Investigating the nucleolar epigenetic code at ultrastructural level

• Published in: Biopolimery i kletka 2019-05, Vol.35 (3), p.243-244
• Main Authors: Zannino, L., Bertone, V., Siciliani, S., Saia, L., Biggiogera, M.
• Format: Article
• Language: English
• Subjects: Apoptosis; Bioinformatics; Cell proliferation; Chromatin; Electron microscopy; Epigenetics; Euchromatin; Gene silencing; Heterochromatin; Immunocytochemistry; Nucleoli; rRNA; Statistical analysis; Transcription; Transfer RNA
• ISSN: 0233-7657; 1993-6842
• DOI: 10.7124/bc.000A05

The nucleolus is a nuclear body where different important molecular processes occurs. Beyond ribosomal biogenesis, other relevant functions were recently assigned to this nuclear region, which are related to cell proliferation control and apoptosis, involvement in telomere formation, transfer RNA modifications and stress sensing. Morphologically it is organized in three main areas: roundish electron-light regions, known as Fibrillar Centers (FCs), surrounded by the Dense Fibrillar Component (DFC). These fibrillary structures lie inside the Granular Component (GC), which constitutes most of the nucleolus. Moreover patches of heterochromatin delimitate the nucleolar periphery, interspaced by euchromatin, with thin strands of condensed chromatin enter the nucleolar body. Some aspects of nucleolar morphology have been correlated to their corresponding molecular activity. It is established that rDNA is present within the FC, DFC, in the perinucleolar heterochromatin and in its intranucleolar strands, whereas ribosomal RNA was localized to DFC and GC. rRNA transcription occurs in the FC/DFC complex, while outside the nucleolus reside transcriptionally inactive rDNA repeats. However we still have little knowledge about the condensed regions of perinucleolar heterochromatin. In order to characterize the molecular activity of this area, we decided to investigate its epigenetics status. We hypothesized that, being a condensed region, it would show the classical markers of repression find in the other nuclear regions characterized by compact chromatin. Indeed we analysed at ultrastructural level the distribution of the histone markers H3K27me3 and H3K9me3, which are known to be involved in chromatin condensation and gene silencing. This study was carried out by immunocytochemistry of these histone marker distributions at electron microscopy. Moreover quantifications and statistics of the marker distributions using bioinformatics tools were carried out. We were able to highlight that not only in all compact regions of the nuclear and nucleolar heterochromatin these two repressive histone markers were present, but also that they were specifically confined to the heterochromatin. From our analysis no significant difference in their density or distributions were found between the nucleolar associated and nuclear heterochromatin. Considering these results, we hypothesize that the general mechanisms of chromatin condensation which involve H3K27me3 and H3K9me3 could be simi