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. 2020 Feb 7;12(2):389. doi: 10.3390/cancers12020389

Figure 3.

Figure 3

Impairment of the ATR-CHK1 signaling triggers a WRNIP1-mediated ATM activation. (A) WB analysis of chromatin binding of WRNIP1, performed as in Figure 2A. WSWRN cells were transfected with GFP, Claspin, or TopBP1 siRNA for 48 h and treated with Aph. The membrane was probed with the indicated antibodies. The normalized ratio of the chromatin-bound WRNIP1/LAMIN B1 is reported; (B) IF analysis of cells depleted of Claspin or TopBP1 as in (A) and stained for pATM (S1981). Bar graph shows pATM intensity per nucleus. Error bars represent standard error. WB using the indicated antibodies confirms depletion of Claspin and TopBP1; (C) WB analysis of the presence of activated, i.e., phosphorylated, CHK1 assessed using S345 phospho-specific antibody (pS345) in WSWRN cells depleted for TopBP1 as in (A) and treated with Aph. ATMi was added 1 h prior to Aph and used as a negative control. The membrane was probed with the indicated antibodies. The normalized ratio of the phosphorylated CHK1/total CHK1 is given.