Mosaic Arrangement of the 5S rDNA in the Aquatic Plant Landoltia punctata (Lemnaceae)

Abstract

Duckweeds are a group of monocotyledonous aquatic plants in the Araceae superfamily, represented by 37 species divided into five genera. Duckweeds are the fastest growing flowering plants and are distributed around the globe; moreover, these plants have multiple applications, including biomass production, wastewater remediation, and making pharmaceutical proteins. Dotted duckweed (Landoltia punctata), the sole species in genus Landoltia, is one of the most resilient duckweed species. The ribosomal DNA (rDNA) encodes the RNA components of ribosomes and represents a significant part of plant genomes but has not been comprehensively studied in duckweeds. Here, we characterized the 5S rDNA genes in L. punctata by cloning and sequencing 25 PCR fragments containing the 5S rDNA repeats. No length variation was detected in the 5S rDNA gene sequence, whereas the nontranscribed spacer (NTS) varied from 151 to 524 bp. The NTS variants were grouped into two major classes, which differed both in nucleotide sequence and the type and arrangement of the spacer subrepeats. The dominant class I NTS, with a characteristic 12-bp TC-rich sequence present in 3–18 copies, was classified into four subclasses, whereas the minor class II NTS, with shorter, 9-bp nucleotide repeats, was represented by two identical sequences. In addition to these diverse subrepeats, class I and class II NTSs differed in their representation of cis-elements and the patterns of predicted G-quadruplex structures, which may influence the transcription of the 5S rDNA. Similar to related duckweed species in the genus Spirodela, L. punctata has a relatively low rDNA copy number, but in contrast to Spirodela and the majority of other plants, the arrangement of the 5S rDNA units demonstrated an unusual, heterogeneous pattern in L. punctata, as revealed by analyzing clones containing double 5S rDNA neighboring units. Our findings may further stimulate the research on the evolution of the plant rDNA and discussion of the molecular forces driving homogenization of rDNA repeats in concerted evolution.

The Ribosomal DNA Loci of the Ancient Monocot Pistia stratiotes L. (Araceae) Contain Different Variants of the 35S and 5S Ribosomal RNA Gene Units

“…LC213012 ) and the eudicot Gossypium ( Cronn et al, 1996 ). The minor group (Type II) is represented by two of the 64 NTSs (clones Ps19 and Ps20), which start with the classic 5S rDNA terminator motif TTTTT found in most of the plants analyzed, including the duckweeds Spirodela polyrhiza ( Borisjuk et al, 2018 ) and Landoltia punctata ( Chen et al, 2021 ). Types I and II also have a slightly different nucleotide arrangement at their 3′-termini in front of the 5S rRNA gene ( Figure 5A ).…”

“…To the best of our knowledge, this work represents the first molecular study of 5S rDNA in Araceae considering the related duckweeds, for which data are available ( Borisjuk et al, 2018 ; Hoang et al, 2020 ; Chen et al, 2021 ), as a separate family. The sequencing of multiple 5S rDNA gene units, containing 78 sequences of full length 5S rRNA gene and 64 full-length intergenic spacers of P. stratiotes , revealed a molecular arrangement common for the majority of analyzed plants ( Zhu et al, 2008 ), with a conserved 119-bp sequence for 5S rRNA and a variable non-transcribed spacer (NTS).…”

“…For analysis of P. stratiotes 5S rRNA genes, the specific DNA fragments were amplified from genomic DNA by PCR using two pairs of primers specific for 5S rRNA gene sequence DW-5S-F/DW-5S-R and cn-5S-for/cn-5S-rev ( Supplementary Table 1 ) as previously described ( Chen et al, 2021 ). In order to increase the chance of amplifying DNA fragments with multiple 5S rDNA units, the elongation time of PCR was prolonged to 1 min 30 s. The generated DNA fragments, cloned into the vector pMD19 (Takara, Dalian, China) were custom sequenced (Sangon Biotech, Shanghai, China), and the obtained nucleotide sequences were analyzed using the CLC Main Workbench (Version 6.9.2, Qiagen) software.…”

Personal Perspectives on Plant Ribosomal RNA Genes Research: From Precursor-rRNA to Molecular Evolution

“…The duckweed species Spirodela polyrhiza and S. intermedia might emerge as a promising new model to study rDNA regulation (in addition to Arabidopsis) because of their compact rDNA loci, composed of no more than a hundred 35S repeated units (Michael et al, 2017;Hoang et al, 2020), even fewer than the 100-200 rDNA copies in yeast (Salim et al, 2017), and an order of magnitude lower than rRNA gene copies in other plants (Wang et al, 2019). The copy number of 5S rDNA is estimated to be about 170 in Landoltia punctuata (Chen et al, 2021) -this issue. This exceptionally low copy number makes the duckweed rDNA locus relatively simple to access applying the third-generation sequencing platforms for ultralong sequencing reads (Jung et al, 2019).…”

Genomic architecture of 5S rDNA cluster and its variations within and between species

“…1 c). A recent study investigated the structure of 5S rDNA units in the Aquatic Plant Landoltia punctata (Lemnaceae) by PCR amplification of 5S rDNA followed by Sanger DNA sequencing [ 43 ]. No length variation was detected in the 5S rDNA gene sequence, whereas the nontranscribed spacer was found to vary from 151 to 524 bp.…”