Christophe E. Redon

GammaH2AX and cancer.

Histone H2AX phosphorylation on a serine four residues from the carboxyl terminus (producing γH2AX) is a sensitive marker for DNA double-strand breaks (DSBs). DSBs may lead to cancer but, paradoxically, are also used to kill cancer cells. Using γH2AX detection to determine the extent of DSB induction may help to detect precancerous cells, to stage cancers, to monitor the effectiveness of cancer therapies and to develop novel anticancer drugs.

γH2AX and cancer

Histone H2AX phosphorylation on a serine four residues from the carboxyl terminus (producing γH2AX) is a sensitive marker for DNA double-strand breaks (DSBs). DSBs may lead to cancer but, paradoxically, are also used to kill cancer cells. Using γH2AX detection to determine the extent of DSB induction may help to detect precancerous cells, to stage cancers, to monitor the effectiveness of cancer therapies and to develop novel anticancer drugs.

Histone H2A variants H2AX and H2AZ

Two of the nucleosomal histone families, H3 and H2A, have highly conserved variants with specialized functions. Recent studies have begun to elucidate the roles of two of the H2A variants, H2AX and H2AZ. H2AX is phosphorylated on a serine four residues from the carboxyl terminus in response to the introduction of DNA double-strand breaks, whether these breaks are a result of environmental insult, metabolic mistake, or programmed process. H2AZ appears to alter nucleosome stability, is partially redundant with nucleosome remodeling complexes, and is involved in transcriptional control.

AID is required to initiate Nbs1/γ-H2AX focus formation and mutations at sites of class switching

Class switch recombination (CSR) is a region-specific DNA recombination reaction that replaces one immunoglobulin heavy-chain constant region (Ch) gene with another. This enables a single variable (V) region gene to be used in conjunction with different downstream Ch genes, each having a unique biological activity. The molecular mechanisms that mediate CSR have not been defined, but activation-induced cytidine deaminase (AID), a putative RNA-editing enzyme, is required for this reaction. Here we report that the Nijmegen breakage syndrome protein (Nbs1) and phosphorylated H2A histone family member X (γ-H2AX, also known as γ-H2afx), which facilitate DNA double-strand break (DSB) repair, form nuclear foci at the Ch region in the G1 phase of the cell cycle in cells undergoing CSR, and that switching is impaired in H2AX-/- mice. Localization of Nbs1 and γ-H2AX to the Igh locus during CSR is dependent on AID. In addition, AID is required for induction of switch region (Sµ)-specific DNA lesions that precede CSR. These results place AID function upstream of the DNA modifications that initiate CSR.

Histone H2AX in DNA damage and repair.

No abstract available.

Role of oxidatively induced DNA lesions in human pathogenesis

Genome stability is essential for maintaining cellular and organismal homeostasis, but it is subject to many threats. One ubiquitous threat is from a class of compounds known as reactive oxygen species (ROS), which can indiscriminately react with many cellular biomolecules including proteins, lipids, and DNA to produce a variety of oxidative lesions. These DNA oxidation products are a direct risk to genome stability, and of particular importance are oxidative clustered DNA lesions (OCDLs), defined as two or more oxidative lesions present within 10 bp of each other. ROS can be produced by exposure of cells to exogenous environmental agents including ionizing radiation, light, chemicals, and metals. In addition, they are produced by cellular metabolism including mitochondrial ATP generation. However, ROS also serve a variety of critical cellular functions and optimal ROS levels are maintained by multiple cellular antioxidant defenses. Oxidative DNA lesions can be efficiently repaired by base excision repair or nucleotide excision repair. If ROS levels increase beyond the capacity of its antioxidant defenses, the cells DNA repair capacity can become overwhelmed, leading to the accumulation of oxidative DNA damage products including OCDLs, which are more difficult to repair than individual isolated DNA damage products. Here we focus on the induction and repair of OCDLs and other oxidatively induced DNA lesions. If unrepaired, these lesions can lead to the formation of mutations, DNA DSBs, and chromosome abnormalities. We discuss the roles of these lesions in human pathologies including aging and cancer, and in bystander effects.

H2AX: functional roles and potential applications.

