Figure 1. DUBs activity is important for maintenance of DNA methylation, but not for DNA replication, in Xenopus egg extracts. (A,B) Sperm chromatin was incubated with interphase Xenopus egg extracts supplemented with buffer (+buffer), 20 μM Ub-VS (+Ub-VS), or 20 μM Ub-VS and 0.2 mg/ml ubiquitin (+Ub-VS +Ub). Prior to addition of sperm chromatin, radiolabeled [α-32P]dCTP (A) or S-[methyl-3H]-adenosyl-L-methionine (B) was added to extracts. Purified DNA samples were analyzed to determine the efficiency of DNA replication (A) and DNA methylation at 90 min after addition of sperm DNA (B), as described in Experimental Procedures. (C) Interphase egg extracts were treated as in (A) and chromatin fractions were isolated and subjected to immunoblotting using the antibodies indicated. (D) Mock- or Uhrf1-depleted interphase egg extracts were treated with Ub-VS and ubiquitin as in (A). ∆ Denotes immunodepletion of the protein indicated. The resultant chromatin fractions were subjected to immunoblotting using the antibodies indicated. Source data are available online for this figure.
Image published in: Yamaguchi L et al. (2017)
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