Abstract
One of the major quantitative trait loci for flowering time in maize, the Vegetative to generative transition 1 (Vgt1) locus, corresponds to an upstream (70 kb) noncoding regulatory element of ZmRap2.7, a repressor of flowering. At Vgt1, a miniature transposon (MITE) insertion into a conserved noncoding sequence was previously found to be highly associated with early flowering in independent studies. Because cytosine methylation is known to be associated with transposons and to influence gene expression, we aimed to investigate how DNA methylation patterns in wild-type and mutant Vgt1 correlate with ZmRap2.7 expression. The methylation state at Vgt1 was assayed in leaf samples of maize inbred and F1 hybrid samples, and at the syntenic region in sorghum. The Vgt1-linked conserved noncoding sequence was very scarcely methylated both in maize and sorghum. However, in the early maize Vgt1 allele, the region immediately flanking the highly methylated MITE insertion was significantly more methylated and showed features of methylation spreading. Allele-specific expression assays revealed that the presence of the MITE and its heavy methylation appear to be linked to altered ZmRap2.7 transcription. Although not providing proof of causative connection, our results associate transposon-linked differential methylation with allelic state and gene expression at a major flowering time quantitative trait locus in maize.
Highlights
One of the major quantitative trait loci for flowering time in maize, the Vegetative to generative transition 1 (Vgt1) locus, corresponds to an upstream (70 kb) noncoding regulatory element of ZmRap2.7, a repressor of flowering
A maize–sorghum–rice evolutionarily conserved noncoding sequence (CNS) (Freeling and Subramaniam 2009) was identified within Vgt1; in early Vgt1 alleles, this CNS is interrupted by the insertion of a miniature transposon (MITE) belonging to the Tourist family (Salvi et al 2007), and this insertional polymorphism has been repeatedly identified as strongly associated with flowering time in independent studies (Salvi et al 2007; Ducrocq et al 2008; Buckler et al 2009; Hung et al 2012; Truntzler et al 2012)
Temporal profile of ZmRap2.7 expression Leaf tissues sampled at four different developmental time points (V1, V3, V5, and V7) were used to determine ZmRap2.7 expression by means of qPCR in five different maize lines: B73, N28, R22 and Gaspé Flint and C22-4
Summary
One of the major quantitative trait loci for flowering time in maize, the Vegetative to generative transition 1 (Vgt1) locus, corresponds to an upstream (70 kb) noncoding regulatory element of ZmRap2.7, a repressor of flowering. Not providing proof of causative connection, our results associate transposon-linked differential methylation with allelic state and gene expression at a major flowering time quantitative trait locus in maize. A maize–sorghum–rice evolutionarily conserved noncoding sequence (CNS) (Freeling and Subramaniam 2009) was identified within Vgt; in early Vgt alleles, this CNS is interrupted by the insertion of a miniature transposon (MITE) belonging to the Tourist family (Salvi et al 2007), and this insertional polymorphism has been repeatedly identified as strongly associated with flowering time in independent studies (Salvi et al 2007; Ducrocq et al 2008; Buckler et al 2009; Hung et al 2012; Truntzler et al 2012). We focused in particular on the CNS/ MITE region with the objective of improving our understanding of the involvement of conserved DNA elements in determining phenotypic variation
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