Xenopus laevis Ctc1-Stn1-Ten1 (xCST) Protein Complex Is Involved in Priming DNA Synthesis on Single-stranded DNA Template in Xenopus Egg Extract

Background: The Ctc1-Stn1-Ten1 (CST) complex has been identified as a telomere-associated single-stranded (ss) DNA-binding protein complex. Results: De novo priming on ssDNA template in Xenopus egg extracts was inefficient in the absence of CST. Conclusion: CST regulates pre-RC (pre-replication complex)-independent DNA replication initiation. Significance: This study contributes to our understanding of the replication mechanism of telomere DNA. The Ctc1-Stn1-Ten1 (CST) complex is an RPA (replication protein A)-like protein complex that binds to single-stranded (ss) DNA. It localizes at telomeres and is involved in telomere end protection in mammals and plants. It is also known to stimulate DNA polymerase α-primase in vitro. However, it is not known how CST accomplishes these functions in vivo. Here, we report the identification and characterization of Xenopus laevis CST complex (xCST). xCST showed ssDNA binding activity with moderate preference for G (guanine)-rich sequences. xStn1-immunodepleted Xenopus egg extracts supported chromosomal DNA replication in in vitro reconstituted sperm nuclei, suggesting that xCST is not a general replication factor. However, the immunodepletion or neutralization of xStn1 compromised DNA synthesis on ssDNA template. Because primed ssDNA template was replicated in xStn1-immunodepleted extracts as efficiently as in control ones, we conclude that xCST is involved in the priming step on ssDNA template. These results are consistent with the current model that CST is involved in telomeric C-strand synthesis through the regulation of DNA polymerase α-primase.

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