Human rDNA and Cancer

In human cells, each rDNA unit consists of the ~13 kb long ribosomal part and ~30 kb long intergenic spacer (IGS). The ribosomal part, transcribed by RNA polymerase I (pol I), includes genes coding for 18S, 5.8S, and 28S RNAs of the ribosomal particles, as well as their four transcribed spacers. Being highly repetitive, intensively transcribed, and abundantly methylated, rDNA is a very fragile site of the genome, with high risk of instability leading to cancer. Multiple small mutations, considerable expansion or contraction of the rDNA locus, and abnormally enhanced pol I transcription are usual symptoms of transformation. Recently it was found that both IGS and the ribosomal part of the locus contain many functional/potentially functional regions producing non-coding RNAs, which participate in the pol I activity regulation, stress reactions, and development of the malignant phenotype. Thus, there are solid reasons to believe that rDNA locus plays crucial role in carcinogenesis. In this review we discuss the data concerning the human rDNA and its closely associated factors as both targets and drivers of the pathways essential for carcinogenesis. We also examine whether variability in the structure of the locus may be blamed for the malignant transformation. Additionally, we consider the prospects of therapy focused on the activity of rDNA.

Comparative Analysis of the nrDNA Repeat Unit of Manila Clam Ruditapes philippinarum and Quahog Mercenaria mercenaria

Ruditapes philippinarum and Mercenaria mercenaria are economically important bivalve species. The complete ribosomal DNA (rDNA) unit sequences of R. philippinarum and M. mercenaria, with as-sembled rDNA unit lengths of 12,910 and 12,100 bp, respectively, were obtained in this study for the first time. The rDNA unit structural organisation was similar to that in other eukaryotes, in-cluding the following elements in order: 18S rRNA-internal transcribed spacer 1 (ITS1); 5.8S rRNA-ITS2-28S rRNA-intergenic spacer (IGS) (3′ external transcribed spacer (ETS); non-transcribed spacer (NTS)-5′ ETS). The genetic differences between R. philippinarum and M. mercenaria were mainly attributable to non-coding regions (ITS1, ITS2 and IGS), especially the IGS region. The boundaries of putative 3′ ETS, NTS and 5′ ETS were confirmed. Seven and three sub-repeat fragments were found in R. philippinarum and M. mercenaria, respectively. These frag-ments ranged from 4 to 154 bp in length, and were located at the NTS and 5′ ETS regions. Five and six cytosine–guanine (CpG) islands were detected in R. philippinarum and M. mercenaria, respec-tively, and these covered 85.58% and 79.29% of the entire IGS sequence, respectively. The phylo-genetic tree was constructed based on Veneridae ITS and 18S rRNA sequences using the maxi-mum likelihood (ML) method. The ML tree based on ITS revealed that species within the same genus clearly clustered together with relatively high supporting values, and all the genera were recovered as monophyletic. The phylogenetic analyses using 18S rRNA provided a weaker phy-logenetic signal than ITS.

Multiple horizontal transfers of nuclear ribosomal genes between phylogenetically distinct grass lineages

The movement of nuclear DNA from one vascular plant species to another in the absence of fertilization is thought to be rare. Here, nonnative rRNA gene [ribosomal DNA (rDNA)] copies were identified in a set of 16 diploid barley (Hordeum) species; their origin was traceable via their internal transcribed spacer (ITS) sequence to five distinct Panicoideae genera, a lineage that split from the Pooideae about 60 Mya. Phylogenetic, cytogenetic, and genomic analyses implied that the nonnative sequences were acquired between 1 and 5 Mya after a series of multiple events, with the result that some current Hordeum sp. individuals harbor up to five different panicoid rDNA units in addition to the native Hordeum rDNA copies. There was no evidence that any of the nonnative rDNA units were transcribed; some showed indications of having been silenced via pseudogenization. A single copy of a Panicum sp. rDNA unit present in H. bogdanii had been interrupted by a native transposable element and was surrounded by about 70 kbp of mostly noncoding sequence of panicoid origin. The data suggest that horizontal gene transfer between vascular plants is not a rare event, that it is not necessarily restricted to one or a few genes only, and that it can be selectively neutral.

Genetic and epigenetic changes of rDNA in a synthetic allotetraploid, Aegilops sharonensis × Ae. umbellulata

The synthetic allotetraploid Aegilops sharonensis × Ae. umbellulata (genomic formula SshU) was used to study inheritance and expression of 45S rDNA during early stages of allopolyploid formation. Using silver staining, we revealed suppression of the NORs (nucleolar organizing regions) from the Ssh genome in response to polyploidization. Most allopolyploid plants of the S2–S4 generations retained the chromosomal location of 45S rDNA typical for the parental species, except for two S3 plants in which a deletion of the rDNA locus on one of the homologous 6Ssh chromosomes was revealed. In addition, we found a decrease in NOR signal intensity on both 6Ssh chromosomes in a portion of the S3 and S4 allopolyploid plants. As Southern hybridization showed, the allopolyploid plants demonstrated additive inheritance of parental rDNA units together with contraction of copy number of some rDNA families inherited from Ae. sharonensis. Also, we identified a new variant of amplified rDNA unit with MspAI1 restriction sites characteristic of Ae. umbellulata. These genetic alterations in the allopolyploid were associated with comparative hypomethylation of the promoter region within the Ae. umbellulata–derived rDNA units. The fast uniparental elimination of rDNA observed in the synthetic allopolyploid agrees well with patterns observed previously in natural wheat allotetraploids.

