Title: Accidental Amplification and Inactivation of a Methyltransferase Gene Eliminates Cytosine Methylation in Mycosphaerella Graminicola Authors
Submitted to: Genetics
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: July 8, 2010
Publication Date: September 10, 2010
Citation: Dhillon, B., Cavaletto, J.R., Wood, K.V., Goodwin, S.B. 2010. Accidental Amplification and Inactivation of a Methyltransferase Gene Eliminates Cytosine Methylation in Mycosphaerella Graminicola. Genetics. 186:67-77. Interpretive Summary: Repetitive DNA occurs in the genomes of almost all organisms analyzed so far, yet its origin and function usually are not known. Transposable elements (TEs) are a subset of the repetitive DNA that are capable of self replication and movement, and in plants are known occasionally to move parts of gene sequences. To test for this phenomenon in fungi, the genome sequence of Mycosphaerella graminicola, the septoria tritici blotch pathogen of wheat, was scanned for repetitive elements. The analysis identified a gene for DNA modification that occurred in many copies and seemed to be part of a repetitive element. However, this gene was unusual and there were no other characteristics of known TEs. All copies except one occurred at the end of a chromosome, and all appeared to have been inactivated by a process known as Repeat-Induced Point Mutation (RIP). Testing of the DNA confirmed that the gene was inactivated; the predicted modification was missing in M. graminicola, but was normal in the related species M. fijiensis from banana and two very closely related, undescribed species from wild grasses in Iran that are thought to have diverged from M. graminicola within the past 11,000 years. Accidental amplification through replication of chromosome ends and inactivation of a usually single-copy gene by RIP appear to form a novel mechanism of gene inactivation in fungi, which seems to have occurred in M. graminicola within the past 11,000 years, concomitant with the domestication of its wheat host as a crop. This information will be useful to evolutionary biologists studying the evolution of fungal genomes and to fungal geneticists analyzing mechanisms of gene regulation and inactivation.
Technical Abstract: A de novo search for repetitive elements in the genome sequence of the wheat pathogen Mycosphaerella graminicola identified a family of repeats containing a DNA methyltransferase sequence (MgDNMT), which is a homologue of the Neurospora crassa Dim-2 gene. A total of 28 MgDNMT sequences was identified, all of which carried signatures of Repeat Induced Point mutations (RIP). All copies of MgDNMT were subtelomeric in location, except for one on chromosome 6. Synteny of the non-telomeric MgDNMT region to the genome of a closely related fungus, M. fijiensis, implied that the copy on chromosome 6 was the original sequence that was amplified. Southern analysis revealed that the MgDNMT sequence also was multicopy in 15 other M. graminicola isolates from various geographical regions. In contrast, an in silico search for MgDNMT homologues identified a single, unRIPed copy in eight ascomycete genomes, suggesting that the amplification event might be specific to M. graminicola. A genome-wide methylation assay revealed that M. graminicola lacks cytosine methylation, as expected if its MgDNMT gene is inactivated. However, methylation appeared to be normal in several other species tested, including the two undescribed sibling species S1 and S2, the closest known relatives of M. graminicola. Since S1 is thought to have diverged from M. graminicola within the past 11,000 years, we suggest that a recent amplification of the single-copy MgDNMT gene made it susceptible to RIP, resulting in complete loss of cytosine methylation in M. graminicola. This change may have occurred concomitant with the domestication of wheat as a crop. This is the first report of accidental amplification followed by RIP as a novel mechanism for inactivation of single-copy genes in fungi.