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At least 775 records · Page 43

Dietary flavonoid derivatives enhance chemotherapeutic effect by inhibiting the DNA damage response pathway

Flavonoids are the most common group of polyphenolic compounds and abundant in dietary fruits and vegetables. Diet high in vegetables or dietary flavonoid supplements is associated with reduced mortality rate for patients with breast cancer. Many studies have been proposed for mechanisms linking flavonoids to improving chemotherapy efficacy in many types of cancers, but data on this issue is still limited. Herein, we report on a new mechanism through which dietary flavonoids inhibit DNA damage checkpoints and repair pathways. We found that dietary flavonoids could inhibit Chk1 phosphorylation and decrease clonogenic cell growth once breast cancer cells receive ultraviolet irradiation, cisplatin, or etoposide treatment. Since the ATR-Chk1 pathway mainly involves response to DNA replication stress, we propose that flavonoid derivatives reduce the side effect of chemotherapy by improving the sensitivity of cycling cells. Therefore, we propose that increasing intake of common dietary flavonoids is beneficial to breast cancer patients who are receiving DNA-damaging chemotherapy, such as cisplatin or etoposide-based therapy. - Highlights: • First report on inhibition of both DNA damage and repair by dietary flavonoids • Dietary flavonoids inhibit cisplatin- and UV-induced Chk1 phosphorylation. • Flavonoids combined with cisplatin or UV treatment show notable growth inhibition. • Promising treatment proposal for patients who are receiving adjuvant chemotherapy.

60 APPLIED LIFE SCIENCES↗

Wasatch Solar Project Final Report

The Wasatch Solar Project was a collaborative effort to conduct stakeholder engagement and provide technical assistance and analysis to support solar market development in Utah. The Wasatch Solar Project addressed four topic areas: 1) technical analysis to inform an assessment of the net benefits and costs of solar in Utah; 2) development of a 10-Year Solar Deployment Plan to create a clear path for continued solar deployment in Utah; 3) demonstration of the costs and resiliency benefits of solar and storage for resiliency at emergency response facilities or public facilities; and 4) technical assistance to create a replicable Commercial Property Assessed Clean Energy (C-PACE) financing program in Utah. The Wasatch Solar Project identified that new technical new technical challenges arise as nascent markets grow to higher penetrations of solar, requiring more detailed interconnection standards and improved efficiency in the interconnection process; that it is difficult for entities served by utilities that do not collect hourly usage data to assess the economic value of solar and storage; that despite significant resiliency benefits, the high cost of battery storage limits its adoption at healthcare and emergency response facilities in states like Utah that have low electricity rates; that administration of C-PACE financing at a local jurisdictional level can be complex, cumbersome, and a barrier to widespread utilization; and that investments in distributed generation can result in long-term savings for utility customers.

14 SOLAR ENERGY↗

Kivalina Biomass Reactor

This report summarizes work performed under DOE Award DE-EE00010149 to support the reliable operation of a community-scale biochar reactor system in Kivalina, Alaska. The project focused on improving sanitation and waste management in a remote community by assessing the installed system, identifying spare parts, defining key performance indicators (KPIs), preparing operator and maintenance manuals, and developing mobile reporting tools for operational data and KPI tracking. The team also produced training materials and recorded videos to support operator onboarding and continuity. The project demonstrated progress in system readiness, documentation, and digital reporting, while also identifying challenges common to remote deployments, including travel constraints, upstream system failures, and local resource limitations. This work provides a practical framework for improving the operation, monitoring, and future replication of biomass reactor systems in remote communities.

