Telomerase diffuses rapidly to find and engage telomeres in transient and stable interactions.
Major finding: Telomerase diffuses rapidly to find and engage telomeres in transient and stable interactions.
Approach: CRISPR/Cas9 genome editing allows for single-molecule live cell imaging to study telomerase dynamics.
Impact: Telomere dynamics suggest a model by which short telomeres may be preferentially elongated.
Telomerase is recruited to telomeres during the S-phase of the cell cycle, where it adds repetitive DNA sequences to single-stranded 3′ overhangs of chromosome ends, preventing telomere attrition. Telomerase recruitment to telomeres involves an interaction between a component of the shelterin complex, TPP1 (also known as ACD), and the catalytic component of telomerase, TERT. However, it is unclear how telomerase finds the telomeres requiring elongation. The use of live cell imaging to understand telomerase dynamics has been limited by the low abundance of telomerase, prompting Schmidt and colleagues to use CRISPR/Cas9 to enable single-molecule live cell imaging to visualize telomerase trafficking. Genome editing generated cell lines with labeling of the endogenous TERT locus, telomeres, and Cajal bodies (where telomerase is predominantly localized outside of S-phase). Live cell imaging revealed that telomerase diffused quickly throughout the nucleus, excluding nucleoli, with a small proportion bound to telomeres or Cajal bodies. Telomerase formed both frequent short, dynamic interactions, and rare static, long-lasting interactions with telomeres. During the short interactions, telomerase probed chromosome ends thousands of times during a single S-phase. Both the short and static interactions required association between TPP1 and the TEN-domain of TERT. The observed dynamic telomerase–telomere interactions suggest a model in which telomerase frequently and transiently interacts with TPP1 at telomeres, only rarely forming stable interactions sufficiently long for telomere elongation, when 3′ overhangs are available for binding. This provides a potential mechanism for preferential elongation of short telomeres where telomerase would be in closer proximity to the 3′ overhang.
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