The TPR-containing domain within Est1 homologs exhibits species-specific roles in telomerase interaction and telomere length homeostasis.
BACKGROUND: The first telomerase-associated protein (Est1) was isolated in yeast due to its essential role in telomere maintenance. The human counterparts EST1A, EST1B, and EST1C perform diverse functions in nonsense-mediated mRNA decay (NMD), telomere length homeostasis, and telomere transcription. Although Est1 and EST1A/B interact with the catalytic subunit of yeast and human telomerase (Est2 and TERT, respectively), the molecular determinants of these interactions have not been elaborated fully. RESULTS: To investigate the functional conservation of the EST1 protein family, we performed protein-protein interaction mapping and structure-function analysis. The domain in hEST1A most conserved between species, containing a TPR (tricotetrapeptide repeat), was sufficient for interaction of hEST1A with multiple fragments of hTERT including the N-terminus. Two mutations within the hTERT N-terminus that perturb in vivo function (NAAIRS(92), NAAIRS(122)) did not affect this protein interaction. ScEst1 hybrids containing the TPR of hEST1A, hEST1B, or hEST1C were expressed in yeast strains lacking EST1, yet they failed to complement senescence. Point mutations within and outside the cognate ScEst1 TPR, chosen to disrupt a putative protein interaction surface, resulted in telomere lengthening or shortening without affecting recruitment to telomeres. CONCLUSIONS: These results identify a domain encompassing the TPR of hEST1A as an hTERT interaction module. The TPR of S. cerevisiae Est1 is required for telomerase-mediated telomere length maintenance in a manner that appears separable from telomere recruitment. Discrete residues in or adjacent to the TPR of Est1 also regulate telomere length homeostasis.
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