09 BIOMASS FUELS↗

Data for Development, Optimization, and Application of an Episomal Plasmid System for Rhodotorula toruloides

Rhodotorula toruloides is an emerging oleaginous yeast with strong potential as a microbial cell factory for the production of acetyl-CoA-derived bioproducts. However, engineering of this organism has been limited by the absence of a functional episomal plasmid system, a foundational genetic tool for rapid gene expression, pathway testing, and CRISPR-based genome engineering. Here, we report the first episomal plasmid system for R. toruloides . Through systematic screening of candidate autonomously replicating sequences (ARSs) from diverse sources, we identified multiple functional ARS elements and selected C63F4, a fragment derived from Contig 63 of R. toruloides CBS14, because of its stable performance. The resulting pC63F4 plasmid was maintained episomally, supported GFP reporter expression, exhibited a copy number of 2.39 ± 0.13, and showed good stability during long term cultivation. To overcome poor transformation efficiency, we developed a Cre-loxP-mediated in vivo re-circularization strategy that enabled reliable delivery of the episomal plasmid. Using this improved system, we demonstrated functional episomal expression of metabolic engineering genes and multi-gene pathways for the production of triacetic acid lactone, fatty alcohols, and limonene. Finally, we leveraged this platform to establish a redesigned CRISPR system that enables seamless genome editing in R. toruloides for the first time, while also simplifying marker recycling. Together, this work establishes a long-needed episomal plasmid platform and associated CRISPR toolkit that will accelerate metabolic engineering, synthetic biology, and fundamental studies in R. toruloides .

Gene Editing↗

Comparison of gamma-ray spectral analysis methods for thick-walled UF 6 cylinders

Three data analysis methods were compared for gamma ray spectra collected by planar and coaxial high-purity germanium detectors (HPGe) on thick-walled uranium hexafluoride cylinders. The cylinder contents range from natural to low-enriched uranium (4.95% 235 U). Each spectrum was analyzed with the traditional 185.7 keV enrichment meter method, a 185.7/1001 keV ratio method, and the Fixed Energy Response Function Analysis with Multiple Efficiency (FRAM) software that utilizes low- and high-energy gamma rays and X-rays. When results for all cylinders were compared, the enrichment meter method and the 185.7/1001 keV peak ratio method consistently provided the most accurate data for all detector types. The error on the enrichment calculations analyzed with FRAM did not meet the international target value whereas those calculated with the enrichment meter and ratio method did. Enrichments calculated by FRAM were found to skew to higher values on natural uranium and to lower values on low-enriched uranium. FRAM calculated the least-precise enrichments; replicate analyses of single cylinders resulted in 235 U values that varied by as much as 19%. FRAM does have the significant advantages in that it does not require the use of calibration cylinders, infinite thickness criteria are not necessary, and it can be applied to any HPGe detector. Finally, if calibration cylinders are available, the enrichment meter and 185.7/1001 peak ratio are the recommended methods as they are the most accurate and precise.

46 INSTRUMENTATION RELATED TO NUCLEAR SCIENCE AND ↗

Therapeutic doses of irradiation activate viral transcription and induce apoptosis in HIV-1 infected cells

The highly active antiretroviral therapy reduces HIV-1 RNA in plasma to undetectable levels. However, the virus continues to persist in the long-lived resting CD4{sup +} T cells, macrophages and astrocytes which form a viral reservoir in infected individuals. Reactivation of viral transcription is critical since the host immune response in combination with antiretroviral therapy may eradicate the virus. Using the chronically HIV-1 infected T lymphoblastoid and monocytic cell lines, primary quiescent CD4{sup +} T cells and humanized mice infected with dual-tropic HIV-1 89.6, we examined the effect of various X-ray irradiation (IR) doses (used for HIV-related lymphoma treatment and lower doses) on HIV-1 transcription and viability of infected cells. Treatment of both T cells and monocytes with IR, a well-defined stress signal, led to increase of HIV-1 transcription, as evidenced by the presence of RNA polymerase II and reduction of HDAC1 and methyl transferase SUV39H1 on the HIV-1 promoter. This correlated with the increased GFP signal and elevated level of intracellular HIV-1 RNA in the IR-treated quiescent CD4{sup +} T cells infected with GFP-encoding HIV-1. Exposition of latently HIV-1infected monocytes treated with PKC agonist bryostatin 1 to IR enhanced transcription activation effect of this latency-reversing agent. Increased HIV-1 replication after IR correlated with higher cell death: the level of phosphorylated Ser46 in p53, responsible for apoptosis induction, was markedly higher in the HIV-1 infected cells following IR treatment. Exposure of HIV-1 infected humanized mice with undetectable viral RNA level to IR resulted in a significant increase of HIV-1 RNA in plasma, lung and brain tissues. Collectively, these data point to the use of low to moderate dose of IR alone or in combination with HIV-1 transcription activators as a potential application for the “Shock and Kill” strategy for latently HIV-1 infected cells. - Highlights: • X-ray irradiation (IR) increases HIV-1 transcription in latently-infected cells. • IR enhances activating effect of bryostatin 1 on HIV-1 transcription in monocytes. • IR induces apoptosis in HIV-1 infected cells via phosphorylation of p53 Ser46. • IR of HIV-1 infected humanized mice increases HIV-1 RNA in plasma, lung and brain.

