Abstract
In eukaryotes, scores of excess ribosomal RNA (rRNA) genes are silenced by repressive chromatin modifications. Given the near sequence identity of rRNA genes within a species, it is unclear how specific rRNA genes are reproducibly chosen for silencing. Using Arabidopsis thaliana ecotype (strain) Col-0, a systematic search identified sequence polymorphisms that differ between active and developmentally silenced rRNA gene subtypes. Recombinant inbred mapping populations derived from three different ecotype crosses were then used to map the chromosomal locations of silenced and active RNA gene subtypes. Importantly, silenced and active rRNA gene subtypes are not intermingled. All silenced rRNA gene subtypes mapped to the nucleolus organizer region (NOR) on chromosome 2 (NOR2). All active rRNA gene subtypes mapped to NOR4. Using an engineered A. thaliana line in which a portion of Col-0 chromosome 4 was replaced by sequences of another ecotype, we show that a major rRNA gene subtype silenced at NOR2 is active when introgressed into the genome at NOR4. Collectively, these results reveal that selective rRNA gene silencing is not regulated gene by gene based on mechanisms dependent on subtle gene sequence variation. Instead, we propose that a subchromosomal silencing mechanism operates on a multimegabase scale to inactivate NOR2.
Highlights
Eukaryotes have hundreds, sometimes thousands, of nearly identical copies of ribosomal RNA genes that are transcribed in the nucleolus by RNA polymerase I (Pol I) (Long and Dawid 1980; Gerbi 1985; Hannan et al 2013; Viktorovskaya and Schneider 2015)
In the ecotype Col-0, whose genome provides the primary reference sequence for A. thaliana as a species, and in ecotype Ler, pulsed-field gel electrophoresis studies indicated that NOR on chromosome 2 (NOR2) and NOR4 consist of ∼350–400 ribosomal RNA (rRNA) genes each
VAR1 genes fail to be silenced if seeds are germinated on medium containing the cytosine methylation inhibitor 5-aza-2′-deoxycytosine or if plants are defective for repressive chromatin-modifying activities, such as HISTONE DEACETYLASE 6 (HDA6)
Summary
In A. thaliana, NORs are the most distal genetic loci on the northern ends of chromosomes two and four (NOR2 and NOR4, respectively), where the terminal rRNA genes are capped directly by telomere repeats (Fig. 1A; Copenhaver et al 1995; Copenhaver and Pikaard 1996a). 1A,B; Pontvianne et al 2010) These four gene types are revealed by PCR amplification of genomic DNA using a primer pair that flanks a 3′ ETS variable region (Fig. 1B; primer sequences are provided in Supplemental Fig. S1). All four classes of rRNA genes are expressed in newly germinated seeds, but, by 10–14 d after germination and throughout the remainder of vegetative development, the VAR1 class, accounting for ∼50% of the total rRNA gene pool, is selectively silenced (Earley et al 2006, 2010; Pontvianne et al 2010). We searched for rRNA gene sequence variation within pre-rRNA transcription units by analyzing short read genome sequence data for potential single-nucleotide polymorphisms (SNPs) relative to a consensus 45S rRNA gene reference sequence (Supplemental Fig. S2). Other sequence polymorphisms allowed for derived CAPS (dCAPS) assays (Neff et al 1998) in which the primer sequence, in conjunction with the polymorphism, contributes to the creation or abolition of a restriction endonuclease
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
