Limits...
Molecular organization of the 25S-18S rDNA IGS of Fagus sylvatica and Quercus suber: a comparative analysis.

Inácio V, Rocheta M, Morais-Cecílio L - PLoS ONE (2014)

Bottom Line: In the NTS: the sub-repeats structure in Beech is more organized than in Cork Oak, sharing some short motifs which results in the lowest sequence similarity of the entire IGS; the AT-rich region differed in both spacers by a GC-rich block inserted in Cork Oak.Methylation of 25S Bam HI site was confirmed in all species and detected for the first time in the 18S of Q. suber and Q. faginea.These results provide important clues for the evolutionary trends of the rDNA 25S-18S IGS in the Fagaceae family.

View Article: PubMed Central - PubMed

Affiliation: Centre for Botany Applied to Agriculture (CBAA), Instituto Superior de Agronomia, University of Lisbon, Lisbon, Portugal.

ABSTRACT
The 35S ribosomal DNA (rDNA) units, repeated in tandem at one or more chromosomal loci, are separated by an intergenic spacer (IGS) containing functional elements involved in the regulation of transcription of downstream rRNA genes. In the present work, we have compared the IGS molecular organizations in two divergent species of Fagaceae, Fagus sylvatica and Quercus suber, aiming to comprehend the evolution of the IGS sequences within the family. Self- and cross-hybridization FISH was done on representative species of the Fagaceae. The IGS length variability and the methylation level of 18 and 25S rRNA genes were assessed in representatives of three genera of this family: Fagus, Quercus and Castanea. The intergenic spacers in Beech and Cork Oak showed similar overall organizations comprising putative functional elements needed for rRNA gene activity and containing a non-transcribed spacer (NTS), a promoter region, and a 5'-external transcribed spacer. In the NTS: the sub-repeats structure in Beech is more organized than in Cork Oak, sharing some short motifs which results in the lowest sequence similarity of the entire IGS; the AT-rich region differed in both spacers by a GC-rich block inserted in Cork Oak. The 5'-ETS is the region with the higher similarity, having nonetheless different lengths. FISH with the NTS-5'-ETS revealed fainter signals in cross-hybridization in agreement with the divergence between genera. The diversity of IGS lengths revealed variants from ∼ 2 kb in Fagus, and Quercus up to 5.3 kb in Castanea, and a lack of correlation between the number of variants and the number of rDNA loci in several species. Methylation of 25S Bam HI site was confirmed in all species and detected for the first time in the 18S of Q. suber and Q. faginea. These results provide important clues for the evolutionary trends of the rDNA 25S-18S IGS in the Fagaceae family.

Show MeSH
Sequence alignment of the R1 and R2 sub-repeats of Fagus sylvatica.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4043768&req=5

pone-0098678-g003: Sequence alignment of the R1 and R2 sub-repeats of Fagus sylvatica.

Mentions: Self-comparison of each IGS using dot plot analysis revealed that each spacer was composed of an initial repeated region (Figure 2) flanked by two unique regions. The repeated regions showed a typical arrangement of Quercus spp. 25S-18S IGSs, with a size of 544 bp in Fs1.9 (position 132 to 685 downstream the 25S end) and 522 bp in Qs2 (position 74 to 595 downstream the 25S end). In Fs1.9, this region consisted of conserved and imperfect copies of two types of sub-repeats disposed uninterruptedly: R1 and R2 (Figure 3). We have found 33 copies of the R1 sub-repeats (around 10 bp in length) showing substitutions in one, two or three nucleotides. The 10 bp long R2 sub-repeat (27 copies of R2 in total) showed variants differing by one or two base substitutions, although some truncated and partially deleted copies are also present. In Qs2, the SR pattern is more intricate, showing conserved and imperfect copies of three types of sub-repeats (A-sub-repeats, B-sub-repeats, and C-sub-repeats) interrupted by a less or unrelated sequence. The A-sub-repeats were around 21 bp long, varying in one, two or three nucleotides (Figure 4). The B-sub-repeats were the most variable with a maximum of 5 base substitutions in seven copies of around 17 bp in length (Figure 4). Nine copies of C-sub-repeats (around 11 bp long) and its variants, differing by one nucleotide substitution and insertions/deletions of the C-stretch and G-stretch, were also found (Figure 4).


