An under-methylated region in the spacer of ribosomal RNA genes of Lilium henryi (original) (raw)
Related papers
2012
Summary. The methylation pattern of rRNA genes in reconstructed barley karyotypes ( Hordeum vulgare L.) with altered position or structure of the two nucleolus organizers (NORs) was studied by double digestion with EcoRI and HhaI and molecular hybridization with specifi c rDNA probes. The lack of the whole rRNA gene cluster residing in chromosome 6H in the homozygous deletion line T-35 led to an increased hypomethylation of the rRNA genes in the remaining NOR5H. On the other hand, repositioning by translocation of the distant part of the split NOR6H to the short arm of the chromosome 5H in the translocation line T-21 did not correlate with any signifi cant changes in the methylation of –GCGC– sequences in rDNA units. In addition, the length of the intergenic spacer (IGS) in both reconstructed barley lines T35 and T21 was also analyzed. Our results showed the same length of IGS in the long rDNA repeat as compared with previously published rDNA clone (GenBank HQ825319). However, the s...
Pattern and degree of methylation in ribosomal RNA genes of Cucurbita pepo L
Plant Molecular Biology, 1994
Methylation with respect to its degree and distribution throughout the 18S, 5.8S and 25S rRNA gene clusters (rDNA) and within single rDNA repeats in seedlings of the higher plant Cucurbita pepo L. (zucchini) was investigated. In this plant, which is characterized by several thousand repeats, at least 70 ~o are completely or nearly completely methylated in CpGs and to a lower degree in CpNpGs. Detailed methylation analysis revealed that a fraction of about 3-4 ~o of all repeats is hypomethylated near the transcription initiation site (TIS) which may indicate the fraction of active repeats in C. pepo. However, a different fraction (3-4 ~o of all repeats) which is not methylated in all sites tested (including those at the TIS) is present in C. pepo and may thus represent active but differentially methylated rDNA. The results are discussed in context of recent models on methytation and gene activity.
Comparative analysis of DNA methylation in tobacco heterochromatic sequences
2000
Cytosine methylation levels and susceptibility to drug-induced hypomethylation have been studied in several Nicotiana tabacum (tobacco) DNA repetitive sequences. It has been shown using HapII, MspI, BamHI and Sau3AI methylation-sensitive restriction enzymes that the degree of 5 Hm C m CG-3 H methylation varied signi¢cantly between different repeats. There were almost saturation levels of 5-methylcytosine at the inner (3 H) cytosine position and variable degrees of methylation at the outer (5 H) cytosine at the enzyme recognition sites. The non-transcribed high copy satellite sequences (HRS60, GRS) displayed signi¢cant heterogeneity in methylation of their basic units while middle repetitive sequences (R8.1, GRD5, 5S rDNA) were more uniformly modi¢ed at both cytosine residues. Dihydroxypropyladenine (DHPA) treatment, which is thought to reduce DNA methyltransferase activity by increasing S-adenosylhomocysteine levels, resulted in extensive demethylation of the outer cytosine in all repeats, and the partial hypomethylation of cytosines at the inner positions in less densely methylated repeats such as HRS60 and GRS. The results suggest that hypomethylation of 5 Hm C m CG-3 H sites with DHPA is a gradual non-random process proceeding in the direction m C m CG 3 C m CG 3 CCG. The 18S-5.8S-25S rDNA was remarkably hypomethylated relative to the 5S rDNA at all restriction sites studied. Fluorescence in-situ hybridization showed that DNA decondensation within and between the 18S-5.8S-25S and 5S rDNA loci was variable in different nuclei. All nuclei had condensed and decondensed sequence. The chromatin of 18S-5.8S-25S rDNA was more readily digested with micrococcal nuclease than the 5S rDNA suggesting that the overall levels of decondensation were higher for 18S-5.8S-25S rDNA. Variable decondensation patterns within and between loci were also observed for GRS and HRS60. Cytosine methylation of the tobacco repeats is discussed with respect to transcription, overall levels of condensation and overall structure.
Euphytica, 2000
The 18s-5.8s-25s ribosomal gene (18s-25s rDNA) in higher plants is present in multiple copies, on different chromosomes, as tandemly repeated units. Among the multiple BamHI sites that occur in the repeat unit, only the site in the middle of the 25s rRNA coding region is methylated in most cereals and pulses. BamHI restriction enzyme analyses of the mungbean 18s-25s rDNA showed the presence of two populations. Nearly 50% of the 18s-25s rDNA population had BamHI site situated in the internal transcribed spacer region (ITS-BamHI) resistant to cleavage by BamHI. The amplified ITS fragment was completely digestible by BamHI showing that partial cleavage by BamHI is not due to variation in the recognition sequence but most probably due to methylation. The complete cleavage of the ITS-BamHI site by BstYI that recognizes BamHI site but is insensitive to methylation confirms that the ITS-BamHI site is methylated. Methylation is probably due to the presence of a guanosine residue adjacent to the 3′ cytosine in the recognition sequence.