60 APPLIED LIFE SCIENCES↗

The role of signal transducer and activator of transcription 3 in Rift Valley fever virus infection

Rift Valley fever (RVF) is a zoonotic disease that can cause severe illness in humans and livestock, triggering spontaneous abortion in almost 100% of pregnant ruminants. In this study, we demonstrate that signal transducer and activator of transcription 3 (STAT3) is phosphorylated on its conserved tyrosine residue (Y705) following RVFV infection. This phosphorylation was dependent on a major virulence factor, the viral nonstructural protein NSs. Loss of STAT3 had little effect on viral replication, but rather resulted in cells being more susceptible to RVFV-induced cell death. Phosphorylated STAT3 translocated to the nucleus, coinciding with inhibition of fos, jun, and nr4a2 gene expression, and the presence of STAT3 and NSs at the nr4a2 promoter. NSs was found predominantly in the cytoplasm of STAT3 null cells, indicating that STAT3 influences NSs nuclear localization. Collectively, these data demonstrate that STAT3 functions in a pro-survival capacity through modulation of NSs localization. - Highlights: • STAT3 is phosphorylated on tyrosine residue 705 following RVFV infection. • Phosphorylation of STAT3 was dependent on the viral protein NSs. • STAT3 -/- MEFs were more susceptible to RVFV-induced cell death. • Loss of STAT3 led to an increase in pro-apoptotic gene expression. • STAT3 functions in a pro-survival capacity by modulation of NSs localization.

60 APPLIED LIFE SCIENCES↗

Processing behavior evolution of recycled polypropylene: An integrated experimental and Computer-Aided engineering simulation study

Polypropylene (PP) comprises 21% of global plastics production and 18% of plastics waste, yet less than 1% of solid-waste PP is recycled in the United States (U.S.), representing significant environmental and economic challenges. Mechanical recycling, the most prevalent recycling method, subject's materials to thermomechanical stresses, which typically degrade polymer properties, affecting the quality of polymer products. This study replicates the impact of mechanical recycling through multiple extrusion cycles to examine the effects on PP's processing behavior. Dynamic scanning calorimetry (DSC) measurements showed stable melting behavior across all processing conditions, while crystallization analysis exhibited consistent shifts in kinetic parameters. Rheological characterization demonstrated progressive viscosity reductions through successive cycles, particularly pronounced at elevated reprocessing temperatures. Here, the integration of this experimental data into injection molding simulations showed that recycled PP maintains viable processing characteristics. Our findings establish quantitative correlations between processing history and material behavior, enabling optimization of processing parameters directly rather than relying on trial-and-error approaches. While these results reflect idealized recycling conditions with minimal contamination, they provide a framework for understanding fundamental property evolution during mechanical recycling.

