Histone deacetylase 4 interacts with 53BP1 to mediate the DNA damage response

Anumber of proteins are recruited to nuclear foci upon exposure to double-strand DNA damage, including 53BP1 and Rad51, but the precise role of these DNA damage–induced foci remain unclear. Here we show in a variety of human cell lines that histone deacetylase (HDAC) 4 is recruited to foci with kinetics similar to, and colocalizes with, 53BP1 after exposure to agents causing double-stranded DNA breaks. HDAC4 foci gradually disappeared in repair-proficient cells but persisted in repair-deficient cell lines or cells irradiated with a lethal dose, suggesting that resolution of HDAC4 foci is linked to repair. Silencing of HDAC4 via RNA interference surprisingly also decreased levels of 53BP1 protein, abrogated the DNA damage–induced G2 delay, and radiosensitized HeLa cells. Our combined results suggest that HDAC4 is a critical component of the DNA damage response pathway that acts through 53BP1 and perhaps contributes in maintaining the G2 cell cycle checkpoint

Genome-wide determination of drug localization

A vast number of small-molecule ligands, including therapeutic drugs under development and in clinical use, elicit their effects by binding specific proteins associated with the genome. An ability to map the direct interactions of a chemical entity with chromatin genome-wide could provide new and important insights into chemical perturbation of cellular function. Here we describe a method that couples ligand-affinity capture and massively parallel DNA sequencing (Chem-seq) to identify the sites bound by small chemical molecules throughout the human genome. We show how Chem-seq can be combined with ChIP-seq to gain unique insights into the interaction of drugs with their target proteins throughout the genome of tumor cells. These methods provide a powerful approach to enhance understanding of therapeutic action and characterize the specificity of chemical entities that interact with DNA or genome-associated proteins

Derivation of Pre-X Inactivation Human Embryonic Stem Cells under Physiological Oxygen Concentrations

The presence of two active X chromosomes (XaXa) is a hallmark of the ground state of pluripotency specific to murine embryonic stem cells (ESCs). Human ESCs (hESCs) invariably exhibit signs of X chromosome inactivation (XCI) and are considered developmentally more advanced than their murine counterparts. We describe the establishment of XaXa hESCs derived under physiological oxygen concentrations. Using these cell lines, we demonstrate that (1) differentiation of hESCs induces random XCI in a manner similar to murine ESCs, (2) chronic exposure to atmospheric oxygen is sufficient to induce irreversible XCI with minor changes of the transcriptome, (3) the Xa exhibits heavy methylation of the XIST promoter region, and (4) XCI is associated with demethylation and transcriptional activation of XIST along with H3K27-me3 deposition across the Xi. These findings indicate that the human blastocyst contains pre-X-inactivation cells and that this state is preserved in vitro through culture under physiological oxygen.Susan WhiteheadHillel and Liliana Bachrac

Human Auditory Cortical Activation during Self-Vocalization

During speaking, auditory feedback is used to adjust vocalizations. The brain systems mediating this integrative ability have been investigated using a wide range of experimental strategies. In this report we examined how vocalization alters speech-sound processing within auditory cortex by directly recording evoked responses to vocalizations and playback stimuli using intracranial electrodes implanted in neurosurgery patients. Several new findings resulted from these high-resolution invasive recordings in human subjects. Suppressive effects of vocalization were found to occur only within circumscribed areas of auditory cortex. In addition, at a smaller number of sites, the opposite pattern was seen; cortical responses were enhanced during vocalization. This increase in activity was reflected in high gamma power changes, but was not evident in the averaged evoked potential waveforms. These new findings support forward models for vocal control in which efference copies of premotor cortex activity modulate sub-regions of auditory cortex

Quantum state preparation and macroscopic entanglement in gravitational-wave detectors

