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Tyrosine dephosphorylation of H2AX modulates apoptosis and survival decisions.

Cook PJ, Ju BG, Telese F, Wang X, Glass CK, Rosenfeld MG - Nature (2009)

Bottom Line: Life and death fate decisions allow cells to avoid massive apoptotic death in response to genotoxic stress.Here we report that a protein tyrosine phosphatase, EYA, is involved in promoting efficient DNA repair rather than apoptosis in response to genotoxic stress in mammalian embryonic kidney cells by executing a damage-signal-dependent dephosphorylation of an H2AX carboxy-terminal tyrosine phosphate (Y142).This post-translational modification determines the relative recruitment of either DNA repair or pro-apoptotic factors to the tail of serine phosphorylated histone H2AX (gamma-H2AX) and allows it to function as an active determinant of repair/survival versus apoptotic responses to DNA damage, revealing an additional phosphorylation-dependent mechanism that modulates survival/apoptotic decisions during mammalian organogenesis.

View Article: PubMed Central - PubMed

Affiliation: Howard Hughes Medical Institute School of Medicine, University of California, San Diego, California 92037, USA.

ABSTRACT
Life and death fate decisions allow cells to avoid massive apoptotic death in response to genotoxic stress. Although the regulatory mechanisms and signalling pathways controlling DNA repair and apoptosis are well characterized, the precise molecular strategies that determine the ultimate choice of DNA repair and survival or apoptotic cell death remain incompletely understood. Here we report that a protein tyrosine phosphatase, EYA, is involved in promoting efficient DNA repair rather than apoptosis in response to genotoxic stress in mammalian embryonic kidney cells by executing a damage-signal-dependent dephosphorylation of an H2AX carboxy-terminal tyrosine phosphate (Y142). This post-translational modification determines the relative recruitment of either DNA repair or pro-apoptotic factors to the tail of serine phosphorylated histone H2AX (gamma-H2AX) and allows it to function as an active determinant of repair/survival versus apoptotic responses to DNA damage, revealing an additional phosphorylation-dependent mechanism that modulates survival/apoptotic decisions during mammalian organogenesis.

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Loss of Eya leads to increased γH2AX-posititve apoptotic cells. (a) TUNEL staining reveals apoptotic cells within the developing kidney of Eya1−/− embryos at e11.5 not present in wild type littermates. (b) Abnormal morphology and loss of developing renal tubules (white arrows) within the urogenital ridge (red dotted line) in Eya1−/−embryos coincides with increased γH2AX-positive nuclei by immunostaining. (c) In culture, 293T human embryonic kidney cells depleted for Eya1/3 using siRNA displayed increased apoptotic response to hypoxia for 20hrs (2% O2). Cell counts were performed on TUNEL stained cells co-stained with DAPI in triplicate to identify the proportion of TUNEL-positive nuclei. Basal level of apoptosis under these conditions was 1.4% TUNEL-positive/total nuclei. Bar graphs represent mean +/− SEM of fold apoptotic cells normalized to control siRNA from triplicate samples. “*”p- <.05.
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Figure 1: Loss of Eya leads to increased γH2AX-posititve apoptotic cells. (a) TUNEL staining reveals apoptotic cells within the developing kidney of Eya1−/− embryos at e11.5 not present in wild type littermates. (b) Abnormal morphology and loss of developing renal tubules (white arrows) within the urogenital ridge (red dotted line) in Eya1−/−embryos coincides with increased γH2AX-positive nuclei by immunostaining. (c) In culture, 293T human embryonic kidney cells depleted for Eya1/3 using siRNA displayed increased apoptotic response to hypoxia for 20hrs (2% O2). Cell counts were performed on TUNEL stained cells co-stained with DAPI in triplicate to identify the proportion of TUNEL-positive nuclei. Basal level of apoptosis under these conditions was 1.4% TUNEL-positive/total nuclei. Bar graphs represent mean +/− SEM of fold apoptotic cells normalized to control siRNA from triplicate samples. “*”p- <.05.

Mentions: We noticed that increased apoptosis and loss of renal tubules seen in the developing kidney of Eya1−/− mouse embryos coincided with increased immunostaining for serine139-phosphorylated H2AX (γH2AX) (Supplementary Fig. 1, Fig. 1a, b). Nuclear phosphorylation of the histone variant H2AX was recently shown to be a crucial component of apoptosis induced by the activation the JNK/SAPK stress response pathway[10], in addition to having a well-studied role in DNA damage repair [11–14].


