repeat induced point mutation
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2021 ◽  
Author(s):  
Florian Carlier ◽  
Tinh-Suong Nguyen ◽  
Alexey K. Mazur ◽  
Eugene Gladyshev

ABSTRACTRepeat-induced point mutation (RIP) is a genetic process that creates cytosine-to-thymine (C-to-T) transitions in duplicated genomic sequences in fungi. RIP detects duplications irrespective of their origin, particular sequence, coding capacity, or genomic positions. Previous studies suggested that RIP involves a cardinally new mechanism of sequence recognition that operates on intact double-stranded DNAs. In the fungus Neurospora crassa, RIP can be mediated by a putative C5-cytosine methyltransferase (CMT) RID or/and a canonical CMT DIM-2. These distinct RIP pathways feature opposite substrate preferences: RID-dependent RIP is largely limited to the duplicated sequences, whereas DIM-2-dependent RIP preferentially mutates adjacent non-repetitive regions. Using DIM-2-dependent RIP as a principal readout of repeat recognition, here we show that GC-rich repeats promote stronger RIP compared to AT-rich repeats (independently of their intrinsic propensities to become mutated), with the relative contribution of AT base-pairs being close to zero. We also show that direct repeats promote much more efficient DIM-2-dependent RIP than inverted repeats; both the spacer DNA between the repeat units (the linker) and the flanking regions are similarly affected by this process. These and other results support the idea that repeat recognition for RIP involves formation of many short interspersed quadruplexes between homologous double-stranded DNAs, which need to undergo concomitant changes in their linking number to accommodate pairing.SUMMARYDuring repeat-induced point mutation (RIP) gene-sized duplications of genomic DNA are detected by a mechanism that likely involves direct pairing of homologous double-stranded DNAs. We show that DIM-2-dependent RIP, triggered by closely-positioned duplications, is strongly affected by their relative orientations (direct versus inverted). We also show that GC-rich repeats promote RIP more effectively than AT-rich repeats. These results support a model in which homologous dsDNAs can pair by establishing interspersed quadruplex-based contacts with concomitant changes in their supercoiling status.


2020 ◽  
Vol 2 (3) ◽  
Author(s):  
Wan-Chen Li ◽  
Hou-Cheng Liu ◽  
Ying-Jyun Lin ◽  
Shu-Yun Tung ◽  
Ting-Fang Wang

Abstract Generation of new genetic diversity by crossover (CO) and non-crossover (NCO) is a fundamental process in eukaryotes. Fungi have played critical roles in studying this process because they permit tetrad analysis, which has been used by geneticists for several decades to determine meiotic recombination products. New genetic variations can also be generated in zygotes via illegitimate mutation (IM) and repeat-induced point mutation (RIP). RIP is a genome defense mechanism for preventing harmful expansion of transposable elements or duplicated sequences in filamentous fungi. Although the exact mechanism of RIP is unknown, the C:G to T:A mutations might result from DNA cytosine methylation. A comprehensive approach for understanding the molecular mechanisms underlying these important processes is to perform high-throughput mapping of CO, NCO, RIP and IM in zygotes bearing large numbers of heterozygous variant markers. To this aim, we developed ‘TSETA’, a versatile and user-friendly pipeline that utilizes high-quality and chromosome-level genome sequences involved in a single meiotic event of the industrial workhorse fungus Trichoderma reesei. TSETA not only can be applied to most sexual eukaryotes for genome-wide tetrad analysis, it also outcompetes most currently used methods for calling out single nucleotide polymorphisms between two or more intraspecies strains or isolates.


Author(s):  
Mareike Möller ◽  
Michael Habig ◽  
Cécile Lorrain ◽  
Alice Feurtey ◽  
Janine Haueisen ◽  
...  

AbstractDNA methylation is found throughout all domains of life, yet the extent and function of DNA methylation differ between eukaryotes. Strains of the plant pathogenic fungus Zymoseptoria tritici appeared to lack cytosine DNA methylation (5mC) because gene amplification followed by Repeat-Induced Point mutation (RIP) resulted in the inactivation of the dim2 DNA methyltransferase gene. 5mC is, however, present in closely related sister species. We demonstrate that inactivation of dim2 occurred recently as some Z. tritici isolates carry a functional dim2 gene. Moreover, we show that dim2 inactivation occurred by a different path than previously hypothesized. We mapped the genome-wide distribution of 5mC in strains with and without functional dim2. Presence of functional dim2 correlates with high levels of 5mC in transposable elements (TEs), suggesting a role in genome defense. We identified low levels of 5mC in strains carrying inactive dim2 alleles, suggesting that 5mC is maintained over time, presumably by an active Dnmt5 DNA methyltransferase. Integration of a functional dim2 allele in strains with mutated dim2 restored normal 5mC levels, demonstrating de novo cytosine methylation activity of dim2. To assess the importance of 5mC for genome evolution, we performed an evolution experiment, comparing genomes of strains with high levels of 5mC to genomes of strains lacking dim2. We found that the presence of dim2 alters nucleotide composition by promoting C to T transitions (C→T) specifically at CpA (CA) sites during mitosis, likely contributing to TE inactivation. Our results show that 5mC density at TEs is a polymorphic trait in Z. tritici populations that can impact genome evolution.Author SummaryCytosine DNA methylation (5mC) is known to silence transposable elements in fungi and thereby appears to contribute to genome stability. The genomes of plant pathogenic fungi are highly diverse, differing substantially in transposon content and distribution. Here, we show extensive differences of 5mC levels within a single species of an important wheat pathogen. These differences were caused by inactivation of the DNA methyltransferase Dim2 in the majority of studied isolates. Presence of widespread 5mC increased point mutation rates in regions with active or mutated transposable elements during mitosis. The mutation pattern is dependent on the presence of Dim2 and resembles a mitotic version of Repeat-Induced Point mutation (RIP). Thus, loss of 5mC may represent an evolutionary trade-off offering adaptive potential at the cost of transposon control.


