Centromeric and pericentric transcription and transcripts: their intricate relationships, regulation, and functions

Abstract

Centromeres are no longer considered to be silent. Both centromeric and pericentric transcription have been discovered, and their RNA transcripts have been characterized and probed for functions in numerous monocentric model organisms recently. Here, we will discuss the challenges in centromere transcription studies due to the repetitive nature and sequence similarity in centromeric and pericentric regions. Various technological breakthroughs have helped to tackle these challenges and reveal unique features of the centromeres and pericentromeres. We will briefly introduce these techniques, including third-generation long-read DNA and RNA sequencing, protein-DNA and RNA-DNA interaction detection methods, and epigenomic and nucleosomal mapping techniques. Interestingly, some newly analyzed repeat-based holocentromeres also resemble the architecture and the transcription behavior of monocentromeres. We will summarize evidences that support the functions of the transcription process and stalling, and those that support the functions of the centromeric and pericentric RNAs. The processing of centromeric and pericentric RNAs into multiple variants and their diverse structures may also provide clues to their functions. How future studies may address the separation of functions of specific centromeric transcription steps, processing pathways, and the transcripts themselves will also be discussed.

Keywords: Centromere; Centromeric and pericentric RNAs; Centromeric and pericentric transcription; Chromatin environment; Epigenetics; Non-coding RNA.

Fig. 1

A schematic diagram outlining how centromeric transcription and stalling can regulate centromere CENP-A loading. A The chromatin environment changes and the “transcription bubble” generated by RNA Pol II passage favors the incorporation of CENP-A nucleosomes at centromere regions. Transcription-associated chromatin remodeling factors, including FACT complex and CHD1, and CENP-A chaperone such as CAL1 or HJURP together with MIS18 have been shown to be important for CENP-A deposition during the centromeric transcription. RNA Pol II phosphorylated at Ser2 of the C-terminal domain (CTD) is present at centromeres, indicating active transcription elongation through the action of RNA Pol II. The nucleosome destabilization activity of FACT complex, which consists of SSRP1 and SUPT16H subunits, could promote RNA Pol II elongation through the compact chromatin, while RNA Pol II transcription could drive further loosening at the centromere domain. FACT has been shown to interact with the CENP-A protein by interacting with CAL1, the CENP-A loading factor in Drosophila. FACT destabilizes H3 nucleosomes in order to promote CENP-A loading. RNA Pol II transcription could also recruit HAT complexes to the kinetochore to generate an acetylated chromatin environment, which has been shown to be favorable for CENP-A loading. Unknown factors involved in transcription elongation may also facilitate CENP-A deposition. B RNA Pol II stalling allows de novo establishment of CENP-A chromatin. Serine 2 in the CTD heptad repeat of RNA Pol II is phosphorylated in elongating RNA Pol II, and this RNA Pol II becomes ubiquitylated upon stalling. In S. pombe, newly introduced plasmid with the core centromere region will lead to transient stalling of RNA Pol II, but it can be efficiently cleared with the aid of factors such as TFIIS and Ubp3 (Kulish et al. ; Kvint et al. ; Martinez-Rucobo & Cramer 2013). TFIIS promotes transcriptional elongation by cleaving nascent transcripts in the context of stalled RNA Pol II. Mutants that lack Ubp3 or TFIIS compromise the restarting process of stalled RNA Pol II, resulting in the accumulation of stalled RNA Pol II complexes, prolonged stalling, and leading to recruitment of unknown factors that promote CENP-A deposition. The RNA Pol II stalling environment causes H3 nucleosomes to be efficiently evicted. CENP-A N-terminal tail lacks the conserved lysine residues of H3 (e.g., K9), and thus does not have H3K9ac-like modification that aids transcription. Therefore, CENP-A nucleosomes are likely to present a greater barrier to transcription than H3 nucleosomes. Thus, once CENP-A nucleosomes are loaded, it might exacerbate the transcriptional stalling, creating conditions permissive for recruitment of more CENP-A in a self-perpetuating way

Fig. 2

Mechanisms that contribute to cenRNA and pericenRNA variants and their potential functions. A RNA transcription could be initiated and terminated at different sites. CenRNA isoforms cover part or all of the core centromere regions, while pericenRNA isoforms are proximal to but do not cover the centromere regions. Pericentric RNAs may cover part of nearby genes, as observed in budding yeast (Hedouin et al. 2022). B CenRNAs and pericenRNAs could be processed by splicing (Neumann et al. 2007), m6A methylation (Xiao et al. 2016), 5′-capping (Choi et al. 2011), and polyadenylation (Arunkumar and Melters ; Choi et al. ; Ling & Yuen ; Neumann et al. 2007). The biological functions of these cenRNA processing processes and products are less certain, but could resemble those of mRNAs or sRNAs