Upon DNA double-strand break (DSB) induction in mammals, the histone H2A variant, H2AX, becomes rapidly phosphorylated at serine 139. This modified form, termed γ-H2AX, is easily identified with antibodies and serves as a sensitive indicator of DNA DSB formation. This review focuses on the potential clinical applications of γ-H2AX detection in cancer and in response to other cellular stresses. In addition, the role of H2AX in homeostasis and disease will be discussed. Recent work indicates that γ-H2AX detection may become a powerful tool for monitoring genotoxic events associated with cancer development and tumor progression.

Histone H2A phosphorylation controls Crb2 recruitment at DNA breaks, maintains checkpoint arrest, and influences DNA repair in fission yeast.

ABSTRACT Mammalian ATR and ATM checkpoint kinases modulate chromatin structures near DNA breaks by phosphorylating a serine residue in the carboxy-terminal tail SQE motif of histone H2AX. Histone H2A is similarly regulated in Saccharomyces cerevisiae. The phosphorylated forms of H2AX and H2A, known as γ-H2AX and γ-H2A, are thought to be important for DNA repair, although their evolutionarily conserved roles are unknown. Here, we investigate γ-H2A in the fission yeast Schizosaccharomyces pombe. We show that formation of γ-H2A redundantly requires the ATR/ATM-related kinases Rad3 and Tel1. Mutation of the SQE motif to AQE (H2A-AQE) in the two histone H2A genes caused sensitivity to a wide range of genotoxic agents, increased spontaneous DNA damage, and impaired checkpoint maintenance. The H2A-AQE mutations displayed a striking synergistic interaction with rad22Δ (Rad52 homolog) in ionizing radiation (IR) survival. These phenotypes correlated with defective phosphorylation of the checkpoint proteins Crb2 and Chk1 and a failure to recruit large amounts of Crb2 to damaged DNA. Surprisingly, the H2A-AQE mutations substantially suppressed the IR hypersensitivity of crb2Δ cells by a mechanism that required the RecQ-like DNA helicase Rqh1. We propose that γ-H2A modulates checkpoint and DNA repair through large-scale recruitment of Crb2 to damaged DNA. This function correlates with evidence that γ-H2AX regulates recruitment of several BRCA1 carboxyl terminus domain-containing proteins (NBS1, 53BP1, MDC1/NFBD1, and BRCA1) in mammals.

Yeast histone 2A serine 129 is essential for the efficient repair of checkpoint‐blind DNA damage

Cells maintain genomic stability by the coordination of DNA‐damage repair and cell‐cycle checkpoint control. In replicating cells, DNA damage usually activates intra‐S‐phase checkpoint controls, which are characterized by delayed S‐phase progression and increased Rad53 phosphorylation. We show that in budding yeast, the intra‐S‐phase checkpoint controls, although functional, are not activated by the topoisomerase I inhibitor camptothecin (CPT). In a CPT‐hypersensitive mutant strain that lacks the histone 2A (H2A) phosphatidylinositol‐3‐OH kinase (PI(3)K) motif at Ser 129 (h2a‐s129a), the hypersensitivity was found to result from a failure to process full‐length chromosomal DNA molecules during ongoing replication. H2A Ser 129 is not epistatic to the RAD24 and RAD9 checkpoint genes, suggesting a non‐checkpoint role for the H2A PI(3)K site. These results suggest that H2A Ser 129 is an essential component for the efficient repair of DNA double‐stranded breaks (DSBs) during replication in yeast, particularly of those DSBs that do not induce the intra‐S‐phase checkpoint.

Repair of Topoisomerase I-Mediated DNA Damage

Publisher Summary This chapter discusses the use of inhibitors of tyrosyl-DNA phosphodiesterase (Tdp1) and Chk1/2 in combination with Topoisomerase I (TopI) inhibitors. TopI is an abundant and essential enzyme. It is the selective target of camptothecins, which are effective anticancer agents. TopI–DNA cleavage complexes can also be trapped by various endogenous and exogenous DNA lesions, including mismatches, abasic sites, and carcinogenic adducts. Tdp1 is one of the repair enzymes for Top1–DNA covalent complexes. It forms a multiprotein complex that includes poly (Adenosine diphosphate (ADP)–ribose) polymerase (PARP). PARP-deficient cells are hypersensitive to camptothecins and functionally deficient for Tdp1. This chapter reviews the developments in several pathways involved in the repair of Top1 cleavage complexes and the role of Chk1 and Chk2 checkpoint kinases in the cellular responses to Top1 inhibitors. The genes conferring camptothecin hypersensitivity are compiled for humans, budding yeast, and fission yeast.