42 ENGINEERING↗

Analysis of Sludge Batch 10 Samples from Tank 51H (HTF-51-18-94, -95)

Savannah River National Laboratory characterized sludge samples from Tank 51H that is being used to generate Sludge Batch 10. The composite slurry sample from Tank 51H was analyzed and determined to contain a concentrated salt solution with a sodium concentration of 5.79 M (1.33E+05 mg/L) and insoluble sludge solids content of 7.20 wt% (using all nine replicates from the 3 wt% total solids measurements). Hydroxide is the main anionic species in the salt solution with a concentration of 3.00 M followed by nitrate (0.728 M) and nitrite (0.657 M). The sum of the major cations versus the sum of the major anions from the analysis of the filtered supernatant liquid phase shows a difference of <5% indicating good data quality. The results from the analysis of the digestions of the well mixed composite slurry sample show sodium to be the main component of the total dried solids of the sample followed by aluminum, iron, manganese, and thorium. Additional measurements of the weight percent total solids and density using hand shaking of the sample bottle and recirculation through a peristaltic pump indicate both methods provide adequate mixing of the sludge slurry sample.

12 MANAGEMENT OF RADIOACTIVE AND NON-RADIOACTIVE W↗

Fabrication of Nb3Sn by Magnetron Sputtering for Superconducting Radiofrequency Application

Particle accelerators are considered as an important device that has wide applications in cancer treatment, sterilizing waste, preserving foods, ion implantation in semiconductor industry, and in production of isotopes for medical applications. Superconducting radiofrequency (SRF) cavities are the building blocks of a linear particle accelerator. Current particle accelerators use niobium (Nb) superconductors as the sheet material to fabricate a single SRF cavity for particle acceleration. With better superconducting properties (critical temperature Tc ~ 18.3 K, superheating field Hsh~ 400 mT), Nb3Sn is considered a potential candidate in SRF technology. Magnetron sputtering is a promising deposition method to fabricate Nb3Sn thin films inside SRF cavities. Superconducting Nb3Sn films were fabricated on Nb and sapphire substrates by magnetron sputtering from a single stoichiometric Nb3Sn target, by multilayer sputtering of Nb and Sn followed by annealing, and by co-sputtering of Nb and Sn followed by annealing. The variation of morphological and superconducting properties was investigated for different substrate temperatures, annealing temperatures, annealing durations, and thicknesses. The film properties were characterized by X-ray diffraction (XRD), scanning electron microscopy (SEM), atomic force microscopy (AFM), and energy dispersive X-ray spectroscopy (EDS). The films had crystalline Nb3Sn structure without any presence of poor superconducting Nb6Sn5 and NbSn2 phases. The highest Tc of the films fabricated from the stoichiometric target, multilayer sputtering and co-sputtering were 17.44, 17.93, and 17.66 K respectively. Finally, a cylindrical sputter coater with two identical magnetrons was designed and commissioned to fabricate Nb3Sn films inside a 2.6 GHz SRF cavity. The magnetrons were installed facing opposite to each other in a custom designed vacuum chamber and multilayers of Nb and Sn films on 1 cm2 Nb substrates replicating the beam tubes and equator locations of the cavity and the coated multilayered films were annealed at 950 °C for 3 h. The XRD of the as?deposited and annealed films confirmed the formation of Nb3Sn after the annealing. The dissertation discusses the fabrication process, characterized results of the fabricated films, the design of the cylindrical sputter coater and the preliminary data obtained from the sputter coater.

