Isolation of the RAD18 gene of Saccharomyces cerevisiae and construction of rad18 deletion mutants

Fabre, Francis; Magaña‐Schwencke, Nieve; Chanet, Roland

doi:10.1007/bf00427039

Cited by 20 publications

(13 citation statements)

References 32 publications

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“…RAD18 is expressed in multiple tissues, but the highest level is found in testis (van der Laan et al, 2000), and in S. cerevisiae, RAD18 gene expression increases during meiosis, suggesting a specific function for RAD18 in this process (Jones and Prakash, 1991). Deletion of RAD18 in yeast does not affect either meiosis or meiotic recombination (Dowling et al, 1985; Fabre et al, 1989; Game and Mortimer, 1974) but double mutants of rad18 and various genes that act during excision–repair show a drastic reduction in spore viability, compared with the single mutants (Dowling et al, 1985). This result indicates that Rad18 does perform a specific function during meiosis in S. cerevisiae .…”

Section: Introductionmentioning

confidence: 99%

Meiotic functions of RAD18

Inagaki

Sleddens-Linkels

Wassenaar

et al. 2011

Journal of Cell Science

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RAD18 is an ubiquitin ligase that is involved in replication damage bypass and DNA double-strand break (DSB) repair processes in mitotic cells. Here, we investigated the testicular phenotype of Rad18-knockdown mice to determine the function of RAD18 in meiosis, and in particular, in the repair of meiotic DSBs induced by the meiosis-specific topoisomerase-like enzyme SPO11. We found that RAD18 is recruited to a specific subfraction of persistent meiotic DSBs. In addition, RAD18 is recruited to the chromatin of the XY chromosome pair, which forms the transcriptionally silent XY body. At the XY body, RAD18 mediates the chromatin association of its interaction partners, the ubiquitin-conjugating enzymes HR6A and HR6B. Moreover, RAD18 was found to regulate the level of dimethylation of histone H3 at Lys4 and maintain meiotic sex chromosome inactivation, in a manner similar to that previously observed for HR6B. Finally, we show that RAD18 and HR6B have a role in the efficient repair of a small subset of meiotic DSBs.

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Section: Introductionmentioning

confidence: 99%

Meiotic functions of RAD18

Inagaki

Sleddens-Linkels

Wassenaar

et al. 2011

Journal of Cell Science

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“…The RAD18::ADE2 plasmid (pCB432) was made as follows. The 2.3-kb ClaI-XbaI fragment containing sequences in and upstream of RAD18 from pJH830 (Fabre et al 1989) was subcloned into Bluescript SK(+) (Stratagene) to generate pCB430. A 2-kb BglII fragment containing ADE2 from pASZ11 (Stotz and Linder 1990) was inserted into the BglII sites of pSL301 (Invitrogen) to generate pCB429.…”

Section: Plasmidsmentioning

confidence: 99%

Tam1, a telomere-associated meiotic protein, functions in chromosome synapsis and crossover interference.

Chua¹,

Roeder²

1997

Genes Dev.

159

198

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The TAM1 gene of Saccharomyces cerevisiae is expressed specifically during meiosis and encodes a protein that localizes to the ends of meiotic chromosomes. In a taml null mutant, there is an increase in the frequency of chromosomes that fail to recombine and an associated increase in homolog nondisjunction at meiosis I. The taml mutant also displays an increased frequency of precocious separation of sister chromatids and a reduced efficiency of distributive disjunction. The defect in distributive disjunction may be attributable to overloading of the distributive system by the increased number of nonrecombinant chromosomes. Recombination is not impaired in the taml mutant, but crossover interference is reduced substantially. In addition, chromosome synapsis is delayed in taml strains. The combination of a defect in synapsis and a reduction in interference is consistent with previous studies suggesting a role for the synaptonemal complex in regulating crossover distribution, taml is the only known yeast mutant in which the control of crossover distribution is impaired, but the frequency of crossing over is unaffected. We discuss here possibilities for how a telomere-associated protein might function in chromosome synapsis and crossover interference. During meiosis, a single round of DNA duplication is followed by two successive nuclear divisions to generate four haploid progeny from a single diploid cell. The meiosis II division resembles mitotic chromosome segregation , but the meiosis I division is unique in that homologous chromosomes disjoin from each other. A complex series of events unique to prophase of meiosis I ensures that homologs segregate reductionally at the first division. One important aspect of meiotic prophase is a high rate of recombination between homologous chromosomes. Crossing over establishes chiasmata, which are physical connections between homologous chromosomes that persist until metaphase. Chiasmata ensure the proper orientation of chromosomes on the meiosis I spindle and therefore promote reductional chromosome segregation (for review, see Carpenter 1994). The distribution of crossovers, and the chiasmata to which they give rise, is nonrandom in two respects. First, two cross-overs rarely occur closely together-a phenomenon known as crossover interference. Second, every chromosome pair (no matter how small) almost always sustains at least one crossover-referred to as obligate chiasma. A number of studies suggest that crossover interference and obligate chiasma are different manifestations of the 1Corresponding author. E-MAIL: shirleen.roeder@yale.edu; FAX (203) 432-3263. same underlying mechanism (Jones 1967; Kaback et al.

