Abstract
POT1 is the single stranded telomeric overhang binding protein, and is part of the shelterin complex, a group of six proteins essential for proper telomere function. The reduction or abrogation of POT1 DNA binding activity in mammalian cells results in telomere elongation, or activation of the ATR DNA damage response at telomeres. Therefore, overhang binding represents the functionally relevant activity of POT1. To better understand the roles of POT1, we sought to isolate proteins that interact with the DNA binding domain of the protein. A yeast two-hybrid screen was implemented using a C-terminal truncation termed POT1DeltaC, retaining the DNA binding domain. This screen yielded a partial cDNA corresponding to TRIP6, a member of the LIM domain protein family. TRIP6 could co-immunoprecipitate with POT1, TRF2 and TIN2 in human cells, arguing for association with the whole shelterin complex, and was detected at telomeres by ChIP. TRIP6 depletion by siRNA led to the induction of telomere dysfunction induced foci (TIFs), indicating a role in telomere protection. A closely related LIM domain protein, LPP, was also found at telomeres and was also important for repressing the DNA damage response. We propose that TRIP6 and LPP are both required for telomere protection.
Highlights
Telomeres are essential chromosomal elements, which ensure proper replication and protection of chromosome ends
The screen was expected to yield clones that associate with the DNA binding domain of POT1, and to exclude TPP1, which interacts with the Cterminus of the molecule [15]
We describe a novel association between shelterin and LIM domain proteins at telomeres
Summary
Telomeres are essential chromosomal elements, which ensure proper replication and protection of chromosome ends. Telomeres prevent an inappropriate DNA damage response by recruiting a six-protein complex called shelterin, which is able to inhibit the induction of ATM and ATR responses [1] They are part of a feedback loop that regulates the ability of telomerase to add TTAGGG repeats preferentially to short telomeres [2]. POT1 itself is able to prevent or limit telomere elongation through its DNA binding activity [4], and TPP1 possesses a recruitment domain for telomerase providing a link between the enzyme and the chromosome’s end [5] It is with two N-terminal OB folds that POT1 binds the telomeric overhang, with high affinity and sequence specificity [6, 7]
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