Elsevier

Neurobiology of Disease

Volume 73, January 2015, Pages 220-228
Neurobiology of Disease

Repression of rRNA transcription by PARIS contributes to Parkinson's disease

https://doi.org/10.1016/j.nbd.2014.10.003Get rights and content

Highlights

  • PARIS interacts with MYBBP1A.

  • PARIS occupies rDNA promoter and represses its transcription.

  • Overexpression of PARIS leads to p53 upregulation, a marker of nucleolar stress.

  • Reduction of rRNA and increase of p53 in conditional parkin KO and sporadic PD patients

Abstract

The nucleolus is a compartment for the transcription of ribosomal RNA (rRNA) and assembly of ribosome subunits. Dysregulation of the nucleolus is considered to be a cellular stress event associated with aging and neurodegenerative disease, including Parkinson's disease (PD). We previously demonstrated that PARIS (PARkin Interacting Substrate, ZNF746) transcriptionally suppresses peroxisome proliferator-activated receptor gamma (PPARγ) coactivator-1α (PGC-1α) in PD and its accumulation results in selective dopaminergic neuronal death. However, functional knowledge of PARIS is limited, and no other studies have been performed to elucidate its function. Here, we used tandem-affinity purification to identify the binding partners of PARIS, showing that PARIS interacts with 160-kDa Myb-binding protein 1α (MYBBP1A), which suppresses rRNA transcription and the rRNA editing process. Interestingly, PARIS was also found to interact with the components of RNA polymerase I, occupied the promoter of rDNA, and suppressed rDNA transcription in vivo. Accordingly, we observed a reduction of rRNA levels and increased expression of p53, a molecular marker of nucleolar stress, in the substantia nigra of conditional parkin knockout mice, AAV-mediated PARIS overexpression mice, and in patients with sporadic PD. Together, our results suggest that dysfunction of the Parkin–PARIS pathway may play a deleterious role in rRNA transcription and contribute to PD pathogenesis.

Introduction

The major function of the nucleolus is to transcribe ribosomal RNA (rRNA) and to assemble ribosome subunits; this process must be tightly regulated to achieve proper cellular proliferation and growth (Boisvert et al., 2007). Indeed, the biogenesis of rRNA is a complex process that requires the transcription of rDNA genes, nucleolytic cleavage events, and chemical modification of precursor rRNAs, ultimately leading to the assembly of mature ribosomes (Boulon et al., 2010). In addition, the nucleolus is involved in directing the cellular response to a diverse array of stressors (Boulon et al., 2010). Nucleolar stress is defined as decreased rRNA synthesis and nucleolar disruption, and is considered as a primary sign of cellular stress associated with aging and neurodegenerative disorders. Notably, epigenetic silencing of rDNA has been observed during the early stages of Alzheimer's disease (AD) (Pietrzak et al., 2011). In addition, altered postmortem nucleolar size and nucleolar damage have also been observed in several neurodegenerative diseases, including Parkinson's disease (PD) (Hetman and Pietrzak, 2012, Rieker et al., 2011). Furthermore, mutation (L166P) of the PD-associated gene DJ-1 causes its misfolding, resulting in alteration of rRNA biogenesis by inhibiting TRAF and TNF Receptor Associated Protein (TTRAP) localization to the nucleolus (Vilotti et al., 2012). Despite the emerging evidence of nucleolar stress in PD, the exact mechanism of neuronal degeneration remains poorly understood.

The 160-kDa Myb-binding protein 1α (MYBBP1A) was recently shown to interact with both the RNA polymerase 1 complex and the ribosome biogenesis machinery to repress rRNA gene transcription. In addition, the absence of MYBBP1A leads to the accumulation of rRNA precursors and cell cycle arrest (Hochstatter et al., 2012). Furthermore, nucleolar stress induces translocation of MYBBP1A from the nucleolus to the nucleoplasm and enhances p53 activity (Kuroda et al., 2011), and MYBBP1A is required to activate p53 by its acetylation and tetramerization under nucleolar stress (Ono et al., 2014).

Previously, we identified a new parkin-interacting substrate, PARIS (ZNF746), and suggested its involvement in the pathomechanism of neuronal dysfunction that occurs after inactivation of parkin in PD. PARIS is a major transcriptional repressor of peroxisome proliferator-activated receptor gamma (PPARγ) coactivator-1α (PGC-1α) expression and accumulation of PARIS may lead to dopaminergic neuronal death via suppression of mitochondrial biogenesis, ROS defense, and oxidative phosphorylation in pathogenic condition (Shin et al., 2011). Interestingly, MYBBP1A has also been found to act as a transcriptional suppressor of PGC-1α (Fan et al., 2004). Thus, we hypothesized that PARIS might play a new function in rRNA transcription and is associated with the nucleolar stress observed in PD pathogenesis. Here, we report for the first time that PARIS co-localizes with RNA polymerase I and suppresses rDNA transcription.

Section snippets

Tandem affinity purification (TAP) and mass spectrometry

PARIS interacting proteins were identified by pull-down assay using the Interplay mammalian TAP system in SH-SY5Y cells (Stratagene, Santa Clara, CA). Briefly, pNTAP plasmid produces N-terminus streptavidin and calmodulin peptides and SH-SY5Y cells were transfected with pNTAP alone (as a control) or pNTAP-PARIS. Two days after transfection, cells were collected at 500 ×g at 4 °C and lysed in the supplied buffer. The TAP procedure was then performed according to the manufacturer's instructions.

PARIS interacts with MYBBP1A

To investigate the physiological roles of PARIS, we utilized tandem affinity purification (TAP) to identify PARIS interacting partners (Supplementary Table 1 and Fig. 1A, left panel). We exclude the TAP-PARIS interacting proteins that are commonly identified in TAP-control and CrapDB-TAP contaminant (Contaminant Repository for Affinity Purification, www.crapome.org) to eliminate false interactors.

The mass spectrometry analysis identified 318 proteins, including ribosome biogenesis and protein

Discussion

Several cellular stressors such as DNA damage, hypoxia, and nutrient deprivation inhibit rRNA synthesis, leading to nucleolar disruption and release of nucleolar proteins into the nucleoplasm. Mislocalization of nucleolar proteins in the nucleoplasm interferes with p53 degradation by MDM2, resulting in the accumulation of p53 and apoptosis (Deisenroth and Zhang, 2010). Accordingly, decreased rRNA synthesis has been observed in neurodegenerative diseases such as Alzheimer's, Huntington's, and PD

Acknowledgments

This research was supported by grants from the NRF (NRF-2012R1A1A1012435) funded by the Korea Ministry of Science, ICT & Future Planning (MSIP) and was also supported by a Samsung Biomedical Research Institute grant (SBRI, SMX1132521). We appreciate for providing valuable human tissue (Department of Pathology, Johns Hopkins University School of Medicine). We would like to thank Dr. Yoon Ki Kim and Sungjin Park (School of Life Science and Biotechnology, Korea University) for performing polysome

References (24)

  • R. Cong et al.

    Interaction of nucleolin with ribosomal RNA genes and its role in RNA polymerase I transcription

    Nucleic Acids Res.

    (2012)
  • C. Deisenroth et al.

    Ribosome biogenesis surveillance: probing the ribosomal protein–Mdm2–p53 pathway

    Oncogene

    (2010)
  • Cited by (0)

    View full text