speckles. The timescale on which quasi-stati c Speckles evolve, is determined by various factors, among others mechanical or thermal deformations. Aims. Understanding of these time-variable instrumental Speckles, and especially their interaction with other aberration s, referred to as the pinning effect, is paramount for the search of faint stellar companions. The temporal evolution of quasi-static Speckles is for instance required for a quantification of the gain expect ed when using angular differential imaging (ADI), and to determine the interval on which Speckle nulling techniques must be carried out. Methods. Following an early analysis of a time series of adaptively corrected, coronagraphic images obtained in a laboratory condition with the High-Order Test bench (HOT) at ESO Headquarters, we confirm our results with new measurements carried out with th e SPHERE instrument during its final test phase in Europe. The a nalysis of the residual Speckle pattern in both direct and differential coronagraphic images enables the characterization of the temporal stability of quasi-static Speckles. Data were obta ined in a thermally actively controlled environment reproducing realistic conditions encountered at the telescope. Results. The temporal evolution of the quasi-static wavefront error exhibits linear power law, which can be used to model quasi-static Speckle evolution in the context of forthcoming high-contrast imaging instruments, with implications for instrumentation (design, observing strategies, data reduction). Such a model can be used for instance to derive the timescale on which non-common path aberrations must be sensed and corrected. We found in our data that quasi-static wavefront error increases with∼0.7A per minute.
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Speckle temporal stability in xao coronagraphic images ii refine model for quasi static Speckle temporal evolution for vlt sphere
arXiv: Instrumentation and Methods for Astrophysics, 2013Co-Authors: P Martinez, M Kasper, A Costille, J F Sauvage, K Dohlen, P Puget, J L BeuzitAbstract:Observing sequences have shown that the major noise source limitation in high-contrast imaging is due to the presence of quasi-static Speckles. The timescale on which quasi-static Speckles evolve, is determined by various factors, among others mechanical or thermal deformations. Understanding of these time-variable instrumental Speckles, and especially their interaction with other aberrations, referred to as the pinning effect, is paramount for the search of faint stellar companions. The temporal evolution of quasi-static Speckles is for instance required for a quantification of the gain expected when using angular differential imaging (ADI), and to determine the interval on which Speckle nulling techniques must be carried out. Following an early analysis of a time series of adaptively corrected, coronagraphic images obtained in a laboratory condition with the High-Order Test bench (HOT) at ESO Headquarters, we confirm our results with new measurements carried out with the SPHERE instrument during its final test phase in Europe. The analysis of the residual Speckle pattern in both direct and differential coronagraphic images enables the characterization of the temporal stability of quasi-static Speckles. Data were obtained in a thermally actively controlled environment reproducing realistic conditions encountered at the telescope. The temporal evolution of the quasi-static wavefront error exhibits linear power law, which can be used to model quasi-static Speckle evolution in the context of forthcoming high-contrast imaging instruments, with implications for instrumentation (design, observing strategies, data reduction). Such a model can be used for instance to derive the timescale on which non-common path aberrations must be sensed and corrected. We found in our data that quasi-static wavefront error increases with ~0.7 angstrom per minute.