Single ribosomal transcription units are linear, compacted Christmas trees in plant nucleoli

Abstract

SummaryThe rDNA transcription units are enormous macromolecular structures located in the nucleolus and containing 50±100 RNA polymerases together with the nascent pre-rRNA attached to the rDNA. It has not previously been possible to visualize nucleolar transcription units directly in intact nucleoli, although highly spread preparations in the electron microscope have been imaged as`Christmas trees' 2±3 mm long. Here we determine the relative conformation of individual transcription units in Pisum sativum plant nucleoli using a novel labelling technique. Nascent transcripts were detected by a highly sensitive silver-enhanced 1 nm gold procedure, followed by 3D electron microscopy of entire nucleoli. Individual transcription units are seen as conical, elongated clusters approximately 300 nm in length and 130 nm in width at the thickest end. We further show that there were approximately 300 active ribosomal genes in the nucleoli examined. The underlying chromatin structure of the transcribing rDNA was directly visualized by applying a novel limited extraction procedure to ®xed specimens in order to wash out the proteins and RNA, thus speci®cally revealing DNA strands after uranyl acetate staining. Using this technique, followed by post-embedding in situ hybridization, we observed that the nucleolar rDNA ®bres are not extended but show a coiled, thread-like appearance. Our results show for the ®rst time that native rDNA transcription units are linear, compacted Christmas trees.

Characterization of AtNUC-L1 Reveals a Central Role of Nucleolin in Nucleolus Organization and Silencing of AtNUC-L2 Gene in Arabidopsis

“…AtNUC-L1 is localized in the nucleolus in the DFC surrounding FC at the site of rRNA transcription (which occurs in the border zone between FC and DFC; Gonzalez-Melendi et al, 2001, and references therein); we also demonstrate that AtNUC-L1 disruption has a major effect on nucleolus structure. We have observed that the nucleolus is completely disorganized in the Atnuc-L1 plant, reminiscent of the ultrastructural changes observed in the yeast nucleolus of gar2 (Leger-Silvestre et al, 1997).…”

“…Occasionally, depending on cell type and stage, other structures such as vacuoles can also be observed in plant nucleolus (Gonzalez-Melendi et al, 2001). It has been accepted for some time that ribosome biosynthesis occurs in a vectorial pattern within the nucleolus.…”

Concerted Evolution of rDNA in Recently Formed Tragopogon Allotetraploids Is Typically Associated With an Inverse Correlation Between Gene Copy Number and Expression

“…In A. thaliana, which has $800 rRNA genes per diploid cell, $50% of the units are estimated to be active. Electron microscopy observations indicate that there are $300 units active in pea nucleoli (Gonzalez-Melendi et al 2001). By extension, nearly all units of T. dubius origin might be active in the 2603-33 individual.…”

Chromosome banding and essential oils composition of Brazilian accessions of Lippia alba (Verbenaceae)

“…In Pisum sativum, only about 5% of the units are transcribed (González-Melendi et al 2001). Thus, a small fraction of rRNA genes from only one locus seems to be sufficient to supply all the ribosomal machinery of a cell.…”