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The Nucleosome Remodeling and Deacetylating Complex (NuRD) is ubiquitously expressed in all metazoans. It combines nucleosome remodeling and histone deacetylating activities to generate inaccessible chromatin structures and to repress gene transcription. NuRD is involved in the generation and maintenance of a wide variety of lineage-specific gene expression programs during differentiation and in differentiated cells. A close cooperation with a large number of lineage-specific transcription factors is key to allow NuRD to function in many distinct differentiation contexts. The molecular nature of this interplay between transcription factors and NuRD is complex and not well understood. This review uses hematopoiesis as a paradigm to highlight recent advances in our understanding of how transcription factors and NuRD cooperate at the molecular level during differentiation. A comparison of vertebrate and invertebrate systems serves to identify the conserved and fundamental concepts guiding functional interactions between transcription factors and NuRD. We also discuss how the transcription factor-NuRD axis constitutes a potential therapeutic target for the treatment of hemoglobinopathies.

The last review on transient-state kinetic methods in The Enzymes was published three decades ago (Johnson, K.A., 1992. The Enzymes, XX, 1-61). In that review the foundations were laid out for the logic behind the design and interpretation of experiments. In the intervening years the instrumentation has improved mainly in providing better sample economy and shorter dead times. More significantly, in 1992 we were just introducing methods for fitting data based on numerical integration of rate equations, but the software was slow and difficult to use. Today, advances in numerical integration methods for data fitting have led to fast and dynamic software, making it easy to fit data without simplifying approximations. This approach overcomes multiple disadvantages of traditional data fitting based on equations derived by analytical integration of rate equations, requiring simplifying approximations. Mechanism-based fitting using computer simulation resolves mechanisms by accounting for the concentration dependence of the rates and amplitudes of the reaction to find a set of intrinsic rate constants that reproduce the experimental data, including details about how the experiment was performed in modeling the data. Rather than discuss how to design and interpret rapid-quench and stopped-flow experiments individually, we now focus on how to fit them simultaneously so that the quench-flow data anchor the interpretation of fluorescence signals. Computer simulation streamlines the fitting of multiple experiments globally to yield a single unifying model to account for all available data.

DNA is under a variety of assaults. As a result, different damages accumulate on DNA. These include base changes, single-strand breaks and double-strand breaks. In this volume and also briefly in the following volume, we discuss DNA damage and double-strand breaks. In particular, we focus on double-strand breaks. We discuss types of double-strand breaks as well as methods to detect them. We also discuss how DNA breaks are formed.

Tritium is a radioisotope of hydrogen emitting low energy β-rays in disintegration to ³He. DNA molecules are damaged mainly by β-ray irradiation, and additional damages can be induced by break of chemical bond by nuclear transmutation to inert ³He. Deep knowledges of the mechanisms underlying DNA damages lead to better understanding of biological effects of tritium. This chapter reviews recent experimental and computer simulation activities on quantitative evaluation of damage rates by β-ray irradiation and nuclear transmutation. The rate of DNA double-strand breaks in tritiated water has been examined using a single molecule observation method. The effects of β-ray irradiation were not noticeable at the level of tritium concentration of ∼kBq/cm³, while the irradiation effects were clear at tritium concentrations of ∼MBq/cm³. The factors affecting on the DSB rate are discussed. A new image processing method for the automatic measurement of DNA length using OpenCV and deep learning is also introduced. The effects of tritium transmutation on hydrogen bonds acting between the two main strands of DNA have been examined using molecular dynamics simulations. The study showed that the collapsing of DNA structure by the transmutation can be quantitatively evaluated using the root mean square deviation of atomic positions.

Since the application of ultrasound for clinical diagnosis and therapeutic purposes has been increased rapidly, the effects of exposure to ultrasound on DNA molecules were studied. In this chapter, we introduced various effects of DNA damages caused by different conditions of exposure of ultrasound. Ultrasound with different sound pressure and pulse transmission conditions have been applied in our study. We discussed the threshold of sound pressure of ultrasound-induced DNA damages. Different kinds of pulses of ultrasound and microbubbles' influences on DNA double-strand breaks were also shown.

