CTC1-STN1 terminates telomerase while STN1-TEN1 enables C-strand synthesis during telomere replication in colon cancer cells

Xuyang Feng,Shih-Jui Hsu,Jiajie Diao,Yongyao Wang,Anukana Bhattacharjee,Carolyn M Price

doi:10.1038/s41467-018-05154-z

Xuyang Feng, Shih-Jui Hsu + Show 4 more

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https://doi.org/10.1038/s41467-018-05154-z

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Telomerase elongates the telomeric G-strand to prevent telomere shortening through conventional DNA replication. However, synthesis of the complementary C-strand by DNA polymerase α is also required to maintain telomere length. Polymerase α cannot perform this role without the ssDNA binding complex CST (CTC1-STN1-TEN1). Here we describe the roles of individual CST subunits in telomerase regulation and G-overhang maturation in human colon cancer cells. We show that CTC1-STN1 limits telomerase action to prevent G-overhang overextension. CTC1−/− cells exhibit telomeric DNA damage and growth arrest due to overhang elongation whereas TEN1−/− cells do not. However, TEN1 is essential for C-strand synthesis and TEN1−/− cells exhibit progressive telomere shortening. DNA binding analysis indicates that CTC1-STN1 retains affinity for ssDNA but TEN1 stabilizes binding. We propose CTC1-STN1 binding is sufficient to terminate telomerase action but altered DNA binding dynamics renders CTC1-STN1 unable to properly engage polymerase α on the overhang for C-strand synthesis.

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Telomerase elongates the telomeric G-strand to prevent telomere shortening through conventional DNA replication
To better understand how CST performs its various roles in telomere replication, we generated human HCT116 cells with a conditional TEN1 gene disruption so we could compare the effects of TEN1 and CTC1 loss[4]
As the stability of individual CST subunits can depend on expression of the other two subunits[15,29,30], we examined whether TEN1 removal causes a decrease in STN1 or CTC1

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Introduction

Telomerase elongates the telomeric G-strand to prevent telomere shortening through conventional DNA replication. Polymerase α cannot perform this role without the ssDNA binding complex CST (CTC1-STN1-TEN1). We show that CTC1-STN1 limits telomerase action to prevent G-overhang overextension. We propose CTC1-STN1 binding is sufficient to terminate telomerase action but altered DNA binding dynamics renders CTC1-STN1 unable to properly engage polymerase α on the overhang for C-strand synthesis. TRF1 and TRF2 bind the TTAGGGAATCCC repeats of the telomere duplex, POT1 binds the 3ʹ ssDNA extension on the G-rich strand (termed the G-overhang) while TPP1 dimerizes with POT1 and links it to TRF1/2 via TIN2. Other players are required, including the ssDNA-binding trimeric complex CST (CTC1-STN1-TEN1) which participates in multiple aspects of telomere replication[4,5]. The DNA termini are processed by nucleases to generate the 3ʹ overhang necessary for telomerase action[7,8]

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