In plants, gametogenesis occurs late in development, and somatic mutations can therefore be transmitted to the next generation. Longer periods of growth are believed to result in an increase in the number of cell divisions before gametogenesis, with a concomitant increase in mutations arising due to replication errors. However, there is little experimental evidence addressing how many cell divisions occur before gametogenesis. Here, we measured loss of telomeric DNA and accumulation of replication errors in Arabidopsis with short and long life spans to determine the number of replications in lineages leading to gametes. Surprisingly, the number of cell divisions within the gamete lineage is nearly independent of both life span and vegetative growth. One consequence of the relatively stable number of replications per generation is that older plants may not pass along more somatically acquired mutations to their offspring. We confirmed this hypothesis by genomic sequencing of progeny from young and old plants. This independence can be achieved by hierarchical arrangement of cell divisions in plant meristems where vegetative growth is primarily accomplished by expansion of cells in rapidly dividing meristematic zones, which are only rarely refreshed by occasional divisions of more quiescent cells. We support this model by 5-ethynyl-2'-deoxyuridine retention experiments in shoot and root apical meristems. These results suggest that stem-cell organization has independently evolved in plants and animals to minimize mutations by limiting DNA replication.

• Keywords
• germline, mismatch repair, mutation rate, shoot apical meristem, telomeres,
• MeSH
• Mutation Accumulation MeSH
• Arabidopsis genetics   growth & development   MeSH
• Diploidy MeSH
• Genome, Plant genetics   MeSH
• Plant Roots genetics   growth & development   MeSH
• Meristem genetics   growth & development   MeSH
• Mutation genetics   MeSH
• Gene Expression Regulation, Plant MeSH
• DNA Replication genetics   MeSH
• Plant Cells MeSH
• Sequence Analysis, DNA MeSH
• Plant Stems genetics   growth & development   MeSH
• Plant Shoots genetics   growth & development   MeSH
• Germ Cells growth & development   MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH

We studied the in trans-silencing capacities of a transgene locus that carried the neomycin phosphotransferase II reporter gene linked to the 35S promoter in an inverted repeat (IR). This transgene locus was originally posttranscriptionally silenced but switched to a transcriptionally silenced epiallele after in vitro tissue culture. Here, we show that both epialleles were strongly methylated in the coding region and IR center. However, by genomic sequencing, we found that the 1.0 kb region around the transcription start site was heavily methylated in symmetrical and non-symmetrical contexts in transcriptionally but not in posttranscriptionally silenced epilallele. Also, the posttranscriptionally silenced epiallele could trans-silence and trans-methylate homologous transgene loci irrespective of their genomic organization. We demonstrate that this in trans-silencing was accompanied by the production of small RNA molecules. On the other hand, the transcriptionally silenced variant could neither trans-silence nor trans-methylate homologous sequences, even after being in the same genetic background for generations and meiotic cycles. Interestingly, 5-aza-2-deoxy-cytidine-induced hypomethylation could partially restore signaling from the transcriptionally silenced epiallele. These results are consistent with the hypothesis that non-transcribed highly methylated IRs are poor silencers of homologous loci at non-allelic positions even across two generations and that transcription of the inverted sequences is essential for their trans-silencing potential.

• MeSH
• Alleles MeSH
• Epigenesis, Genetic * MeSH
• Plants, Genetically Modified genetics   metabolism   MeSH
• Kanamycin Kinase genetics   metabolism   MeSH
• DNA Methylation MeSH
• RNA, Untranslated analysis   MeSH
• Repetitive Sequences, Nucleic Acid * MeSH
• Genes, Reporter MeSH
• Nicotiana genetics   MeSH
• Transgenes * MeSH
• Gene Silencing * MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Kanamycin Kinase MeSH
• RNA, Untranslated MeSH