Unique Epigenetic Features of Ribosomal RNA Genes (rDNA) in Early Diverging Plants (Bryophytes)
Frontiers in Plant Science, 2019
Introduction: In plants, the multicopy genes encoding ribosomal RNA (rDNA) typically exhibit heterochromatic features and high level of DNA methylation. Here, we explored rDNA methylation in early diverging land plants from Bryophyta (15 species, 14 families) and Marchantiophyta (4 species, 4 families). DNA methylation was investigated by methylation-sensitive Southern blot hybridization in all species. We also carried out whole genomic bisulfite sequencing in Polytrichum formosum (Polytrichaceae) and Dicranum scoparium (Dicranaceae) and used available model plant methyloms (Physcomitrella patents and Marchantia polymorpha) to determine rDNA unit-wide methylation patterns. Chromatin structure was analyzed using fluorescence in situ hybridization (FISH) and immunoprecipitation (CHIP) assays. Results: In contrast to seed plants, bryophyte rDNAs were efficiently digested with methylation-sensitive enzymes indicating no or low levels of CG and CHG methylation in these loci. The rDNA methylom analyses revealed variation between species ranging from negligible (<3%, P. formosum, P. patens) to moderate (7 and 17% in M. polymorpha and D. scoparium, respectively) methylation levels. There were no differences between coding and noncoding parts of rDNA units and between gametophyte and sporophyte tissues. However, major satellite repeat and transposable elements were heavily methylated in P. formosum and D. scoparium. In P. formosum rDNA, the euchromatic H3K4m3 and heterochromatic H3K9m2 histone marks were nearly balanced contrasting the angiosperms data where H3K9m2 typically dominates rDNA chromatin. In moss interphase nuclei, rDNA was localized at the nucleolar periphery and its condensation level was high. Conclusions: Unlike seed plants, the rRNA genes seem to escape global methylation machinery in bryophytes. Distinct epigenetic features may be related to rDNA expression and the physiology of these early diverging plants that exist in haploid state for most of their life cycles.
Adaptive methylation pattern of ribosomal DNA in wild barley from Israel
2005
Forty four accessions of wild barley, H. spontaneum were assayed to study the methylation status of ribosomal DNA repeat units. For this purpose, BamHI and HpaII, which are, methylation sensitive restriction enzymes and MspI, which is methyl insensitive enzyme, were used for restriction digestion. Southern blots were hybridized with pTa71 probe, which represented a complete ribosomal DNA repeat unit from bread wheat. Wild barley material studied belonged to two ecologically diverse climatic and edaphic microsites (the "Evolution Canyon" at Lower Nahal Oren, Mount Carmel, and Tabigha, Eastern Upper Galilee Mountains), from Israel, each having two different ecogeographcally contrasting microniches. HpaII did not cleave ribosomal DNA, while MspI gave a pattern typical of ribosomal DNA. In contrast, a total of 23 BamHI phenotypes were observed due to methylation. Same rDNA repeat length units exhibited different methylation status at different microsites and miocroniches, as inferred from RFLPs observed. Differential methylation status of same rDNA repeat unit seems to be associated with ecological conditions dominating at a particular microsite or microniche suggesting a role of natural selection in determining methylation patterns.
Structure and variation in ribosomal RNA genes of pea
Plant Molecular Biology, 1987
A complete ribosomal DNA (rDNA) repeat unit has been cloned from the genome of Pisum sativum (garden pea) and used to construct a map containing a total of 58 cleavage sites for 23 different restriction enzymes. Regions encoding 18s and 25s ribosomal RNA (rRNA) were identified by R-loop analysis. A 180 bp sequence element is repeated eight times in the intergenic 'nontranscribed spacer' (NTS) region, as defined by eight evenly spaced RsaI cleavage sites. Sequence heterogeneity among these elements (subrepeats) is indicated by the presence of an NcoI site within the five RsaI subrepeats distal to the 25s rRNA gene but not in the three subrepeats proximal to this gene, and also by the presence of an additional RsaI cleavage site in one subrepeat.
Frontiers in Genetics, 2020
Apomixis, an asexual mode of reproduction through seeds, has immense scope for crop improvement due to its ability to fix hybrid vigor. In C. ciliaris, a predominantly apomictically reproducing range grass, apomixis is genetically controlled by an apospory-specific-genomic-region (ASGR) which is enriched with retrotransposons. Earlier studies showed insertional polymorphisms of a few ASGRspecific retrotransposons between apomictic and sexual plants of C. ciliaris. REs are mainly regulated at the transcriptional level through cytosine methylation. To understand the possible association of ASGR-specific retrotransposon to apomixis, the extent and pattern of differential methylation of Gy163 RE and its impact on transcription were investigated in two genotypes each of apomictic and sexual plants of C. ciliaris. We observed that Gy163 encodes for an integrase domain of RE Ty3-Gypsy, is differentially methylated between reproductive tissues of apomictic and sexual plants. However, leaf tissues did not exhibit differential methylation between apomictic and sexual plants. Among the three contexts (CG, CHG, and CHH) of cytosine methylation, the maximum variation was observed in CHH context in reproductive (at aposporous initial and mature embryo sac stages) tissues of apomictic plants implicating RdDM pathway in methylation of Gy163. Quantitative PCR analysis showed that Gy163 transcripts are expressed more in the reproductive tissues of apomictic plants compared to that in the sexual plants, which was negatively correlated with the methylation level. Thus, the study helps in understanding the role of RE present in ASGR in epigenetic regulation of apomictic mode of reproduction in C. ciliaris.
Non-symmetrical cytosine methylation in tobacco pollen DNA
Plant Molecular Biology, 1996
We have detected sequence-specific non-symmetrical cytosine methylation within a t40 bp region of the promoter for the tobacco auxin-binding protein gene T85 in pollen DNA. Direct sequencing of the population of bisulphite reaction products showed that, in this region, 10 out of a possible 49 cytosine residues were methylated at a high frequency in pollen whereas the corresponding region from somatic cells (leaf DNA) did not show a detectable level of methylation. The context of these sites was I x mSCpTpC, 1 x mSCpGpT, l x mSCpCpT, 2x mSCpTpT, 2x mSCpGpG, and 3x mSCpApT of which only mSCpGpG and mSCpGpT fitted the consensus sequence for symmetrical methylation in plants.