Nucleolus

About: Nucleolus is a research topic. Over the lifetime, 5873 publications have been published within this topic receiving 232435 citations. The topic is also known as: GO:0005730 & cell nucleolus.

The nucleolar ubiquitin-specific protease USP36 deubiquitinates and stabilizes c-Myc

Abstract: c-Myc protein stability and activity are tightly regulated by the ubiquitin-proteasome system. Aberrant stabilization of c-Myc contributes to many human cancers. c-Myc is ubiquitinated by SCFFbw7 (a SKP1-cullin-1-F-box complex that contains the F-box and WD repeat domain-containing 7, Fbw7, as the F-box protein) and several other ubiquitin ligases, whereas it is deubiquitinated and stabilized by ubiquitin-specific protease (USP) 28. The bulk of c-Myc degradation appears to occur in the nucleolus. However, whether c-Myc is regulated by deubiquitination in the nucleolus is not known. Here, we report that the nucleolar deubiquitinating enzyme USP36 is a novel c-Myc deubiquitinase. USP36 interacts with and deubiquitinates c-Myc in cells and in vitro, leading to the stabilization of c-Myc. This USP36 regulation of c-Myc occurs in the nucleolus. Interestingly, USP36 interacts with the nucleolar Fbw7γ but not the nucleoplasmic Fbw7α. However, it abolished c-Myc degradation mediated both by Fbw7γ and by Fbw7α. Consistently, knockdown of USP36 reduces the levels of c-Myc and suppresses cell proliferation. We further show that USP36 itself is a c-Myc target gene, suggesting that USP36 and c-Myc form a positive feedback regulatory loop. High expression levels of USP36 are found in a subset of human breast and lung cancers. Altogether, these results identified USP36 as a crucial and bono fide deubiquitinating enzyme controlling c-Myc’s nucleolar degradation pathway.

Epigenetic disruption of ribosomal RNA genes and nucleolar architecture in DNA methyltransferase 1 (Dnmt1) deficient cells

Abstract: The nucleolus is the site of ribosome synthesis in the nucleus, whose integrity is essential. Epigenetic mechanisms are thought to regulate the activity of the ribosomal RNA (rRNA) gene copies, which are part of the nucleolus. Here we show that human cells lacking DNA methyltransferase 1 (Dnmt1), but not Dnmt33b, have a loss of DNA methylation and an increase in the acetylation level of lysine 16 histone H4 at the rRNA genes. Interestingly, we observed that SirT1, a NAD+-dependent histone deacetylase with a preference for lysine 16 H4, interacts with Dnmt1; and SirT1 recruitment to the rRNA genes is abrogated in Dnmt1 knockout cells. The DNA methylation and chromatin changes at ribosomal DNA observed are associated with a structurally disorganized nucleolus, which is fragmented into small nuclear masses. Prominent nucleolar proteins, such as Fibrillarin and Ki-67, and the rRNA genes are scattered throughout the nucleus in Dnmt1 deficient cells. These findings suggest a role for Dnmt1 as an epigenetic caretaker for the maintenance of nucleolar structure.

The structure of NoRC-associated RNA is crucial for targeting the chromatin remodelling complex NoRC to the nucleolus.

Abstract: Silencing of ribosomal RNA genes (rDNA) requires binding of the chromatin remodelling complex NoRC to RNA that is complementary to the rDNA promoter. NoRC-associated RNA (pRNA) folds into a conserved stem–loop structure that is required for nucleolar localization and rDNA silencing. Mutations that disrupt the stem–loop structure impair binding of TIP5, the large subunit of NoRC, to pRNA and abolish targeting of NoRC to nucleoli. Binding to pRNA results in a conformational change of TIP5, as shown by enhanced sensitivity of TIP5 towards trypsin digestion. Our results indicate an RNA-dependent mechanism that targets NoRC to chromatin and facilitates the interaction with co-repressors that promote heterochromatin formation and rDNA silencing.

In vivo evidence that TATA-binding protein/SL1 colocalizes with UBF and RNA polymerase I when rRNA synthesis is either active or inactive.

Abstract: Here we show that the TATA-binding protein (TBP) is localized in the nucleoplasm and in the nucleolus of mammalian cells, consistent with its known involvement in transcription by RNA polymerase I, II, and III. In the nucleolus of actively growing cells, TBP colocalizes with upstream binding factor (UBF) and RNA polymerase I at the sites of rRNA transcription. During mitosis, when rRNA synthesis is down-regulated, TBP colocalizes with TBP-associated factors for RNA polymerase I (TAF(I)s), UBF, and RNA polymerase I on the chromosomal regions containing the rRNA genes. Treatment of cells with a low concentration of actinomycin D inhibits rRNA synthesis and causes a redistribution of the rRNA genes that become concentrated in clusters at the periphery of the nucleolus. A similar redistribution was observed for the major components of the rRNA transcription machinery (i.e., TBP, TAF(I)s, UBF, and RNA polymerase I), which still colocalized with each other. Furthermore, anti-TBP antibodies are shown to coimmunoprecipitate TBP and TAF(I)63 in extracts prepared from untreated and actinomycin D-treated cells. Collectively, the data indicate that in vivo TBP/promoter selectivity factor, UBF, and RNA polymerase I remain associated with both active and inactive rRNA genes.