Sayeed, Md Nizam↗

Clean Energy Innovation Ecosystems Discovery - Measuring Innovation Through Data Analytics (MIDAS)

As part of the Mission Innovation rollout at the Paris COP 21, 20 countries announced to double their R&D budgets over 5 years to accelerate lab-to-market clean energy innovations. These innovations must address technical urgency and navigate complex institutional, social and financial environments. To inform budget and program development, policymakers need to understand existing clean energy innovation ecosystems (IE) and their characteristics at the local, state, and regional levels within the US. Researchers at ORNL have implemented a ground-breaking application of natural language processing, link analysis, and other computational techniques to transform text and numerical data into metrics on clean energy innovation activity and geography. The 'Measuring Innovation through Data Analytics' (MIDAS) application uses data to determine where clean energy innovation ecosystems exist and the underlying scoring algorithm "ranks" the strength of the ecosystem for several clean energy technologies. The selection of the data sources is driven by the fact that a clean energy IE can be identified as an intersection of nascent clean technology efforts and financing mechanisms and related large companies located in a geographically concentrated area with an enabling environment to encourage commercialization and networking assets to facilitate communication. MIDAS is a machine-assisted methodology that provides the user with a replicable method to rapidly identify, quantify and characterize clean energy innovation ecosystems. This software application and the underlying datasets have the potential to address many important policy questions. The initial broad list of U.S. clean energy ecosystems, with geographic area, technology focus, and list and types of involved organizations can help describe regional tech activities and capabilities. Other potential applications include understanding the effect of private sector investments on ecosystem activity, and trends analysis.

Chinthavali, Supriya↗

Compilation of LLNL CUP-2 Data

The CUP-2 uranium ore concentrate (UOC) standard reference material, a powder, was produced at the Blind River uranium refinery of Eldorado Resources Ltd. in Canada in 1986. This material was produced as part of a joint effort by the Canadian Certified Reference Materials Project and the Canadian Uranium Producers Metallurgical Committee to develop a certified reference material for uranium concentration and the concentration of several impurity constituents. This standard was developed to satisfy the requirements of the UOC mining and milling industry, and was characterized with this purpose in mind. To produce CUP-2, approximately 25 kg of UOC derived from the Blind River uranium refinery was blended, homogenized, and assessed for homogeneity by X-ray fluorescence (XRF) analysis. The homogenized material was then packaged into bottles, containing 50 g of material each, and distributed for analysis to laboratories in 1986. The CUP-2 UOC standard was characterized by an interlaboratory analysis program involving eight member laboratories, six commercial laboratories, and three additional volunteer laboratories. Each laboratory provided five replicate results on up to 17 analytes, including total uranium concentration, and moisture content. The selection of analytical technique was left to each participating laboratory. Uranium was reported on an “as-received” basis; all other analytes (besides moisture content) were reported on a “dry-weight” basis. A bottle of 25g of CUP-2 UOC standard as described above was purchased by LLNL and characterized by the LLNL Nuclear Forensics Group. Non-destructive and destructive analytical techniques were applied to the UOC sample. Information obtained from short-term techniques such as photography, gamma spectrometry, and scanning electron microscopy were used to guide the performance of longer-term techniques such as ICP-MS. Some techniques, such as XRF and ICP-MS, provided complementary types of data. The results indicate that the CUP-2 standard has a natural isotopic ratio, and does not appear to have been isotopically enriched or depleted in any way, and was not contaminated by a source of uranium with a non-natural isotopic composition. Furthermore, the lack of 233 U and 236 U above the instrumental detection limit indicates that this sample was not exposed to a neutron flux, which would have generated one or both of these isotopes in measurable concentrations.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗

The Antiviral and Cancer Genomic DNA Deaminase APOBEC3H Is Regulated by an RNA-Mediated Dimerization Mechanism

Human APOBEC3H and homologous single-stranded DNA cytosine deaminases are unique to mammals. These DNA-editing enzymes function in innate immunity by restricting the replication of viruses and transposons. APOBEC3H also contributes to cancer mutagenesis. Here, in this paper, we address the fundamental nature of RNA in regulating human APOBEC3H activities. APOBEC3H co-purifies with RNA as an inactive protein, and RNase A treatment enables strong DNA deaminase activity. RNA-binding-defective mutants demonstrate clear separation of function by becoming DNA hypermutators. Biochemical and crystallographic data demonstrate a mechanism in which double-stranded RNA mediates enzyme dimerization. Additionally, APOBEC3H separation-of-function mutants show that RNA binding is required for cytoplasmic localization, packaging into HIV-1 particles, and antiviral activity. Overall, these results support a model in which structured RNA negatively regulates the potentially harmful DNA deamination activity of APOBEC3H while, at the same time, positively regulating its antiviral activity.

