Reaching for the off switch in nucleolar dominance

Abstract

Nucleolus organizer regions (NORs) are eukaryotic chromosomal loci where ribosomal RNA (rRNA) genes are clustered, typically in hundreds to thousands of copies. Transcription of these rRNA genes by RNA polymerase I and processing of their transcripts results in the formation of the nucleolus, the sub-nuclear domain in which ribosomes are assembled. Approximately 90 years ago, cytogenetic observations revealed that NORs inherited from the different parents of an interspecific hybrid sometimes differ in morphology at metaphase. Fifty years ago, those chromosomal differences were found to correlate with differences in rRNA gene transcription and the phenomenon became known as nucleolar dominance. Studies of the past 30 years have revealed that nucleolar dominance results from selective rRNA gene silencing, involving repressive chromatin modifications, and occurs in pure species as well as hybrids. Recent evidence also indicates that silencing depends on the NOR in which an rRNA gene is located, and not on the gene's sequence. In this perspective, we discuss how our thinking about nucleolar dominance has shifted over time from the kilobase scale of individual genes to the megabase scale of NORs and chromosomes and questions that remain unanswered in the search for a genetic and biochemical understanding of the off switch.

Keywords: Arabidopsis suecica; Arabidopsis thaliana; Brassica allopolyploids; DNA methylation; chromatin; histone modification; nucleolus; nucleolus organizer region; rRNA gene; ribosomal RNA; transcriptional silencing.

Figure 1.. Nucleoli, NORs and rRNA gene expression.

a. Nucleoli form within nuclei as a consequence of rRNA gene transcription and processing. Thus, active rRNA genes (green triangles) are located within the nucleolus whereas inactive rRNA genes are located outside the nucleolus, as confirmed by flow cytometry experiments (Pontvianne et al., 2013). The cartoon depicts a diploid nucleus, at interphase, that has nucleoli on two different chromosomes (as in Arabidopsis thaliana), with the NORs identified by DNA-FISH (fluorescence in situ hybridization) using an rRNA gene probe. Inactive genes co-localize within highly condensed foci, and this includes the excess genes of active NORs. b. At metaphase, secondary constrictions are observed at NOR intervals where rRNA genes were transcriptionally active during the prior interphase. c. Chromatin modifications that correlate with the on and off states of plant rRNA genes, and names of some of the Arabidopsis chromatin modifying enzymes (in parentheses) whose mutation or knockdown disrupts rRNA gene silencing (Lawrence and Pikaard, 2004, Probst et al., 2004, Earley et al., 2006, Preuss et al., 2008, Pontvianne et al., 2012, Mohannath et al., 2023). Note that DNA methyltransferase MET1 primarily catalyzes maintenance methylation in the CG context, but also affects CHG maintenance methylation to some extent. CMT2 primarily catalyzes maintenance methylation in the CHH context. DRM2 catalyzes de novo methylation in all contexts (CG, CHG and CHH). d. rRNA genes that map to NOR2 are selectively silenced whereas genes that map to NOR4 are expressed in the A. thaliana accession, Col-0. The molecular signals and targets that initiate silencing remain undefined but may involve sequences internal to, or flanking, NOR2 and possibly NOR4.