CST does not evict elongating telomerase but prevents initiation by ssDNA binding

Area of Science:

• Molecular Biology
• Genetics
• Biochemistry

Background:

• The CST complex (CTC1-STN1-TEN1) plays a crucial role in telomere maintenance.
• CST is known to inhibit telomerase and facilitate C-strand synthesis by DNA polymerase alpha-primase (pol α-primase).
• Recent cryo-EM structure determination of human CST enabled the design of mutants to probe its function.

Purpose of the Study:

• To reexamine the mechanism by which CST inhibits telomerase.
• To test the primer sequestration model of CST-mediated telomerase inhibition.
• To investigate whether CST can terminate ongoing telomerase extension reactions.

Main Methods:

• Utilized DNA-binding mutants of the CST complex.
• Employed competitive binding simulations.
• Conducted pulse-chase telomerase extension assays in vitro.

Main Results:

• DNA-binding mutants of CST showed significantly reduced telomerase inhibition, supporting the primer sequestration model.
• CST did not terminate ongoing processive telomerase reactions, even when added to established reactions.
• The inhibitory effect of CST was independent of bound pol α-primase during ongoing reactions.

Conclusions:

• The findings strongly support the primer sequestration model for CST's inhibition of telomerase initiation.
• CST does not act as a processivity factor to terminate telomere extension.
• These results provide insights into the mechanism facilitating the switch from telomerase to pol α-primase activity for C-strand synthesis.