Hrq1, a Homolog of the Human RecQ4 Helicase, Acts Catalytically and Structurally to Promote Genome Integrity

Abstract

Summary Human RecQ4 affects cancer and aging, but it is difficult to study because it is a fusion between a helicase and an essential replication factor. Budding yeast Hrq1 is homologous to the disease-linked helicase domain of RecQ4 and, like hRecQ4, was a robust 3’–5’ helicase. Additionally, Hrq1 had the unusual property of forming heptameric rings. Cells lacking Hrq1 exhibited two particularly dangerous DNA damage phenotypes: hypersensitivity to DNA inter-strand crosslinks (ICLs) and telomere addition to DNA breaks. Both activities are rare: their co-existing in a single protein is unprecedented. Resistance to ICLs required helicase activity, but suppression of telomere addition did not. Hrq1 also affected telomere length by a non-catalytic mechanism, as well as telomerase-independent telomere maintenance. As Hrq1 bound telomeres in vivo, it likely affects them directly. Thus, the tumor suppressing activity of RecQ4 could be due to a role in ICL repair and/or suppressing de novo telomere addition.

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

“…This is in disagreement with our previous work demonstrating that the same concentration of S. cerevisiae Hrq1 (50 nM) unwinds a fork substrate with only 25-nt ssDNA tails [8,9]. However, Hrq1 was generated in insect cells by baculovirus over-expression in Ref.…”

“…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.…”

“…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]. Additionally, Hrq1 functions in the repair of DNA inter-strand crosslinks (ICL) in the same pathway as the ICL repair nuclease Pso2 [8,9].…”

“…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].…”

“…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.…”