59 BASIC BIOLOGICAL SCIENCES↗

Combination therapy with lenvatinib and radiation significantly inhibits thyroid cancer growth by uptake of tyrosine kinase inhibitor

Highlights: • Lenvatinib combined with radiation significantly inhibits thyroid cancer growth. • Combined therapy induced apoptosis and G2/M phase cell cycle arrest. • Irradiation increased cellular uptake of lenvatinib into cancer cells. • Combined therapy suggests a new therapeutic strategy for advanced thyroid cancer. Although surgical treatment cures >90% of differentiated thyroid cancer (DTC) patients, the remaining patients, including advanced DTC cases, have poor clinical outcomes. These patients with inoperable disease have only two choices of radioactive iodine therapy and tyrosine kinase inhibitors such as lenvatinib, which have a high incidence of treatment-related adverse events and can only prolong progression free survival by approximately 5–15 months. In this study, we investigated the antitumor effects of combination therapy with lenvatinib and radiation (CTLR) for DTC. CTLR synergistically inhibited cell replication and colony formation in vitro and tumor growth in nude mice without apparent toxicities and suppressed the expression of proliferation marker (Ki-67). CTLR also induced apoptosis and G2/M phase cell cycle arrest. Moreover, quantitative analysis of the intracellular uptake of lenvatinib using liquid chromatography and mass spectrometry demonstrated that intracellular uptake of lenvatinib was significantly increased 48 h following irradiation. These data suggest that increased membrane permeability caused by irradiation increases the intracellular concentration of levatinib, contributing to the synergistic effect. This mechanism-based potential of combination therapy suggests a powerful new therapeutic strategy for advanced thyroid cancer with fewer side effects and might be a milestone for developing a regimen in clinical practice.

62 RADIOLOGY AND NUCLEAR MEDICINE↗

Modeling the Effects of Morphine-Altered Virus Specific Antibody Responses on HIV/SIV Dynamics

Abstract Drugs of abuse, such as opiates, have been widely associated with enhancing HIV replication, accelerating disease progression and diminishing host-immune responses, thereby making it harder to effectively manage HIV infection. It is thus important to study the effects of drugs of abuse on HIV-infection and immune responses. Here, we develop mathematical models that incorporate the effects of morphine-altered antibody responses on HIV/SIV dynamics. Based on fitting the model to experimental data from simian immunodeficiency virus (SIV) infections in control and morphine-addicted macaques, we found that two of the most significant effects of virus specific antibodies are neutralizing viral particles and enhancing viral clearance. Using our model, we quantified how morphine alters virus-specific antibody responses, and how this alteration affects the key components of virus dynamics such as infection rate, virus clearance, viral load, CD4 + T cell count, and CD4 + T cell loss in SIV-infected macaques under conditioning with morphine. We found that in a subpopulation of SIV-infected morphine addicted macaques, the presence of drugs of abuse may cause significantly diminished antibody responses, resulting in more severe infection with increased SIV infectivity, a decreased viral clearance rate, increased viral load, and higher CD4 + T cell loss.

