Abstract
SPO11 introduces double-strand breaks (DSBs) that trigger the phosphorylation of H2AX during meiotic prophase. In mice, SPO11 is strictly required for initiation of meiotic recombination and synapsis, yet SPO11 is still considered to be dispensable for sex-body formation in mouse spermatocytes. We provide conclusive evidence showing that functional SPO11, and consequently recombination and synapsis, are required for phosphorylation of H2AX in the X-Y chromatin and for sex-body formation in mouse spermatocytes. We investigated the role in meiosis of the three kinases [ATM (ataxia telangiectasia mutated), ATR (ataxia-telangiectasia- and Rad-3-related) and DNA-PKcs (DNA-dependent-protein-kinase catalytic subunit)] known to phosphorylate H2AX in mitotic cells. We found that DNA-PKcs can be ruled out as an essential kinase in this process, whereas ATM is strictly required for the chromatin-wide phosphorylation of H2AX occurring in leptotene spermatocytes in response to DSBs. Remarkably, we discovered that Spo11 heterozygosity can rescue the prophase-I-arrest characteristic of ATM-deficient spermatocytes. Characterization of the rescued Atm-/- Spo11+/- mutant indicates that ATM is dispensable for sex-body formation and phosphorylation of H2AX in this subnuclear domain. The co-localization of ATR, phosphorylated H2AX and the sex chromatin observed in the Atm-/- Spo11+/- mutant, along with ATR transcription kinetics during the first wave of spermatogenesis, confirm and expand recent findings indicating that ATR is the kinase involved in H2AX phosphorylation in the sex body.
MeSH terms
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Animals
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Ataxia Telangiectasia Mutated Proteins
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Cell Cycle Proteins / genetics
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Cell Cycle Proteins / physiology*
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Chromosome Pairing / genetics
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DNA-Activated Protein Kinase / deficiency
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DNA-Activated Protein Kinase / genetics
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DNA-Activated Protein Kinase / physiology
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DNA-Binding Proteins / deficiency
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DNA-Binding Proteins / genetics
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DNA-Binding Proteins / physiology*
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Endodeoxyribonucleases
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Esterases
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Gene Expression
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Heterozygote*
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Histones / metabolism
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Male
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Meiosis / genetics
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Meiosis / physiology
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Meiotic Prophase I / genetics
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Meiotic Prophase I / physiology*
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Mice
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Mice, Inbred C57BL
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Mice, Knockout
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Nuclear Proteins / deficiency
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Nuclear Proteins / genetics
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Nuclear Proteins / physiology
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Phosphorylation
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Protein Serine-Threonine Kinases / deficiency
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Protein Serine-Threonine Kinases / genetics
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Protein Serine-Threonine Kinases / physiology*
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Proteins / metabolism
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Proteins / physiology*
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RNA, Messenger / genetics
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Recombinant Proteins / metabolism
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Sex Chromatin / metabolism*
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Spermatocytes / cytology
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Spermatocytes / metabolism*
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Testis / cytology
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Testis / ultrastructure
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Tumor Suppressor Proteins / deficiency
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Tumor Suppressor Proteins / genetics
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Tumor Suppressor Proteins / physiology*
Substances
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Cell Cycle Proteins
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DNA-Binding Proteins
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H2AX protein, mouse
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Histones
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Nuclear Proteins
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Proteins
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RNA, Messenger
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Recombinant Proteins
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Tumor Suppressor Proteins
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Atr protein, mouse
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Ataxia Telangiectasia Mutated Proteins
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Atm protein, mouse
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DNA-Activated Protein Kinase
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Prkdc protein, mouse
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Protein Serine-Threonine Kinases
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Endodeoxyribonucleases
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Esterases
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meiotic recombination protein SPO11