DNA modification of a maize transposable element correlates with loss of activity.

Abstract

An unstable allele of the bronze 2 (bz2) locus was isolated from a Robertson's Mutator Zea mays line containing a family of active transposable Mu elements. This mutation is somatically unstable, resulting in numerous revertant purple sectors on a bronze kernel. By following the variegated kernel phenotype through two generations, several lineages have been identified that have a distorted transmission of the mutant phenotype (fewer variegated kernels are produced than expected). Southern blot analysis ofMu elements in these plants demonstrates a correlation between an inhibition of digestion of Mu elements by certain restriction enzymes and the loss of somatic reversion at the mutant allele. The DNA modification can occur in all the Mu elements in a plant within one generation; however, plants have been identified that contain both modified and unmodified elements, suggesting that the modification can occur in a progressive manner. We hypothesize that the DNA modification results in nonfunctional elements.The Mutator system of maize (Zea mays) We describe here the initial characterization of a Mutatorinduced mutation of bronze 2 (bz2), a gene encoding one of the last steps in the anthocyanin pathway (10). Plants have been identified that have a decreased transmission of the mutant variegated phenotype, and a molecular characterization of the Mu elements within these lines suggests that the elements are inactivated by extensive DNA modification. as indicated in figure legends. Unincorporated nucleotides were removed from nick-translated probes (18) by spin dialysis (19) through Bio-Gel P-10 (Bio-Rad) equilibrated in 10 mM Tris HCl, pH 7.5/1 mM EDTA. Maize DNA fragments for nick-translation were prepared by digestion with restriction enzymes, separated from the plasmid vector DNA by electrophoresis through low-melting agarose (FMC Seaplaque), extracted with phenol, and precipitated with ethanol. Hybridizations were in 50% (vol/vol) formamide (EM Science)/450 mM sodium chloride/45 mM sodium citrate/0.05% Ficoll 400 (Pharmacia)/0.05% polyvinylpyrrolidone/0.05% bovine serum albumin, containing salmon sperm DNA (Sigma) at 100 Ag/ml at 420C for 36-48 hr. Filters were washed in 15 mM sodium chloride/1.5 mM sodium citrate/0.1% NaDodSO4 at 650C (Mul and Adhl probes) or 500C (ars probe) for 3 hr with three buffer changes. The filters Abbreviations: Mu, members of a transposable element family in Mutator lines; ars, autonomously replicating sequence; kb, kilobase(s); bp, base pair(s); APT-paper, aminophenylthioether paper. MATERIALS AND METHODS

Molecular mechanisms in the developmental regulation of the maize Suppressor-mutator transposable element.

“…However, partially methylated Ac elements appear to retain their genetic activity (Schwartz and Dennis 1986). A correlation has also been reported between the extent of methylation and the mobility of Mu element sequences in maize (Bennetzen 1985;Chandler and Walbot 1986;Walbot 1988). However, in the case of the Mu elements, there is insufficient genetic information to permit a distinction to be made between effects on expression of a mobilizing element and on the mobility of a transposition-defective element.…”

“…The underlying genetic mechanism is heritable, reversible, and capable of promoting the incremental transition of the element from a stably active to a stably inactive (cryptic) form, implying the involvement of multiple epigenetic events (Fedoroff and Banks 1988). In view of the accumulating evidence that the inactivation of gene expression, in general (for reviews, see Razin et al 1984;Adams and Burdon 1985;Holliday 1987;Cedar 1988), and maize transposable elements, in particular Bennetzen 1985;Chandler and Walbot 1986;Schwartz and Dennis 1986;Chomet et al 1987;Fedoroff et al 1988a, b), are associated with DNA modification, we have investigated the relationship between Spm expression and the methylation pattern of sequences within and flanking an Spin element inserted at the maize a locus.…”

Expression and Post-Transcriptional Regulation of Maize Transposable Element MuDR and Its Derivatives

“…Epigenetic silencing of Mutator activity negatively affects MuDR transcription and is accompanied by the coordinate DNA methylation in TIRs of MuDR and all other Mu elements (Chandler and Walbot, 1986;Martienssen and Baron, 1994). Silencing initiates when a transcriptionally active MuDR element(s) is present, but maintenance of TIR methylation persists even if MuDR is segregated out of the stock (Lisch et al, 1995).…”

“…These observations suggest that MURA is present and competent in DNA binding early in development. Furthermore, methylation accompanying the epigenetic silencing of Mu and MuDR elements (Chandler and Walbot, 1986;Martienssen and Baron, 1994) is maintained, despite the presence of hMURA proteins in both inbred and inactive Mutator lines. This evidence indicates that homolog-encoded MURA proteins are unable to interact with the TIRs in a manner that results in demethylation.…”

Are imprinting and inbreeding two related phenomena?

“…Such modification may be one mechanism regulating the possible deleterious activity in the cell (Chandler and Walbot, 1986). Transposition rate may also be under genetic control as in the switch in mating type of yeasts, which is normally confined to the mother cell lineage (Hicks et al, 1977).…”