59 BASIC BIOLOGICAL SCIENCES↗

Creation of a prototype biomimetic fish to better understand impact trauma caused by hydropower turbine blade strikes

Biomimetic model organisms could be useful surrogates for live animals in many applications if the models have sufficient biofidelity. One such application is for use in field and laboratory tests of fish mortality associated with passage through hydropower turbines. Laboratory trials suggest that blade strikes are especially injurious and often causes mortality when fish are struck by thinner blades moving at higher velocities. Dose-response relationships have been created from these data, but the exact relationship between fish mortality and the actual forces enacted on fish during simulated blade strike testing remains unknown. Here, we describe the methods used to create a prototype biomimetic model fish composed of ballistic gelatin and covered with a surrogate skin to better approximate the biomechanical properties of a fish body. Frozen fish were scanned with high-fidelity laser scanners, and a 3D-printed, reusable mold was created from which to cast our gelatin model. Computed tomography scan data, imaged directly or taken from online data repositories, were also successfully used to create CAD models for use in additive manufacturing of molds. One 3-axis accelerometer was embedded into the gelatin to compare accelerometer data to dose-response data from previous laboratory research on live fish. The resulting model ( i.e. , Gelfish) had a statistically indistinguishable tissue durometer to that of real fish tissue and preliminary blade strike impact testing suggested its overall flexibility was similar to that of live fish. Gelfish was designed with biofidelity as its guiding principle and our results suggest initial experimentation was successful. Future research will include replication of initial Gelfish test results, quantitative measurement of model flexibility relative to real fish, and inclusion of surrogate skeletal structures to enhance biofidelity. Use of more sophisticated sensors would also better quantify the physical forces of blade strike impact and help determine how said forces correlate with rates of mortality observed during tests on live fish.

36 MATERIALS SCIENCE↗

Data From Experiments on Bubbling Fluidization of Zeolite in a Rectangular Bubbling Fluidized Bed

Fluidization experiments were conducted in a lab-scale rectangular bubbling fluidized bed with the objective of generating a high-quality dataset for model validation and artificial intelligence/machine learning (AI/ML) training. Zeolite was chosen as the bed material, and the fluidizing medium was air as supplied by a compressor. Three different flow rates at the inlet were chosen such that the particles were fluidized but not elutriated from the system. The test matrix involved randomization and replicates to provide uncertainty estimates as well as four different batches of zeolite as the bed material. The quantities of interest obtained from this study were statistics of differential pressures, interface heights, and particle velocities. Considering all the components of the elaborate test plan, the results obtained were consistent and reproducible. Characterization tests were performed to estimate particle properties including size, density, coefficient of friction, coefficient of restitution, and minimum fluidization velocity. In addition, the angle of repose from granular discharge experiments has been reported to account for rolling friction, though its effect on the overall process is expected to be negligible.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗

Structural basis for recognition of H3K56-acetylated histone H3-H4 by the chaperone Rtt106

Dynamic variations in the structure of chromatin influence virtually all DNA-related processes in eukaryotes and are controlled in part by post-translational modifications of histones. One such modification, the acetylation of lysine 56 (H3K56ac) in the amino-terminal α-helix (αN) of histone H3, has been implicated in the regulation of nucleosome assembly during DNA replication and repair, and nucleosome disassembly during gene transcription. In Saccharomyces cerevisiae, the histone chaperone Rtt106 contributes to the deposition of newly synthesized H3K56ac-carrying H3-H4 complex on replicating DNA, but it is unclear how Rtt106 binds H3-H4 and specifically recognizes H3K56ac as there is no apparent acetylated lysine reader domain in Rtt106. Here, we show that two domains of Rtt106 are involved in a combinatorial recognition of H3-H4. An N-terminal domain homodimerizes and interacts with H3-H4 independently of acetylation while a double pleckstrin-homology (PH) domain binds the K56-containing region of H3. Affinity is markedly enhanced upon acetylation of K56, an effect that is probably due to increased conformational entropy of the αN helix of H3. Our data support a mode of interaction where the N-terminal homodimeric domain of Rtt106 intercalates between the two H3-H4 components of the (H3-H4) 2 tetramer while two double PH domains in the Rtt106 dimer interact with each of the two H3K56ac sites in (H3-H4) 2 . We show that the Rtt106-(H3-H4) 2 interaction is important for gene silencing and the DNA damage response.

59 BASIC BIOLOGICAL SCIENCES↗