Tyrosine dephosphorylation of H2AX modulates apoptosis and survival decisions.

Cook PJ, Ju BG, Telese F, Wang X, Glass CK, Rosenfeld MG - Nature (2009)

Loss of Eya leads to increased γH2AX-posititve apoptotic cells. (a) TUNEL staining reveals apoptotic cells within the developing kidney of Eya1−/− embryos at e11.5 not present in wild type littermates. (b) Abnormal morphology and loss of developing renal tubules (white arrows) within the urogenital ridge (red dotted line) in Eya1−/−embryos coincides with increased γH2AX-positive nuclei by immunostaining. (c) In culture, 293T human embryonic kidney cells depleted for Eya1/3 using siRNA displayed increased apoptotic response to hypoxia for 20hrs (2% O2). Cell counts were performed on TUNEL stained cells co-stained with DAPI in triplicate to identify the proportion of TUNEL-positive nuclei. Basal level of apoptosis under these conditions was 1.4% TUNEL-positive/total nuclei. Bar graphs represent mean +/− SEM of fold apoptotic cells normalized to control siRNA from triplicate samples. “*”p- <.05.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2692521&req=5

Figure 1: Loss of Eya leads to increased γH2AX-posititve apoptotic cells. (a) TUNEL staining reveals apoptotic cells within the developing kidney of Eya1−/− embryos at e11.5 not present in wild type littermates. (b) Abnormal morphology and loss of developing renal tubules (white arrows) within the urogenital ridge (red dotted line) in Eya1−/−embryos coincides with increased γH2AX-positive nuclei by immunostaining. (c) In culture, 293T human embryonic kidney cells depleted for Eya1/3 using siRNA displayed increased apoptotic response to hypoxia for 20hrs (2% O2). Cell counts were performed on TUNEL stained cells co-stained with DAPI in triplicate to identify the proportion of TUNEL-positive nuclei. Basal level of apoptosis under these conditions was 1.4% TUNEL-positive/total nuclei. Bar graphs represent mean +/− SEM of fold apoptotic cells normalized to control siRNA from triplicate samples. “*”p- <.05.
Mentions: We noticed that increased apoptosis and loss of renal tubules seen in the developing kidney of Eya1−/− mouse embryos coincided with increased immunostaining for serine139-phosphorylated H2AX (γH2AX) (Supplementary Fig. 1, Fig. 1a, b). Nuclear phosphorylation of the histone variant H2AX was recently shown to be a crucial component of apoptosis induced by the activation the JNK/SAPK stress response pathway[10], in addition to having a well-studied role in DNA damage repair [11–14].

Bottom Line: Life and death fate decisions allow cells to avoid massive apoptotic death in response to genotoxic stress.Here we report that a protein tyrosine phosphatase, EYA, is involved in promoting efficient DNA repair rather than apoptosis in response to genotoxic stress in mammalian embryonic kidney cells by executing a damage-signal-dependent dephosphorylation of an H2AX carboxy-terminal tyrosine phosphate (Y142).This post-translational modification determines the relative recruitment of either DNA repair or pro-apoptotic factors to the tail of serine phosphorylated histone H2AX (gamma-H2AX) and allows it to function as an active determinant of repair/survival versus apoptotic responses to DNA damage, revealing an additional phosphorylation-dependent mechanism that modulates survival/apoptotic decisions during mammalian organogenesis.

View Article: PubMed Central - PubMed

Affiliation: Howard Hughes Medical Institute School of Medicine, University of California, San Diego, California 92037, USA.

ABSTRACT
Life and death fate decisions allow cells to avoid massive apoptotic death in response to genotoxic stress. Although the regulatory mechanisms and signalling pathways controlling DNA repair and apoptosis are well characterized, the precise molecular strategies that determine the ultimate choice of DNA repair and survival or apoptotic cell death remain incompletely understood. Here we report that a protein tyrosine phosphatase, EYA, is involved in promoting efficient DNA repair rather than apoptosis in response to genotoxic stress in mammalian embryonic kidney cells by executing a damage-signal-dependent dephosphorylation of an H2AX carboxy-terminal tyrosine phosphate (Y142). This post-translational modification determines the relative recruitment of either DNA repair or pro-apoptotic factors to the tail of serine phosphorylated histone H2AX (gamma-H2AX) and allows it to function as an active determinant of repair/survival versus apoptotic responses to DNA damage, revealing an additional phosphorylation-dependent mechanism that modulates survival/apoptotic decisions during mammalian organogenesis.

Show MeSH
Related in: MedlinePlus