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“…Genetic studies of Saccharomyces cerevisiae suggest that all known components of TLS belong to the epistasis group of the RAD6-RAD18 genes (21,22), which are conserved from yeast to mammalian cells (23)(24)(25)(26)(27). The Rad6 protein is one of the ubiquitin-conjugating enzymes (E2s) and forms a tight complex with the Rad18 protein, which may have E3 ubiquitin ligase activity (28,29 lymphocyte line DT40 (30).…”

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confidence: 99%

Involvement of Vertebrate Polκ in Rad18-independent Postreplication Repair of UV Damage

Okada¹,

Sonoda²,

Yamashita³

et al. 2002

Journal of Biological Chemistry

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DNA damage, which is left unrepaired by excision repair pathways, often blocks replication, leading to lesions such as breaks and gaps on the sister chromatids. These lesions may be processed by either homologous recombination (HR) repair or translesion DNA synthesis (TLS). Vertebrate Pol belongs to the DNA polymerase Y family, as do most TLS polymerases. However, the role for Pol in vertebrate cells is unclear because of the lack of reverse genetic studies. Here, we generated cells deficient in Pol (pol cells) from the chicken B lymphocyte line DT40. Although purified Pol is unable to bypass ultraviolet (UV) damage, pol cells exhibited increased UV sensitivity, and the phenotype was suppressed by expression of human and chicken Pol, suggesting that Pol is involved in TLS of UV photoproduct. Defects in both Pol and Rad18, which regulates TLS in yeast, in DT40 showed an additive effect on UV sensitivity. Interestingly, the level of sister chromatid exchange, which reflects HR-mediated repair, was elevated in normally cycling pol cells. This implies functional redundancy between HR and Pol in maintaining chromosomal DNA. In conclusion, vertebrate Pol is involved in Rad18-independent TLS of UV damage and plays a role in maintaining genomic stability.A wide range of potential insult to the genomic DNA is contributed not only by the environment, but also by cellular activities. DNA damages, which are left unrepaired by excision repair pathways, may arrest DNA replication, leading to breaks and gaps on the sister chromatids. These lesions are processed by two major postreplication repair pathways: homologous recombination (HR) 1 repair and translesion DNA synthesis (TLS) (1). TLS fills a daughter strand gap that encompasses a DNA damage on the template strand by employing a number of specialized DNA polymerases (Pols) (reviewed in Refs. 2-4). On the other hand, HR functions in processing both gaps and breaks by facilitating recombination between damaged sister chromatids with the other intact ones. Thus, daughter-strand gaps can be processed by both TLS and HR in postreplicational repair.Human cells contain four Y family DNA polymerases, namely Pol (Rad30A), Pol (Rad30B), Pol (DinB1), and Rev1 (5-7). A number of biochemical studies have demonstrated that Y family DNA polymerases can bypass specific lesions on the template strand (8 -12), whereas in vivo function of these polymerases are as yet unclear except for Pol. Pol is mutated in a variant form of xeroderma pigmentosum (XP-V) (13, 14), which is characterized by predisposition to skin cancer and elevated UV sensitivity.

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Isolation of the RAD18 gene of Saccharomyces cerevisiae and construction of rad18 deletion mutants

Cited by 20 publications

References 32 publications

Meiotic functions of RAD18

Meiotic functions of RAD18

Tam1, a telomere-associated meiotic protein, functions in chromosome synapsis and crossover interference.

Involvement of Vertebrate Polκ in Rad18-independent Postreplication Repair of UV Damage

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