2018 ◽  
Author(s):  
Dev Ashish Giri ◽  
Ajith V. Pankajam ◽  
Koodali T. Nishant ◽  
Durgadas P. Kasbekar

AbstractMeiotic silencing by unpaired DNA (MSUD) was discovered in crosses made in the standard Oak Ridge (OR) genetic background of Neurospora crassa. However, MSUD often was decidedly less efficient when the OR-derived MSUD tester strains were crossed with wild-isolated strains (W), which suggested either that sequence heterozygosity in tester x W crosses suppresses MSUD, or that OR represents the MSUD-conducive extreme in the range of genetic variation in MSUD efficiency. Our results support the latter model. MSUD was much less efficient in near-isogenic crosses made in a novel N. crassa B/S1 and the N. tetrasperma 85 genetic backgrounds. Possibly, regulatory cues that in other genetic backgrounds calibrate the MSUD response are missing from OR. The OR versus B/S1 difference appears to be determined by loci on chromosomes 1, 2, and 5. OR crosses heterozygous for a duplicated chromosome segment (Dp) have for long been known to exhibit an MSUD-dependent barren phenotype. However, inefficient MSUD in N. tetrasperma 85 made Dp-heterozygous crosses non-barren. This is germane to our earlier demonstration that Dps can act as dominant suppressors of repeat-induced point mutation (RIP). Occasionally, during ascospore partitioning rare asci contained >8 nuclei, and round ascospores dispersed less efficiently than spindle-shaped ones.General abstractIn crosses made in the standard OR genetic background of Neurospora crassa, an RNAi-mediated process called MSUD efficiently silences any gene not properly paired with its homologue during meiosis. We found that MSUD was not as efficient in comparable crosses made in the N. crassa B/S1 and N. tetrasperma 85 backgrounds, suggesting that efficient MSUD is not necessarily the norm in Neurospora. Indeed, using OR strains for genetic studies probably fortuitously facilitated the discovery of MSUD and its suppressors. As few as three unlinked loci appear to underlie the OR versus B/S1 difference in MSUD.


2017 ◽  
Author(s):  
Wan-Chen Li ◽  
Chien-Hao Huang ◽  
Chia-Ling Chen ◽  
Yu-Chien Chuang ◽  
Shu-Yun Tung ◽  
...  

AbstractTrichoderma reesei (Ascomycota, Pezizomycotina) QM6a is a model fungus for a broad spectrum of physiological phenomena, including plant cell wall degradation, industrial production of enzymes, light responses, conidiation, sexual development, polyketide biosynthesis and plant-fungal interactions. The genomes of QM6a and its high-enzyme producing mutants have been sequenced by second-generation-sequencing methods and are publicly available from the Joint Genome Institute (JGI). While these genome sequences have offered useful information for genomic and transcriptomic studies, their limitations and especially their short read lengths make them poorly suited for some particular biological problems, including assembly, genome-wide determination of chromosome architecture and genetic modification or engineering. We integrated Pacific Biosciences and Illumina sequencing platforms for the highest-quality genome assembly yet achieved, revealing seven telomere-to-telomere chromosomes (34,922,528 bp; 10877 genes) with 1630 newly-predicted genes and >1.5 Mb of new sequences. Most new sequences are located on AT-rich blocks, including 7 centromeres, 14 subtelomeres and 2329 interspersed AT-rich blocks. The seven QM6a centromeres separately consist of 24 conserved repeats and 37 putative centromere-encoded genes. These findings open up a new perspective for future centromere and chromosome architecture studies. Next, we demonstrate that sexual crossing readily induced cytosine-to-thymine point mutations on both tandem and unlinked duplicated sequences. We also show by bioinformatic analysis that Trichoderma reesei has evolved a robust repeat-induced point mutation (RIP) system to accumulate AT-rich sequences, with longer AT-rich blocks having more RIP mutations. The widespread distribution of AT-rich blocks correlates genome-wide partitions with gene clusters, explaining why clustering of genes has been reported to not influence gene expression in Trichoderma reesei. Compartmentation of ancestral gene clusters by AT-rich blocks might promote flexibilities that are evolutionarily advantageous in this fungus’ soil habitats and other natural environments. Our analyses, together with the complete genome sequence, provide a better blueprint for biotechnological and industrial applications.


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