Characterization of the Neurospora crassa mus-25 mutant: the gene encodes a protein which is homologous to the Saccharomyces cerevisiae Rad54 protein

Handa, Nobuhiko; Noguchi, Yoko; Sakuraba, Yoshiyuki; Ballario, Paola; Macino, Giuseppe; Fujimoto, Naomi; Ishii, Chizu; Inoue, Hirokazu

doi:10.1007/s004380000303

Cited by 17 publications

(16 citation statements)

References 43 publications

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“…The rate of homologous integration of plasmid pMTR (12) into the chromosomal mtr locus was assayed in the wild type and in mutants defective in these HR repair genes (11). In the wild type, 3-5% of transformants showed homologous integration, whereas Ͻ0.3% of transformants did so in mei-3 and mus-25 mutants.…”

Section: Highly Efficient Gene Replacements In Neurospora Strains Defmentioning

confidence: 99%

Highly efficient gene replacements in Neurospora strains deficient for nonhomologous end-joining

Ninomiya

Suzuki

Ishii

et al. 2004

Proc. Natl. Acad. Sci. U.S.A.

541

453

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Gene disruption and overexpression play central roles in the analysis of gene function. Homologous recombination is, in principle, the most efficient method of disrupting, modifying, or replacing a target gene. Although homologous integration of exogenous DNA into the genome occurs readily in Saccharomyces cerevisiae , it is rare in many other organisms. We identified and disrupted Neurospora crassa genes homologous to human KU70 and KU80 , which encode proteins that function in nonhomologous end-joining of double-stranded DNA breaks. The resulting mutants, named mus - 51 and mus - 52 , showed higher sensitivity to methyl methanesulfonate, ethyl methanesulfonate, and bleomycin than wild type, but not to UV, 4-nitroquinoline 1-oxide, camptothecin, or hydroxyurea. Vegetative growth, conidiation, and ascospore production in homozygous crosses were normal. The frequency of integration of exogenous DNA into homologous sequences of the genome in the KU disruption strains of N. crassa was compared with that in wild type, mei-3 , and mus - 11 . In mei-3 and mus - 11 , which are defective in homologous recombination, none or few homologous integration events were observed under any conditions. When mtr target DNA with ≈2-kb 5′ and 3′ flanking regions was used for transformation of the KU disruption strains, 100% of transformants exhibited integration at the homologous site, compared to 10 to 30% for a wild-type recipient. Similar results were obtained when the ad-3A gene was targeted for disruption. These results indicate that KU disruption strains are efficient recipients for gene targeting.

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Section: Highly Efficient Gene Replacements In Neurospora Strains Defmentioning

confidence: 99%

Highly efficient gene replacements in Neurospora strains deficient for nonhomologous end-joining

Ninomiya

Suzuki

Ishii

et al. 2004

Proc. Natl. Acad. Sci. U.S.A.

541

453

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“…In N. crassa, MUS-25 and at least three other SNF2-family proteins have been implicated in homologous recombination (Handa et al 2000;Zhang et al 2013). These are swr1 (NCU09993), chd1 (NCU03060), and isw1 (NCU03875), two of which are included in Figure 3 because of their similarity to SAD-6.…”

Section: Discussionmentioning

confidence: 99%

“…have roles in vegetative and sexual phases of the N. crassa life cycle (Handa et al 2000;Zhang et al 2013). Overall, sad-6's expression pattern suggests that it has roles in both vegetative and sexual processes.…”

Section: Sad-6 Transcript Analysis Suggests That It Has Functions Outmentioning

confidence: 94%

“…The N. crassa ortholog of S. cerevisiae Rad54 is MUS-25 (Handa et al 2000;Zhang et al 2013). MUS-25 is the only other SNF2-family protein in N. crassa predicted to be a member of the Rad54-like group (Flaus et al 2006), and it has been shown to be important for quelling, a vegetative RNA silencing process (Chang et al 2012;Zhang et al 2013).…”