Molecular diversity of the 5S rDNA units in the Elymus dahuricus complex (Poaceae: Triticeae) supports the genomic constitution of St, Y, and H haplomes

The phylogenetic analysis of 118 5S rRNA gene sequences cloned from members of the Elymus dahuricus complex containing the St, Y, and H haplomes, and of several related species containing at least one of these three haplomes, is reported. Differences in sequence pattern, primarily within the nontranscribed spacer, enabled the identification of six putative orthologous groups that we refer to as unit classes. In previous publications, we have been able to assign unit classes to haplomes. In addition to four unit classes previously identified in other genera, namely the long H1, long S1, long P1, and long {Y1, here we document two new unit classes called the long S2 and long W1. Most sequences of the E. dahuricus complex and related tetraploid species are classified as long S1 and assigned to the St haplome. Both long S1 and long S2 unit classes were identified in the diploid Pseudoroegneria spicata (Pursh) Á. Löve with the St haplome. The long S2 unit class was also identified in the hexaploid Elymus scabrus (R. Br.) Á. Löve with the St,Y,and W haplomes. The long P1 was known from the diploid Agropyron cristatum Gaertn. with the P haplome, and the long W1 was determined in Australopyrum retrofractum (Vickery) Á. Löve, known to contain the W haplome, but was not yet detected in E. scabrus, a hexaploid species with W being one of the three haplomes. The long H1 reported earlier from Hordeum was identified in several clones of the E. dahuricus complex. As previously reported, the long {Y1 unit class was found to be rare overall, but we identified it in a few clones of Elymus drobovii and in the E. dahuricus complex.Key words: 5S rDNA, unit classes, haplomes, concerted evolution.

Evolution of 5S rDNA unit arrays in the plant genus Nicotiana (Solanaceae)

Nicotiana tabacum (tobacco, Solanaceae) has two 5S ribosomal DNA (rDNA) families, one of unit length ~646 bp and the other ~430 bp, that differ in the length of the 5S rDNA non-transcribed spacer (NTS). The long 5S rDNA family, found on the T genome of tobacco and in Nicotiana tomentosiformis, contains a GC-rich subregion that is absent in the short family. We designed primers for this subregion and generated a probe that we used against a range of Nicotiana and related Solanaceous species. We demonstrated the presence of the GC-rich subregion in a range of Nicotiana species, but it was absent in Nicotiana sylvestris, Nicotiana longiflora, and two closely related genera, Petunia and Solanum. These data suggest that this subregion of the NTS is likely to have evolved with the genus Nicotiana. The absence of the subregion in N. sylvestris and N. longiflora is likely to have arisen by a deletion event in the evolution of section alatae. We demonstrate patterns of evolution in the 5S rDNA unit cluster in relation to a phylogenetic reconstruction of species relationships in section tomentosae. Nicotiana glutinosa diverged early from the section and contains a 5S rDNA family based on a 550-bp unit. After this divergence, 430- and 650-bp rDNA unit families evolved. The 650-bp family is found in all species of tomentosae (except N. glutinosa) and in tobacco. The 430-bp family within tomentosae includes the GC-rich subregion and is thus unrelated to the 430-bp family in N. sylvestris. Nicotiana setchellii is unusual in that it has three 5S rDNA loci, including one locus that is exceptionally large. This species, unique to tomentosae, has a very abundant 900-bp unit family. It is possible that this 900-bp family occurs on this one large locus. In N. tomentosa and N. kawakamii, the 650-bp family is predominant, whereas N. tomentosiformis and N. otophora have only the 650-bp family. There is no clear relationship between the number of 5S families and the number of 5S rDNA loci. Certainly the replacement of 5S rDNA units, perhaps by gene conversion, has occurred repeatedly in the evolution of genus Nicotiana.Key words: 5S rDNA, evolution, Nicotiana, Solanaceae, satellite homogenization.

Patterns of Variation in the Intergenic Spacers of Ribosomal DNA in Drosophila melanogaster Support a Model for Genetic Exchanges During X-Y Pairing

Detailed analysis of variation in intergenic spacer (IGS) and internal transcribed spacer (ITS) regions of rDNA drawn from natural populations of Drosophila melanogaster has revealed contrasting patterns of homogenization although both spacers are located in the same rDNA unit. On the basis of the role of IGS regions in X-Y chromosome pairing, we proposed a mechanism of single-strand exchanges at the IGS regions, which can explain the different evolutionary trajectories followed by the IGS and the ITS regions. Here, we provide data from the chromosomal distribution of selected IGS length variants, as well as the detailed internal structure of a large number of IGS regions obtained from specific X and Y chromosomes. The variability found in the different internal subrepeat regions of IGS regions isolated from X and Y chromosomes supports the proposed mechanism of genetic exchanges and suggests that only the “240” subrepeats are involved. The presence of a putative site for topoisomerase I at the 5′ end of the 18S rRNA gene would allow for the exchange between X and Y chromosomes of some 240 subrepeats, the promoter, and the ETS region, leaving the rest of the rDNA unit to evolve along separate chromosomal lineages. The phenomenon of localized units (modules) of homogenization has implications for multigene family evolution in general.