Single-stranded telomere-binding protein employs a dual rheostat for binding affinity and specificity that drives function

Abstract

SignificanceProteins that bind nucleic acids are frequently categorized as being either specific or nonspecific, with interfaces to match that activity. In this study, we have found that a telomere-binding protein exhibits a degree of specificity for ssDNA that is finely tuned for its function, which includes specificity for G-rich sequences with some tolerance for substitution. Mutations of the protein that dramatically impact its affinity for single-stranded telomeric DNA are lethal, as expected; however, mutations that alter specificity also impact biological function. Unexpectedly, we found mutations that make the protein more specific are also deleterious, suggesting that specificity and nonspecificity in nucleic acid recognition may be achieved through more nuanced mechanisms than currently recognized.

Commentary: Single-stranded telomere-binding protein employs a dual rheostat for binding affinity and specificity that drives function

“…In summary, the results of Glustrom et al (2018) nicely complement the conclusions inferred from previously published studies on the puzzling diversity of yeast telomeres as well as on the evolution of DNA-binding proteins. In more general terms, they also provide additional evidence that increasing perfection of molecular recognition does not necessarily mean an optimal evolutionary strategy (Kováč, 1987; Bar-Even et al, 2011).…”

“…Recently, Glustrom et al performed an elegant systematic analysis of the effects of amino acid substitutions across the ssTBP of S. cerevisiae (Cdc13p) DNA-binding interface on its affinity and specificity toward heterogeneous telomeric repeats of the host cell (Glustrom et al, 2018). The authors showed that, as they expected, a subset of mutants exhibiting a significant loss in affinity in vitro also conferred a profound loss of viability in vivo .…”

Spontaneous replication fork collapse regulates telomere length homeostasis in wild type cells

“…Furthermore, a mutation in CDC13 (cdc13-F539A) that reduced the ability of the t-RPA complex to associate with telomeres had a substantial effect on replication fork collapse. This mutation, located in a DNA contact residue on the surface of the high affinity DNAbinding domain of Cdc13 (19), reduced DNA binding in vitro as well as association of Cdc13 with telomeric chromatin (ref; SI Appendix, Fig. S10).…”

“…The effect was particularly pronounced when comparing sub-populations of the shortest telomeres, as the fraction of telomeres that retained ≤ 125 bp of founder DNA increased from 9.2% in wild type to 31.5% in the cdc13-F539A strain (p < 0.0001). Since the cdc13-F539A strain does not have a resection defect (19), this argues that the increased loss of founder sequence DNA was the consequence of an elevated frequency of fork collapse, rather than increased terminus-specific erosion.…”

“…Cdc13 performs its role at telomeres in association with two other proteins (Stn1 and Ten1) as a heterotrimeric complex with striking structural and architectural similarities to the canonical RPA complex (17,18). Specificity for chromosome ends is mediated by a high affinity DNA binding domain in Cdc13, which exhibits exceptional sequence preference for single-stranded G-rich telomeric sequences in budding yeast (19) and closely related species (20), leading us to refer to it as a t-RPA complex. This contrasts with the genome-wide role performed by this conserved RPA-like complex in multiple other species (21)(22)(23)(24), consistent with the lack of strong specificity for telomeric DNA (25).…”