Dataset Information

DNA methylation associated with repeat-induced point mutation in Neurospora crassa.

ABSTRACT: Repeat-induced point mutation (RIP) is a process that efficiently detects DNA duplications prior to meiosis in Neurospora crassa and peppers them with G:C to A:T mutations. Cytosine methylation is typically associated with sequences affected by RIP, and methylated cytosines are not limited to CpG dinucleotides. We generated and characterized a collection of methylated and unmethylated amRIP alleles to investigate the connection(s) between DNA methylation and mutations by RIP. Alleles of am harboring 84 to 158 mutations in the 2.6-kb region that was duplicated were heavily methylated and triggered de novo methylation when reintroduced into vegetative N. crassa cells. Alleles containing 45 and 56 mutations were methylated in the strains originally isolated but did not become methylated when reintroduced into vegetative cells. This provides the first evidence for de novo methylation in the sexual cycle and for a maintenance methylation system in Neurospora cells. No methylation was detected in am alleles containing 8 and 21 mutations. All mutations in the eight primary alleles studied were either G to A or C to T, with respect to the coding strand of the am gene, suggesting that RIP results in only one type of mutation. We consider possibilities for how DNA methylation is triggered by some sequences altered by RIP.

SUBMITTER: Singer MJ

PROVIDER: S-EPMC230809 | biostudies-other | 1995 Oct

REPOSITORIES: biostudies-other

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DNA methylation associated with repeat-induced point mutation in Neurospora crassa.

Singer M J MJ Marcotte B A BA Selker E U EU

Molecular and cellular biology 19951001 10

Repeat-induced point mutation (RIP) is a process that efficiently detects DNA duplications prior to meiosis in Neurospora crassa and peppers them with G:C to A:T mutations. Cytosine methylation is typically associated with sequences affected by RIP, and methylated cytosines are not limited to CpG dinucleotides. We generated and characterized a collection of methylated and unmethylated amRIP alleles to investigate the connection(s) between DNA methylation and mutations by RIP. Alleles of am harbo ...[more]

PMID: 7565710

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Project description:Numerous cytogenetic observations have shown that homologous chromosomes (or individual chromosomal loci) can engage in specific pairing interactions in the apparent absence of DNA breakage and recombination, suggesting that canonical recombination-mediated mechanisms may not be the only option for sensing DNA/DNA homology. One proposed mechanism for such recombination-independent homology recognition involves direct contacts between intact double-stranded DNA molecules. The strongest in vivo evidence for the existence of such a mechanism is provided by the phenomena of homology-directed DNA modifications in fungi, known as repeat-induced point mutation (RIP, discovered in Neurospora crassa) and methylation-induced premeiotically (MIP, discovered in Ascobolus immersus). In principle, Neurospora RIP can detect the presence of gene-sized DNA duplications irrespectively of their origin, underlying nucleotide sequence, coding capacity or relative, as well as absolute positions in the genome. Once detected, both sequence copies are altered by numerous cytosine-to-thymine (C-to-T) mutations that extend specifically over the duplicated region. We have recently shown that Neurospora RIP does not require MEI-3, the only RecA/Rad51 protein in this organism, consistent with a recombination-independent mechanism. Using an ultra-sensitive assay for RIP mutation, we have defined additional features of this process. We have shown that RIP can detect short islands of homology of only three base-pairs as long as many such islands are arrayed with a periodicity of 11 or 12 base-pairs along a pair of DNA molecules. While the presence of perfect homology is advantageous, it is not required: chromosomal segments with overall sequence identity of only 35-36 % can still be recognized by RIP. Importantly, in order for this process to work efficiently, participating DNA molecules must be able to co-align along their lengths. Based on these findings, we have proposed a model, in which sequence homology is detected by direct interactions between slightly-extended double-stranded DNAs. As a next step, it will be important to determine if the uncovered principles also apply to other processes that involve recombination-independent interactions between homologous chromosomal loci in vivo as well as to protein-free DNA/DNA interactions that were recently observed under biologically relevant conditions in vitro.

Dataset Information

DNA methylation associated with repeat-induced point mutation in Neurospora crassa.

Publications

DNA methylation associated with repeat-induced point mutation in Neurospora crassa.

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