Long-baseline laser-interferometer gravitational-wave detectors are operating at a factor of 10 (in amplitude) above the standard quantum limit (SQL) within a broad frequency band. Such a low classical noise budget has already allowed the creation of a controlled 2.7 kg macroscopic oscillator with an effective eigenfrequency of 150 Hz and an occupation number of 200. This result, along with the prospect for further improvements, heralds the new possibility of experimentally probing macroscopic quantum mechanics (MQM) - quantum mechanical behavior of objects in the realm of everyday experience - using gravitational-wave detectors. In this paper, we provide the mathematical foundation for the first step of a MQM experiment: the preparation of a macroscopic test mass into a nearly minimum-Heisenberg-limited Gaussian quantum state, which is possible if the interferometer's classical noise beats the SQL in a broad frequency band. Our formalism, based on Wiener filtering, allows a straightforward conversion from the classical noise budget of a laser interferometer, in terms of noise spectra, into the strategy for quantum state preparation, and the quality of the prepared state. Using this formalism, we consider how Gaussian entanglement can be built among two macroscopic test masses, and the performance of the planned Advanced LIGO interferometers in quantum-state preparation

Search for gravitational wave bursts in LIGO's third science run

We report on a search for gravitational wave bursts in data from the three LIGO interferometric detectors during their third science run. The search targets subsecond bursts in the frequency range 100-1100 Hz for which no waveform model is assumed, and has a sensitivity in terms of the root-sum-square (rss) strain amplitude of hrss ~ 10^{-20} / sqrt(Hz). No gravitational wave signals were detected in the 8 days of analyzed data.Comment: 12 pages, 6 figures. Amaldi-6 conference proceedings to be published in Classical and Quantum Gravit

Ki67 Proliferation Index as a Tool for Chemotherapy Decisions During and After Neoadjuvant Aromatase Inhibitor Treatment of Breast Cancer: Results From the American College of Surgeons Oncology Group Z1031 Trial (Alliance)

To determine the pathologic complete response (pCR) rate in estrogen receptor (ER) –positive primary breast cancer triaged to chemotherapy when the protein encoded by the MKI67 gene (Ki67) level was > 10% after 2 to 4 weeks of neoadjuvant aromatase inhibitor (AI) therapy. A second objective was to examine risk of relapse using the Ki67-based Preoperative Endocrine Prognostic Index (PEPI)

Evidence That Gene Activation and Silencing during Stem Cell Differentiation Requires a Transcriptionally Paused Intermediate State

A surprising portion of both mammalian and Drosophila genomes are transcriptionally paused, undergoing initiation without elongation. We tested the hypothesis that transcriptional pausing is an obligate transition state between definitive activation and silencing as human embryonic stem cells (hESCs) change state from pluripotency to mesoderm. Chromatin immunoprecipitation for trimethyl lysine 4 on histone H3 (ChIP-Chip) was used to analyze transcriptional initiation, and 3′ transcript arrays were used to determine transcript elongation. Pluripotent and mesodermal cells had equivalent fractions of the genome in active and paused transcriptional states (∼48% each), with ∼4% definitively silenced (neither initiation nor elongation). Differentiation to mesoderm changed the transcriptional state of 12% of the genome, with roughly equal numbers of genes moving toward activation or silencing. Interestingly, almost all loci (98–99%) changing transcriptional state do so either by entering or exiting the paused state. A majority of these transitions involve either loss of initiation, as genes specifying alternate lineages are archived, or gain of initiation, in anticipation of future full-length expression. The addition of chromatin dynamics permitted much earlier predictions of final cell fate compared to sole use of conventional transcript arrays. These findings indicate that the paused state may be the major transition state for genes changing expression during differentiation, and implicate control of transcriptional elongation as a key checkpoint in lineage specification

Upper limits on the strength of periodic gravitational waves from PSR J1939+2134

The first science run of the LIGO and GEO gravitational wave detectors presented the opportunity to test methods of searching for gravitational waves from known pulsars. Here we present new direct upper limits on the strength of waves from the pulsar PSR J1939+2134 using two independent analysis methods, one in the frequency domain using frequentist statistics and one in the time domain using Bayesian inference. Both methods show that the strain amplitude at Earth from this pulsar is less than a few times $10^{-22}$.Comment: 7 pages, 1 figure, to appear in the Proceedings of the 5th Edoardo Amaldi Conference on Gravitational Waves, Tirrenia, Pisa, Italy, 6-11 July 200