title: CaMV-35S promoter sequence-specific DNA methylation in lettuce.
abstract: We found 35S promoter sequence-specific DNA methylation in lettuce. Additionally, transgenic lettuce plants having a modified 35S promoter lost methylation, suggesting the modified sequence is subjected to the methylation machinery. We previously reported that cauliflower mosaic virus 35S promoter-specific DNA methylation in transgenic gentian (Gentiana triflora × G. scabra) plants occurs irrespective of the copy number and the genomic location of T-DNA, and causes strong gene silencing. To confirm whether 35S-specific methylation can occur in other plant species, transgenic lettuce (Lactuca sativa L.) plants with a single copy of the 35S promoter-driven sGFP gene were produced and analyzed. Among 10 lines of transgenic plants, 3, 4, and 3 lines showed strong, weak, and no expression of sGFP mRNA, respectively. Bisulfite genomic sequencing of the 35S promoter region showed hypermethylation at CpG and CpWpG (where W is A or T) sites in 9 of 10 lines. Gentian-type de novo methylation pattern, consisting of methylated cytosines at CpHpH (where H is A, C, or T) sites, was also observed in the transgenic lettuce lines, suggesting that lettuce and gentian share similar methylation machinery. Four of five transgenic lettuce lines having a single copy of a modified 35S promoter, which was modified in the proposed core target of de novo methylation in gentian, exhibited 35S hypomethylation, indicating that the modified sequence may be the target of the 35S-specific methylation machinery.
authors.affiliation: Graduate School of Life and Environmental Sciences, Osaka Prefecture University, 1-1 Gakuen, Nakaku, Sakai, Osaka, 599-8531, Japan.|Miyoshi Co., Ltd., 187 Kamisasao, Kobuchizawa, Hokuto, Yamanashi, 408-0041, Japan.
authors.forename: Azusa
authors.initials: A
authors.lastname: Okumura
authors.affiliation: Graduate School of Life and Environmental Sciences, Osaka Prefecture University, 1-1 Gakuen, Nakaku, Sakai, Osaka, 599-8531, Japan.
authors.forename: Asahi
authors.initials: A
authors.lastname: Shimada
authors.affiliation: Graduate School of Life and Environmental Sciences, Osaka Prefecture University, 1-1 Gakuen, Nakaku, Sakai, Osaka, 599-8531, Japan.|Tempstaff People Co., Ltd, 1-5 Shinsakae, Naka-ku, Nagoya, Aichi, 460-8482, Japan.
authors.forename: Satoshi
authors.initials: S
authors.lastname: Yamasaki
authors.affiliation: Graduate School of Life and Environmental Sciences, Osaka Prefecture University, 1-1 Gakuen, Nakaku, Sakai, Osaka, 599-8531, Japan.
authors.forename: Takuya
authors.initials: T
authors.lastname: Horino
authors.affiliation: Graduate School of Life and Environmental Sciences, Osaka Prefecture University, 1-1 Gakuen, Nakaku, Sakai, Osaka, 599-8531, Japan.
authors.forename: Yuji
authors.initials: Y
authors.lastname: Iwata
authors.affiliation: Graduate School of Life and Environmental Sciences, Osaka Prefecture University, 1-1 Gakuen, Nakaku, Sakai, Osaka, 599-8531, Japan.
authors.forename: Nozomu
authors.initials: N
authors.lastname: Koizumi
authors.affiliation: Iwate Biotechnology Research Center, 22-174-4 Narita, Kitami, Iwate, 024-0003, Japan.
authors.forename: Masahiro
authors.initials: M
authors.lastname: Nishihara
authors.affiliation: Graduate School of Life and Environmental Sciences, Osaka Prefecture University, 1-1 Gakuen, Nakaku, Sakai, Osaka, 599-8531, Japan. mishiba@plant.osakafu-u.ac.jp.
authors.forename: Kei-ichiro
authors.initials: K
authors.lastname: Mishiba
mesh_terms: D001483:Base Sequence; D017796:Caulimovirus / Q000235:genetics*; D019175:DNA Methylation; D018506:Gene Expression Regulation, Plant; D020868:Gene Silencing; D017930:Genes, Reporter; D030012:Gentiana / Q000235:genetics; D018545:Lactuca / Q000235:genetics*; D030821:Plants, Genetically Modified; D011401:Promoter Regions, Genetic / Q000235:genetics*; D017422:Sequence Analysis, DNA
keywords: 35S promoter; DNA methylation; De novo methylation; Gentian; Lettuce; Transgene silencing