Proteasome inhibition alters mitotic progression through the upregulation of centromeric α‐Satellite RNAs

Abstract

Cell cycle progression requires control of the abundance of several proteins and RNAs over space and time to properly transit from one phase to the next and to ensure faithful genomic inheritance in daughter cells. The proteasome, the main protein degradation system of the cell, facilitates the establishment of a proteome specific to each phase of the cell cycle. Its activity also strongly influences transcription. Here, we detected the upregulation of repetitive RNAs upon proteasome inhibition in human cancer cells using RNA‐seq. The effect of proteasome inhibition on centromeres was remarkable, especially on α‐Satellite RNAs. We showed that α‐Satellite RNAs fluctuate along the cell cycle and interact with members of the cohesin ring, suggesting that these transcripts may take part in the regulation of mitotic progression. Next, we forced exogenous overexpression and used gapmer oligonucleotide targeting to demonstrate that α‐Sat RNAs have regulatory roles in mitosis. Finally, we explored the transcriptional regulation of α‐Satellite DNA. Through in silico analyses, we detected the presence of CCAAT transcription factor‐binding motifs within α‐Satellite centromeric arrays. Using high‐resolution three‐dimensional immuno‐FISH and ChIP‐qPCR, we showed an association between the α‐Satellite upregulation and the recruitment of the transcription factor NFY‐A to the centromere upon MG132‐induced proteasome inhibition. Together, our results show that the proteasome controls α‐Satellite RNAs associated with the regulation of mitosis.

Satellite DNAs in Health and Disease

“…Satellite DNA repeats located in (peri)centromeric heterochromatin are actively transcribed with transcripts being necessary for the formation and maintenance of heterochromatin as well as centromere function [ 12 , 13 , 14 ]. The presence of transcription factor binding sites was predicted and experimentally proven for different satellite DNAs [ 15 , 16 ] and transcription, which is often bidirectional, proceeds from internal promoters, usually by RNA polymerase II (RNA Pol II; [ 17 , 18 , 19 , 20 , 21 ]). Under physiological conditions, satellite DNA expression is low, and it is temporally and spatially regulated [ 18 , 22 , 23 ].…”

“…Namely, CENP-B promotes the binding of the zinc-finger transcriptional regulator (ZFAT) which activates RNA Pol II transcription through histone modification H4K8Ac. α satellite RNA level fluctuates throughout the cell cycle, peaking in the G2/M phase [ 21 ], being controlled by the proteasome [ 16 ]. Satellite RNA interacts with members of the cohesin ring, suggesting a role for these transcripts in the regulation of mitotic progression [ 16 ].…”

“…α satellite RNA level fluctuates throughout the cell cycle, peaking in the G2/M phase [ 21 ], being controlled by the proteasome [ 16 ]. Satellite RNA interacts with members of the cohesin ring, suggesting a role for these transcripts in the regulation of mitotic progression [ 16 ]. The abundance of centromeric satellite RNAs seems to be regulated during the cell cycle in other organisms such as mouse, peaking in G2 phase, and long transcripts undergo post-transcriptional processing to generate smaller RNAs from 120 to 150 nucleotides [ 47 ].…”

LncRNAs-associated to genomic instability: A barrier to cancer therapy effectiveness

“…Although the centromere and the pericentromere bear epigenetic repressive marks, they can also display histone post-translational modifications associated with active chromatin. Basal centromeric expression is detectable in normal human cells from different tissues ( Eymery et al, 2009 ), even by Northern blot ( Caceres-Gutierrez et al, 2022 ). But these regions are also transcriptionally dynamic and highly responsive to the cellular context and internal and external stimuli, such as cell cycle progression ( Bury et al, 2020 ), differentiation ( Bouzinba-Segard et al, 2006 ), cancer progression ( Ting et al, 2011 ; Zhu et al, 2011 ), heat shock, osmotic pressure, oxidative stress, and exposure to heavy metals ( Valgardsdottir et al, 2008 ).…”

“…In this regard, we have demonstrated that proteasome inhibitors promote the overexpression of several repetitive RNAs, including the centromeric α-satellites ( Caceres-Gutierrez et al, 2022 ). Centromeric and pericentromeric lncRNAs have also been associated with resistance to different antineoplastic drugs, such as the proteasome inhibitor bortezomib ( Figure 2 ).…”