Molecular organization of the 25S-18S rDNA IGS of Fagus sylvatica and Quercus suber: a comparative analysis.

Inácio V, Rocheta M, Morais-Cecílio L - PLoS ONE (2014)

Sequence alignment of the R1 and R2 sub-repeats of Fagus sylvatica.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4043768&req=5

pone-0098678-g003: Sequence alignment of the R1 and R2 sub-repeats of Fagus sylvatica.
Mentions: Self-comparison of each IGS using dot plot analysis revealed that each spacer was composed of an initial repeated region (Figure 2) flanked by two unique regions. The repeated regions showed a typical arrangement of Quercus spp. 25S-18S IGSs, with a size of 544 bp in Fs1.9 (position 132 to 685 downstream the 25S end) and 522 bp in Qs2 (position 74 to 595 downstream the 25S end). In Fs1.9, this region consisted of conserved and imperfect copies of two types of sub-repeats disposed uninterruptedly: R1 and R2 (Figure 3). We have found 33 copies of the R1 sub-repeats (around 10 bp in length) showing substitutions in one, two or three nucleotides. The 10 bp long R2 sub-repeat (27 copies of R2 in total) showed variants differing by one or two base substitutions, although some truncated and partially deleted copies are also present. In Qs2, the SR pattern is more intricate, showing conserved and imperfect copies of three types of sub-repeats (A-sub-repeats, B-sub-repeats, and C-sub-repeats) interrupted by a less or unrelated sequence. The A-sub-repeats were around 21 bp long, varying in one, two or three nucleotides (Figure 4). The B-sub-repeats were the most variable with a maximum of 5 base substitutions in seven copies of around 17 bp in length (Figure 4). Nine copies of C-sub-repeats (around 11 bp long) and its variants, differing by one nucleotide substitution and insertions/deletions of the C-stretch and G-stretch, were also found (Figure 4).

Bottom Line: In the NTS: the sub-repeats structure in Beech is more organized than in Cork Oak, sharing some short motifs which results in the lowest sequence similarity of the entire IGS; the AT-rich region differed in both spacers by a GC-rich block inserted in Cork Oak.Methylation of 25S Bam HI site was confirmed in all species and detected for the first time in the 18S of Q. suber and Q. faginea.These results provide important clues for the evolutionary trends of the rDNA 25S-18S IGS in the Fagaceae family.

View Article: PubMed Central - PubMed

Affiliation: Centre for Botany Applied to Agriculture (CBAA), Instituto Superior de Agronomia, University of Lisbon, Lisbon, Portugal.

ABSTRACT
The 35S ribosomal DNA (rDNA) units, repeated in tandem at one or more chromosomal loci, are separated by an intergenic spacer (IGS) containing functional elements involved in the regulation of transcription of downstream rRNA genes. In the present work, we have compared the IGS molecular organizations in two divergent species of Fagaceae, Fagus sylvatica and Quercus suber, aiming to comprehend the evolution of the IGS sequences within the family. Self- and cross-hybridization FISH was done on representative species of the Fagaceae. The IGS length variability and the methylation level of 18 and 25S rRNA genes were assessed in representatives of three genera of this family: Fagus, Quercus and Castanea. The intergenic spacers in Beech and Cork Oak showed similar overall organizations comprising putative functional elements needed for rRNA gene activity and containing a non-transcribed spacer (NTS), a promoter region, and a 5'-external transcribed spacer. In the NTS: the sub-repeats structure in Beech is more organized than in Cork Oak, sharing some short motifs which results in the lowest sequence similarity of the entire IGS; the AT-rich region differed in both spacers by a GC-rich block inserted in Cork Oak. The 5'-ETS is the region with the higher similarity, having nonetheless different lengths. FISH with the NTS-5'-ETS revealed fainter signals in cross-hybridization in agreement with the divergence between genera. The diversity of IGS lengths revealed variants from ∼ 2 kb in Fagus, and Quercus up to 5.3 kb in Castanea, and a lack of correlation between the number of variants and the number of rDNA loci in several species. Methylation of 25S Bam HI site was confirmed in all species and detected for the first time in the 18S of Q. suber and Q. faginea. These results provide important clues for the evolutionary trends of the rDNA 25S-18S IGS in the Fagaceae family.

Show MeSH