Synergism of the Two Myb Domains of Tay1 Protein Results in High Affinity Binding to Telomeres

Katarina Visacka,Ctirad Hofr,Smaranda Willcox,Ivona Necasova,Jana Pavlouskova,Regina Sepsiova,Michaela Wimmerova,Lucia Simonicova,Jozef Nosek,Jiri Fajkus,Jack D Griffith,Lubomir Tomaska

doi:10.1074/jbc.m112.385591

Abstract

Double-stranded regions of the telomeres are recognized by proteins containing Myb-like domains conferring specificity toward telomeric repeats. Although biochemical and structural studies revealed basic molecular principles involved in DNA binding, relatively little is known about evolutionary pathways leading to various types of Myb domain-containing proteins in divergent species of eukaryotes. Recently we identified a novel type of telomere-binding protein YlTay1p from the yeast Yarrowia lipolytica containing two Myb domains (Myb1, Myb2) very similar to the Myb domain of mammalian TRF1 and TRF2. In this study we prepared mutant versions of YlTay1p lacking Myb1, Myb2, or both Myb domains and found that YlTay1p carrying either Myb domain exhibits preferential affinity to both Y. lipolytica (GGGTTAGTCA)(n) and human (TTAGGG)(n) telomeric sequences. Quantitative measurements of the protein binding to telomeric DNA revealed that the presence of both Myb domains is required for a high affinity of YlTay1p to either telomeric repeat. Additionally, we performed detailed thermodynamic analysis of the YlTay1p interaction with its cognate telomeric DNA, which is to our knowledge the first energetic description of a full-length telomeric-protein binding to DNA. Interestingly, when compared with human TRF1 and TRF2 proteins, YlTay1p exhibited higher affinity not only for Y. lipolytica telomeres but also for human telomeric sequences. The duplication of the Myb domain region in YlTay1p thus produces a synergistic effect on its affinity toward the cognate telomeric sequence, alleviating the need for homodimerization observed in TRF-like proteins possessing a single Myb domain.

Highlights

In contrast to mammalian TRF1 and TRF2, yeast telomeric protein YlTay1 possesses two Myb domains
Whereas the growth of the transformants carrying the wild-type YlTAY1 (WT) on glucose was similar to the control strain, their growth was dramatically reduced on media containing galactose as a sole carbon source (Fig. 1B)
We tested the mutant version of YlTay1p lacking both putative Myb domains (⌬⌬) and found that their growth on galactose was restored, indicating that the DNA binding activity of YlTay1p is responsible for the growth inhibition (Fig. 1B)

Summary

Introduction

In contrast to mammalian TRF1 and TRF2, yeast telomeric protein YlTay possesses two Myb domains. We identified a novel type of telomere-binding protein YlTay1p from the yeast Yarrowia lipolytica containing two Myb domains (Myb, Myb2) very similar to the Myb domain of mammalian TRF1 and TRF2. Quantitative measurements of the protein binding to telomeric DNA revealed that the presence of both Myb domains is required for a high affinity of YlTay1p to either telomeric repeat. When compared with human TRF1 and TRF2 proteins, YlTay1p exhibited higher affinity for Y. lipolytica telomeres and for human telomeric sequences. The duplication of the Myb domain region in YlTay1p produces a synergistic effect on its affinity toward the cognate telomeric sequence, alleviating the need for homodimerization observed in TRF-like proteins possessing a single Myb domain

Methods

Results

Conclusion