Roles of Heterochromatin and Telomere Proteins in Regulation of Fission Yeast Telomere Recombination and Telomerase Recruitment

Lyne Khair,Lakxmi Subramanian,Bettina A Moser,Toru M Nakamura

doi:10.1074/jbc.m109.078840

Lyne Khair, Lakxmi Subramanian + Show 2 more

Open Access

https://doi.org/10.1074/jbc.m109.078840

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Abstract

When the telomerase catalytic subunit (Trt1/TERT) is deleted, a majority of fission yeast cells survives by circularizing chromosomes. Alternatively, a small minority survives by maintaining telomeric repeats through recombination among telomeres. The recombination-based telomere maintenance in trt1Delta cells is inhibited by the telomere protein Taz1. In addition, catalytically inactive full-length Trt1 (Trt1-CI) and truncated Trt1 lacking the T-motif and reverse transcriptase (RT) domain (Trt1-DeltaT/RT) can strongly inhibit recombination-based survival. Here, we investigated the effects of deleting the heterochromatin proteins Swi6 (HP1 ortholog) and Clr4 (Suv39 family of histone methyltransferases) and the telomere capping complex subunits Poz1 and Ccq1 on Taz1- and Trt1-dependent telomere recombination inhibition. The ability of Taz1 to inhibit telomere recombination did not require Swi6, Clr4, Poz1, or Ccq1. Although Swi6, Clr4, and Poz1 were dispensable for the inhibition of telomere recombination by Trt1-CI, Ccq1 was required for efficient telomere recruitment of Trt1 and Trt1-CI-dependent inhibition of telomere recombination. We also found that Swi6, Clr4, Ccq1, the checkpoint kinase Rad3 (ATR ortholog), and the telomerase regulatory subunit Est1 are all required for Trt1-DeltaT/RT to inhibit telomere recombination. However, because loss of Swi6, Clr4, Rad3, Ccq1, or Est1 did not significantly alter the recruitment efficiency of Trt1-DeltaT/RT to telomeres, these factors are likely to enhance the ability of Trt1-DeltaT/RT to inhibit recombination-based survival by contributing to the negative regulation of telomere recombination.

Highlights

Because taz1⌬ trt1⌬ cells are healthy and maintain stable telomeres by utilizing a recombination-based mechanism, mutations that lead to chromosome circularization in taz1⌬ trt1⌬ cells might identify positive regulators of telomere recombination
Because we had previously established that Taz1, Trt1-CI, and Trt1-⌬T/reverse transcriptase (RT) strongly inhibit telomere recombination, mutations that suppress chromosome circularization upon reintroduction of Taz1, Trt1-CI, or Trt1-⌬T/RT into taz1⌬ trt1⌬ cells may identify factors that contribute to the inhibition of telomere recombination
A subset of mutations that can suppress chromosome circularization induced by Trt1-CI or Trt1⌬T/RT may identify factors involved in the recruitment of telomerase to telomeres

Summary

Regulation of Telomere Recombination in Fission Yeast

The catalytic subunit of telomerase, known as TERT (telomerase reverse transcriptase) [2], is encoded by the trt1ϩ gene in fission yeast [15]. We have previously shown that the telomeric GT-rich repeat-specific double-stranded DNA-binding protein Taz plays a critical role in inhibiting recombination-based telomere maintenance in trt1⌬ cells [23] and that taz1⌬ trt1⌬ cells sustain robust growth with stable linear chromosomes [21, 23]. Reintroduction of catalytically inactive Trt (Trt1-CI) or a C-terminal-truncated Trt, which lacks the telomerase-specific T-motif and the reverse transcriptase domain (Trt1-⌬T/ RT), causes circularization of chromosomes in taz1⌬ trt1⌬ survivor cells, uncovering a RT-independent role for Trt in the inhibition of telomere recombination [23]. We tested if Taz1- or Trt1-dependent inhibition of telomere recombination requires the presence of heterochromatin proteins (Swi and Clr4) or Pot complex components (Ccq and Poz). Our investigations utilizing Trt1⌬T/RT implicate a subtle contribution of heterochromatin and the checkpoint kinase Rad3ATR in repression of telomere recombination in fission yeast

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DISCUSSION