Characterization of the level, target sites and inheritance of cytosine methylation in tomato nuclear DNA

Abstract

The tomato nuclear genome was determined to have a G + C content of 37% which is among the lowest reported for any plant species. Non-coding regions have a G + C content even lower (32% average) whereas coding regions are considerably richer in G + C (46%). 5-methyl cytosine was the only modified base detected and on average 23% of the cytosine residues are methylated. Immature tissues and protoplasts have significantly lower levels of cytosine methylation (average 20%) than mature tissues (average 25%). Mature pollen has an intermediate level of methylation (22%). Seeds gave the highest value (27%), suggesting de novo methylation after pollination and during seed development. Based on isoschizomer studies we estimate 55% of the CpG target sites (detected by Msp I/Hpa II) and 85% of the CpNpG target sites (detected by Bst NI/Eco RI) are methylated. Unmethylated target sites (both CpG and CpNpG) are not randomly distributed throughout the genome, but frequently occur in clusters. These clusters resemble CpG islands recently reported in maize and tobacco. The low G + C content and high levels of cytosine methylation in tomato may be due to previous transitions of 5mC----T. This is supported by the fact that G + C levels are lowest in non-coding portions of the genome in which selection is relaxed and thus transitions are more likely to be tolerated. This hypothesis is also supported by the general deficiency of methylation target sites in the tomato genome, especially in non-coding regions. Using methylation isoschizomers and RFLP analysis we have also determined that polymorphism between plants, for cytosine methylation at allelic sites, is common in tomato. Comparing DNA from two tomato species, 20% of the polymorphisms detected by Bst NI/Eco RII could be attributed to differential methylation at the CpNpG target sites. With Msp I/Hpa II, 50% of the polymorphisms were attributable to methylation (CpG and CpNpG sites). Moreover, these polymorphisms were demonstrated to be inherited in a mendelian fashion and to co-segregate with the methylation target site and thus do not represent variation for transacting factors that might be involved in methylation of DNA. The potential role of heritable methylation polymorphism in evolution of gene regulation and in RFLP studies is discussed.

DNA methylation polymorphism and stability in Chinese indica hybrid rice

“…A further concern for inbred rice breeders is the segregation patterns of heterozygous methylation alleles. In heterozygous methylation alleles of tomato (Solanum lycopersicum) [34] and Arabidopsis [35], both consistency and violation of Mendelian rules have been observed; this was also the case in this study (Table 8; Figure S1 in Supporting Information). Therefore, it is not expected that the frequency of heterozygous methylation alleles in the segregation generation will be the same as that of the genetic alleles.…”

Transposition pattern of a modified Ds element in tomato

“…A comparison between the sequenced plant DNAs did not show any general similarities for either the 8 bp sequences flanking Ds-r or more extended stretches (100-300 bp) of Ds-r flanking DNA. The average CG content of these sequences was 39~o, which is in line with the average CG content of tomato [39]. Homology was, however, found between plant DNAs flanking Ds-r elements clusterly integrated on chromosome 10.…”

Analysis of DNA methylation of perennial ryegrass under drought using the methylation-sensitive amplification polymorphism (MSAP) technique

“…In the control and drought stresstreated rice, the overall level of DNA methylation ranged from 24.55 to 13.52 % . In tomato, immature tissues and protoplasts of cytosine methylation levels, average 20 %, are significantly lower than mature tissues, average 25 % (Messeguer et al 1991). Perennial ryegrass has a large and complex genome that is approximately 2,695 Mb in size (Byrne et al 2011), which most likely contains a large number of duplicated sequences (Lewin 2008).…”