Yeast Hrq1 shares structural and functional homology with the disease-linked human RecQ4 helicase

Abstract

The five human RecQ helicases participate in multiple processes required to maintain genome integrity. Of these, the disease-linked RecQ4 is the least studied because it poses many technical challenges. We previously demonstrated that the yeast Hrq1 helicase displays similar functions to RecQ4 in vivo, and here, we report the biochemical and structural characterization of these enzymes. In vitro, Hrq1 and RecQ4 are DNA-stimulated ATPases and robust helicases. Further, these activities were sensitive to DNA sequence and structure, with the helicases preferentially unwinding D-loops. Consistent with their roles at telomeres, telomeric repeat sequence DNA also stimulated binding and unwinding by these enzymes. Finally, electron microscopy revealed that Hrq1 and RecQ4 share similar structural features. These results solidify Hrq1 as a true RecQ4 homolog and position it as the premier model to determine how RecQ4 mutations lead to genomic instability and disease.

Saccharomyces cerevisiae Hrq1 helicase activity is affected by the sequence but not the length of single-stranded DNA

“…Concerning DNA unwinding by Hrq1, a weak in vitro helicase activity has been reported for the S. pombe Hrq1 helicase [12], but the activity displayed by S. cerevisiae Hrq1 is more robust [8,9,13]. However, there is a discrepancy in the S. cerevisiae Hrq1 literature.…”

“…Hrq1 lacks the Sld2-like domain found in metazoan RecQ4 sub-family helicases, alleviating the issue of replication confounding the DNA repair function of the helicase. Furthermore, Hrq1 over-expression and purification produces abundant amounts of protein relative to RecQ4 [8]. Since its discovery, Hrq1 has been determined to play important roles at telomeres such as maintaining telomere length homeostasis in the absence of the known telomerase inhibitor Pif1 [9].…”

“…Since its discovery, Hrq1 has been determined to play important roles at telomeres such as maintaining telomere length homeostasis in the absence of the known telomerase inhibitor Pif1 [9]. Additionally, Hrq1 functions in the repair of DNA inter-strand crosslinks (ICL) in the same pathway as the ICL repair nuclease Pso2 [8,9]. Telomere maintenance and ICL repair are likewise associated with human RecQ4 [10,11].…”

“…Telomere maintenance and ICL repair are likewise associated with human RecQ4 [10,11]. In vitro, Hrq1 and RecQ4 helicase activity is stimulated by poly(dT) and telomeric repeat ssDNA [8]. Both helicases preferentially bind to Holliday junctions and D-Loop DNA structures, with the latter being a preferred substrate for unwinding.…”

“…In contrast, another group reports an "absolute requirement for a long 3'-tail (!70 nt) for efficient unwinding" by Hrq1 [13]. Similarly, it is unclear if Hrq1 lacks single-stranded DNA (ssDNA) annealing activity [8,9] or if it is an annealase like human RecQ4 [13]. The current dearth of Hrq1 literature amplifies these discrepancies and complicates follow-up research as proper substrate design is pivotal for accurately reconstituting biochemical pathways in vitro.…”