Histone H2AX Phosphorylation: A Marker for DNA Damage
- 23 July 2012
- book chapter
- Published by Springer Science and Business Media LLC
- Vol. 920, 613-626
- https://doi.org/10.1007/978-1-61779-998-3_40
Abstract
The DNA damage response can be initiated in response to a variety of stress signals that are encountered during physiological processes or in response to exogenous cues, such as ionizing radiation or DNA-damaging therapeutic agents. A number of methods have been developed to examine the morphological, biochemical, and molecular changes that take place during the DNA damage response. When cells are exposed to ionizing radiation or DNA-damaging chemotherapeutic agents, double-stranded breaks (DSBs) are generated that rapidly result in the phosphorylation of histone H2A variant H2AX. Because phosphorylation of H2AX at Ser 139 (γ-H2AX) is abundant, fast, and correlates well with each DSB, it is the most sensitive marker that can be used to examine the DNA damage produced and the subsequent repair of the DNA lesion. γ-H2AX can be detected by immunoblotting and immunostaining using microscopic or flow cytometric detection. Since γ-H2AX can be also generated during DNA replication, as a consequence of apoptosis, or as it is found associated with residual DNA damage, it is important to determine the kinetics, number, size, and morphology of γ-H2AX-associated foci. This chapter describes a few standard protocols that we have successfully used in our laboratory for a number of experimental systems, primarily hematologic and epithelial cells grown in culture.Keywords
This publication has 18 references indexed in Scilit:
- The DNA Damage Response: Making It Safe to Play with KnivesMolecular Cell, 2010
- γH2AX foci analysis for monitoring DNA double-strand break repair: Strengths, limitations and optimizationCell Cycle, 2010
- -H2AX in recognition and signaling of DNA double-strand breaks in the context of chromatinNucleic Acids Research, 2008
- Chapter 6 DNA Damage Response and ApoptosisMethods in Enzymology, 2008
- DNA-PK phosphorylates histone H2AX during apoptotic DNA fragmentation in mammalian cellsDNA Repair, 2006
- γ-H2AX Dephosphorylation by Protein Phosphatase 2A Facilitates DNA Double-Strand Break RepairMolecular Cell, 2005
- Evidence for a lack of DNA double-strand break repair in human cells exposed to very low x-ray dosesProceedings of the National Academy of Sciences of the United States of America, 2003
- ATM Phosphorylates Histone H2AX in Response to DNA Double-strand BreaksJournal of Biological Chemistry, 2001
- A critical role for histone H2AX in recruitment of repair factors to nuclear foci after DNA damageCurrent Biology, 2000
- DNA Double-stranded Breaks Induce Histone H2AX Phosphorylation on Serine 139Journal of Biological Chemistry, 1998