Section: Deletion Of Ncu06190 Suppresses Msudmentioning

confidence: 99%

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Efficient Detection of Unpaired DNA Requires a Member of the Rad54-Like Family of Homologous Recombination Proteins

et al. 2014

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Meiotic silencing by unpaired DNA (MSUD) is a process that detects unpaired regions between homologous chromosomes and silences them for the duration of sexual development. While the phenomenon of MSUD is well recognized, the process that detects unpaired DNA is poorly understood. In this report, we provide two lines of evidence linking unpaired DNA detection to a physical search for DNA homology. First, we have found that a putative SNF2-family protein (SAD-6) is required for efficient MSUD in Neurospora crassa. SAD-6 is closely related to Rad54, a protein known to facilitate key steps in the repair of double-strand breaks by homologous recombination. Second, we have successfully masked unpaired DNA by placing identical transgenes at slightly different locations on homologous chromosomes. This masking falls apart when the distance between the transgenes is increased. We propose a model where unpaired DNA detection during MSUD is achieved through a spatially constrained search for DNA homology. The identity of SAD-6 as a Rad54 paralog suggests that this process may be similar to the searching mechanism used during homologous recombination. MEIOSIS is fundamental to sexual reproduction. During meiosis, chromosomes are replicated, aligned, recombined, and segregated to nuclei that will develop into gametes. Two of these key processes, alignment and recombination, likely require a search for DNA homology between chromosomes (Barzel and Kupiec 2008;Moore and Shaw 2009). Such homology searching is necessary because sexual organisms inherit a copy of each chromosome from each of its parents. These chromosomes, referred to as homologs, must somehow find each other so that alignment, recombination, and segregation can occur.Although recent research has improved our understanding of homology search mechanisms (Forget and Kowalczykowski 2012;Renkawitz et al. 2013), there are many questions that remain unanswered. The filamentous fungus Neurospora crassa may be useful for investigating the unknowns of homology searching because it possesses a genetically tractable phenomenon called meiotic silencing by unpaired DNA (MSUD) (Aramayo and Selker 2013;Billmyre et al. 2013). MSUD scans pairs of homologs for segments of DNA that are not accurately paired between them. If improper pairing (i.e., unpairing) is identified, the offending sequences are silenced for the duration of sexual development. For example, if a hypothetical gene called "gene A" is on the left arm of one chromosome but on the right arm of its homolog, it will be silenced. The same holds true if gene A has been lost from one of the homologs.A functional MSUD response can be easily detected with alleles that affect ascospore (sexual spore) color or shape. Indeed, MSUD was discovered during studies of ascospore maturation-1 (asm-1), a gene required for the production of pigmented (black) ascospores . A cross between an asm-1 + strain and an asm-1 D strain produces mostly unpigmented (white) ascospores. This is because MSUD silences the unpaired asm-1 + all...

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“…Epistasis analyses involving mus-10 suggested that it belonged to the uvs-6 epistasis group, which functions in recombination repair (Kafer and Perlmutter 1980;Kafer 1983). However, mus-10 did not display several phenotypes common to other members of the uvs-6 epistasis group: chromosomal instability, a high sensitivity to histidine, and the inability to produce viable ascospores in homozygous crosses Kafer and Perlmutter 1980;Kafer 1981;Schroeder 1986;Watanabe et al 1997;Handa et al 2000;Sakuraba et al 2000). Furthermore, the frequencies of spontaneous and radiation-induced mutation observed in mus-10 were similar to those of a wild-type strain (Kafer 1981).…”

mentioning

confidence: 87%

Deletion of a Novel F-Box Protein, MUS-10, in Neurospora crassa Leads to Altered Mitochondrial Morphology, Instability of mtDNA and Senescence

Kurashima¹,

Chae²,

Sawada³

et al. 2010

Genetics

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While mitochondria are renowned for their role in energy production, they also perform several other integral functions within the cell. Thus, it is not surprising that mitochondrial dysfunction can negatively impact cell viability. Although mitochondria have received an increasing amount of attention in recent years, there is still relatively little information about how proper maintenance of mitochondria and its genomes is achieved. The Neurospora crassa mus-10 mutant was first identified through its increased sensitivity to methyl methanesulfonate (MMS) and was thus believed to be defective in some aspect of DNA repair. Here, we report that mus-10 harbors fragmented mitochondria and that it accumulates deletions in its mitochondrial DNA (mtDNA), suggesting that the mus-10 gene product is involved in mitochondrial maintenance. Interestingly, mus-10 begins to senesce shortly after deletions are visualized in its mtDNA. To uncover the function of MUS-

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Characterization of the Neurospora crassa mus-25 mutant: the gene encodes a protein which is homologous to the Saccharomyces cerevisiae Rad54 protein

Cited by 17 publications

References 43 publications

Highly efficient gene replacements in Neurospora strains deficient for nonhomologous end-joining

Highly efficient gene replacements in Neurospora strains deficient for nonhomologous end-joining

Efficient Detection of Unpaired DNA Requires a Member of the Rad54-Like Family of Homologous Recombination Proteins

Deletion of a Novel F-Box Protein, MUS-10, in Neurospora crassa Leads to Altered Mitochondrial Morphology, Instability of mtDNA and Senescence

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