HERC2 inactivation abrogates nucleolar localization of RecQ helicases BLM and WRN

Mingzhang Zhu,Weixin Liang,Tomohiko Ohta,Wenwen Wu,Yasuo Miyoshi,Yukiko Togashi

doi:10.1038/s41598-020-79715-y

Mingzhang Zhu, Weixin Liang + Show 4 more

Open Access

https://doi.org/10.1038/s41598-020-79715-y

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Abstract

The nucleolus is a nuclear structure composed of ribosomal DNA (rDNA), and functions as a site for rRNA synthesis and processing. The rDNA is guanine-rich and prone to form G-quadruplex (G4), a secondary structure of DNA. We have recently found that HERC2, an HECT ubiquitin ligase, promotes BLM and WRN RecQ DNA helicases to resolve the G4 structure. Here, we report the role of HERC2 in the regulation of nucleolar localization of the helicases. Furthermore, HERC2 inactivation enhances the effects of CX-5461, an inhibitor of RNA polymerase I (Pol I)-mediated transcription of rRNA with an intrinsic G4-stabilizing activity. HERC2 depletion or homozygous deletion of the C-terminal HECT domain of HERC2 prevented the nucleolar localization of BLM and WRN, and inhibited relocalization of BLM to replication stress-induced nuclear RPA foci. HERC2 colocalized with fibrillarin and Pol I subunit RPA194, both of which are required for rRNA transcription. The HERC2 dysfunction enhanced the suppression of pre-rRNA transcription by CX-5461. These results suggest the effect of HERC2 status on the functions of BLM and WRN on rRNA transcription in the nucleolus. Since HERC2 is downregulated in numerous cancers, this effect may be clinically relevant considering the beneficial effects of CX-5461 in cancer treatments.

Highlights

The nucleolus is a non-membranous nuclear organelle, which is involved in ribosome biogenesis and is composed of ribosomal DNA
We report that HERC2 regulates the nucleolar localization of helicases and regulates the effect of CX-5461 on pre-rRNA transcription and cell viability
BLM interacts with polymerase I (Pol I) and is required for efficient rRNA transcription possibly through its helicase activity, as BLM can unwind granular component (GC)-rich ribosomal DNA (rDNA)-like substrates that form in the nucleolus and can normally inhibit progression of the Pol I c omplex[24]

Summary

Introduction

The nucleolus is a non-membranous nuclear organelle, which is involved in ribosome biogenesis and is composed of ribosomal DNA (rDNA) It serves as a site for rRNA synthesis and processing for ribosome assembly[1,2]. The nucleolus is critical in the cellular response to DNA damage, and incorporates substantial range of DNA damage repair proteins, including RecQ DNA helicases BLM and WRN2–5 While these proteins reside in the nucleolus as cellular stock for the damage response, they plays critical roles in the rRNA synthesis[2,3,4,5]. G4s play essential physiological roles, they may interrupt the process of replication or transcription if left unresolved To overcome these obstacles, cells harbor G4 unwinding helicases, such as PIF1, DNA2, FANCJ, DDX11, RTEL1, RHAU/DHX36, BLM and WRN. WRN interacts with Pol I and depletion of WRN results in decreased levels of rRNA transcription and ribosomal s ubunits[25,26]

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