Fig. 4. Localization and transcription kinetics of ATR are consistent with a role for ATR in the phosphorylation of H2AX in the sex body. (A) ATR colocalizes with H2AX in all of the mutants tested. Triple labeling of structurally preserved nuclei from wild-type (a-c), Atm^-/- Spo11^+/- (d-f), Spo11^-/- (g-i) and Atm^-/- Spo11^-/- (j-l) mice (two sets of littermates were analysed), with antibodies against SCP3 (red), ATR (green) and H2AX (blue). Images show the focal plane in which the H2AX-chromatin domain can be visualized. ATR consistently localizes to H2AX-positive chromatin domains, whether these correspond to the X-Y chromatin (as in wild-type and Atm^-/- Spo11^+/- spermatocytes) or to non-sex-chromatin regions (as in Spo11^-/- and Atm^-/- Spo11^-/- mutants). (B) ATR transcription peaks at the stage in prophase when the sex body can first be visualized. The curves represent absolute levels of the mRNAs encoding the three PIKKs (ATM, ATR and DNA-PKcs) and SCP1 (Sycp1, as a marker of meiotic progression) during the first wave of spermatogenesis. Data were extracted from previously published microarray data (Schultz et al., 2003). Triplicates from each developmental time point were averaged and normalized to 1 dpp. The table shows prophase-stage distribution at different times during the first wave of spermatogenesis determined on meiotic spreads [data from Goetz et al. (Goetz et al., 1984)].