In plants, genome duplication followed by genome diversification and selection is recognized as a major evolutionary process. Rapid epigenetic and genetic changes that affect the transcription of parental genes are frequently observed after polyploidization. The pattern of alternative splicing is also frequently altered, yet the related molecular processes remain largely unresolved. Here, we study the inheritance and expression of parental variants of three floral organ identity genes in allotetraploid tobacco. DEFICIENS and GLOBOSA are B-class genes, and AGAMOUS is a C-class gene. Parental variants of these genes were found to be maintained in the tobacco genome, and the respective mRNAs were present in flower buds in comparable amounts. However, among five tobacco cultivars, we identified two in which the majority of paternal GLOBOSA pre-mRNA transcripts undergo exon 3 skipping, producing an mRNA with a premature termination codon. At the DNA level, we identified a G-A transition at the very last position of exon 3 in both cultivars. Although alternative splicing resulted in a dramatic decrease in full-length paternal GLOBOSA mRNA, no phenotypic effect was observed. Our finding likely serves as an example of the initiation of homoeolog diversification in a relatively young polyploid genome.

• Klíčová slova
• Alternative splicing, Floral genes, Flowering, Polyploidy, Tobacco,
• MeSH
• alternativní sestřih genetika   MeSH
• bodová mutace genetika   MeSH
• exony genetika   MeSH
• genetická transkripce * MeSH
• homeodoménové proteiny biosyntéza   genetika   MeSH
• nukleotidy genetika   MeSH
• polyploidie MeSH
• prekurzory RNA genetika   MeSH
• regulace genové exprese u rostlin MeSH
• rostlinné proteiny biosyntéza   genetika   MeSH
• tabák genetika   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• GLOBOSA protein, plant MeSH Prohlížeč
• homeodoménové proteiny MeSH
• nukleotidy MeSH
• prekurzory RNA MeSH
• rostlinné proteiny MeSH

BACKGROUND: DNA methylation plays a key role in development, contributes to genome stability, and may also respond to external factors supporting adaptation and evolution. To connect different types of stimuli with particular biological processes, identifying genome regions with altered 5-methylcytosine distribution at a genome-wide scale is important. Many researchers are using the simple, reliable, and relatively inexpensive Methylation Sensitive Amplified Polymorphism (MSAP) method that is particularly useful in studies of epigenetic variation. However, electrophoretic patterns produced by the method are rather difficult to interpret, particularly when MspI and HpaII isoschizomers are used because these enzymes are methylation-sensitive, and any C within the CCGG recognition motif can be methylated in plant DNA. RESULTS: Here, we evaluate MSAP patterns with respect to current knowledge of the enzyme activities and the level and distribution of 5-methylcytosine in plant and vertebrate genomes. We discuss potential caveats related to complex MSAP patterns and provide clues regarding how to interpret them. We further show that addition of combined HpaII + MspI digestion would assist in the interpretation of the most controversial MSAP pattern represented by the signal in the HpaII but not in the MspI profile. CONCLUSIONS: We recommend modification of the MSAP protocol that definitely discerns between putative hemimethylated mCCGG and internal CmCGG sites. We believe that our view and the simple improvement will assist in correct MSAP data interpretation.

• MeSH
• 5-methylcytosin chemie   MeSH
• DNA rostlinná genetika   MeSH
• epigeneze genetická MeSH
• metylace DNA * MeSH
• obratlovci genetika   MeSH
• polymorfismus genetický * MeSH
• restrikční mapování MeSH
• tabák genetika   MeSH
• techniky amplifikace nukleových kyselin metody   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• 5-methylcytosin MeSH
• DNA rostlinná MeSH

The widespread occurrence of epigenetic alterations in allopolyploid species deserves scrutiny that DNA methylation systems may be perturbed by interspecies hybridization and polyploidization. Here we studied the genes involved in DNA methylation in Nicotiana tabacum (tobacco) allotetraploid containing S and T genomes inherited from Nicotiana sylvestris and Nicotiana tomentosiformis progenitors. To determine the inheritance of DNA methyltransferase genes and their expression patterns we examined three major DNA methyltransferase families (MET1, CMT3 and DRM) from tobacco and the progenitor species. Using Southern blot hybridization and PCR-based methods (genomic CAPS), we found that the parental loci of these gene families are retained in tobacco. Homoeologous expression was found in all tissues examined (leaf, root, flower) suggesting that DNA methyltransferase genes were probably not themselves targets of uniparental epigenetic silencing for over thousands of generations of allotetraploid evolution. The level of CG and CHG methylation of selected high-copy repeated sequences was similar and high in tobacco and its diploid progenitors. We speculate that natural selection might favor additive expression of parental DNA methyltransferase genes maintaining high levels of DNA methylation in tobacco, which has a repeat-rich heterochromatic genome.

• MeSH
• diploidie MeSH
• DNA rostlinná genetika   MeSH
• DNA-(cytosin-5-)methyltransferasa klasifikace   genetika   metabolismus   MeSH
• epigeneze genetická MeSH
• exprese genu MeSH
• fylogeneze MeSH
• genom rostlinný MeSH
• klonování DNA MeSH
• metylace DNA genetika   MeSH
• molekulární sekvence - údaje MeSH
• multigenová rodina * MeSH
• polyploidie MeSH
• repetitivní sekvence nukleových kyselin MeSH
• rostlinné geny * MeSH
• sekvence nukleotidů MeSH
• selekce (genetika) MeSH
• tabák enzymologie   genetika   MeSH
• tkáňová distribuce MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• DNA rostlinná MeSH
• DNA-(cytosin-5-)methyltransferasa MeSH

In plants, 5S rRNA genes (5S rDNA) encoding 120-nt structural RNA molecules of ribosomes are organized in tandem arrays comprising thousands of units. Failure to correctly terminate transcription would generate longer inaccurately processed transcripts interfering with ribosome biogenesis. Hence multiple termination signals occur immediately after the 5S rRNA coding sequence. To obtain information about the efficiency of termination of 5S rDNA transcription in plants we analyzed 5S rRNA pools in three Nicotiana species, N. sylvestris, N. tomentosiformis and N. tabacum. In addition to highly abundant 120-nt 5S rRNA transcripts, we also detected RNA species composed of a genic region and variable lengths of intergenic sequences. These genic-intergenic RNA molecules occur at a frequency severalfold lower than the mature 120-nt transcripts, and are posttranscriptionally modified by polyadenylation at their 3' end in contrast to 120-nt transcripts. An absence of 5S small RNAs (smRNA) argue against a dominant role for the smRNA biosynthesis pathway in the degradation of aberrant 5S rRNA in Nicotiana. This work is the first description of polyadenylated 5S rRNA species in higher eukaryotes originating from a read-through transcription into the intergenic spacer. We propose that polyadenylation may function in a "quality control" pathway ensuring that only correctly processed molecules enter the ribosome biogenesis.

• MeSH
• Arabidopsis genetika   MeSH
• genetická transkripce * MeSH
• intergenová DNA * MeSH
• malá interferující RNA metabolismus   MeSH
• messenger RNA metabolismus   MeSH
• modely genetické MeSH
• molekulární sekvence - údaje MeSH
• polyadenylace * MeSH
• regulace genové exprese u rostlin * MeSH
• RNA ribozomální 5S genetika   MeSH
• sekvence nukleotidů MeSH
• sekvenční homologie nukleových kyselin MeSH
• tabák genetika   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• intergenová DNA * MeSH
• malá interferující RNA MeSH
• messenger RNA MeSH
• RNA ribozomální 5S MeSH