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Fig. 6. Chk1 is required for ionizing-radiation-induced checkpoint maintenance. (A) Asynchronously growing wild-type (chk1:ep) ( ${square}$ ), chk1 ${Delta}$ ( ${blacksquare}$ ) and chk1-ts1 ( ${circ}$ ) cells were incubated at 25°C for the duration of the experiment or shifted to 36°C at 15 minutes after irradiation. The proportion of binucleate cells was counted at the time points indicated by DAPI and calcafluor staining, and normalized to that of t₀ (binucleate index). In each case, shifting the culture to 36°C led to a chk1-independent decrease in the number of binucleate cells as a result the temperature-shift-induced heat shock. (B) The checkpoint response of asynchronously growing wild type (chk1:ep) ( ${square}$ ), chk1 ${Delta}$ ( ${blacksquare}$ ) and chk1-ts1 ( ${circ}$ ) to 150 Gy ionizing radiation. Cultures were incubated at 25°C for the duration of the experiment or shifted to 36°C at 15 minutes after irradiation. The percentage of binucleate cells was counted at the indicated timepoints. In the absence of Chk1 activity (chk1-ts1 and chk1 ${Delta}$ ), the percentage of binucleate cells increased more rapidly than wildtype (chk1:ep) controls. (C) Checkpoint proficiency of asynchronously growing wild-type (chk1:ep) ( ${square}$ ), chk1 ( ${blacksquare}$ ) and chk1-ts1 ( ${circ}$ ) to 150 Gy ionising radiation. Cultures were incubated at 25°C for the duration of the experiment, or shifted to 36°C at 15 minutes post irradiation. The proportion of aberrant mitoses was determined at the indicated time points. The proportion of aberrant mitoses is higher in the absence of Chk1 activity (chk1-ts1 and chk1 ${Delta}$ ).

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