Irradiation of high Z elements such as iodine, gold, gadolinium with monochromatic X-rays causes photoelectric effects that include the release of Auger electrons. Decay of radioactive iodine such as I-123 and I-125 also results in multiple events and some involve the generation of Auger electrons. These electrons have low energy and travel only a short distance but have a strong effect on DNA damage including the generation of double-strand breaks. In this chapter, we focus on iodine and discuss various studies that used iodine-containing chemicals to generate Auger electrons and cause DNA double-strand breaks. First, DNA synthesis precursors containing iodine were used to place iodine on DNA. DNA binding dyes such as iodine Hoechst were investigated for Auger electron generation and DNA breaks. More recently, iodine containing nanoparticles were developed. We describe our study using tumor spheroids loaded with iodine nanoparticles and synchrotron-generated monochromatic X-rays. This study led to the demonstration that an optimum effect on DNA double-strand break formation is observed with a 33.2keV X-ray which is just above the K-edge energy of iodine.

Genomic DNA is organized three-dimensionally in the nucleus as chromatin. Recent accumulating evidence has demonstrated that chromatin organizes into numerous dynamic domains in higher eukaryotic cells, which act as functional units of the genome. These compacted domains facilitate DNA replication and gene regulation. Undamaged chromatin is critical for healthy cells to function and divide. However, the cellular genome is constantly threatened by many sources of DNA damage (e.g., radiation). How do cells maintain their genome integrity when subjected to DNA damage? This chapter describes how the compact state of chromatin safeguards the genome from radiation damage and chemical attacks. Together with recent genomics data, our finding suggests that DNA compaction, such as chromatin domain formation, plays a critical role in maintaining genome integrity. But does the formation of such domains limit DNA accessibility inside the domain and hinder the recruitment of repair machinery to the damaged site(s) during DNA repair? To approach this issue, we first describe a sensitive imaging method to detect changes in chromatin states in living cells (single-nucleosome imaging/tracking). We then use this method to explain how cells can overcome potential recruiting difficulties; cells can decompact chromatin domains following DNA damage and temporarily increase chromatin motion (∼DNA accessibility) to perform efficient DNA repair. We also speculate on how chromatin compaction affects DNA damage-resistance in the clinical setting.

By adapting the method of single molecular observation for individual DNAs, it will be shown that reliable analysis of double-strand breaks, DSBs, becomes possible for various kinds of damage sources. Single DNA above the size of several-tens kilo base-pairs exhibits the length scale above several μm, indicating that their whole conformation is visible with fluorescence microscopy by adding suitable fluoresce dye to the solution. Various examples of the quantitative evaluation on DSBs are described, together with the evaluation of the protective effects of anti-oxidants.

Ionizing radiation causes various types of DNA damage, such as single- (SSBs) and double-strand breaks (DSBs), nucleobase lesions, abasic sites (AP sites), and cross-linking between complementary strands of DNA or DNA and proteins. DSBs are among the most harmful type of DNA damage, inducing serious genetic effects such as cell lethality and mutation. Nucleobase lesions and AP sites, on the other hand, may be less deleterious and are promptly repaired by base excision repair (BER) pathways. Recently, biochemical approaches to quantify nucleobase lesions and AP sites have revealed certain types of non-strand break lesions as harmful DNA damage, called clustered DNA damage. Such clusters can retard nucleobase excision repair enzymes, and can sometimes be converted to DSBs by BER catalysis. This unique character of clustered DNA damage strongly depends on the spatial density of ionization or excitation events occurring at the track end of initial radiation or low energy secondary electrons. In particular, the photoelectric effect of elements comprising biological molecules, followed by emission of Auger electrons, are key factors in determining the future fate of each clustered damage site. This chapter describes biological studies of clustered nucleobase lesions with SSBs or AP sites, and mechanistical studies on core level excitation and Auger relaxation giving rise to clustered DNA damage.

The Enzymes series was initiated in 1950 with the publication of a book entitled, "The Enzymes: Chemistry and Mechanism of Action" edited by James B. Sumner and Karl Myerback. There are two parts, Part 1 and Part 2 and the book contains 78 chapters. Authors and chapter titles for Part 1 and Part 2 are listed.