Evidence that spontaneous mitotic recombination occurs at the two-strand stage.

Abstract

ABSTRACr Spontaneous reciprocal mitotic recombination in the yeast Saccharomyces cerevisiae, associated with heteroallelic recombination, occurs almost exclusively at the twostrand stage and involves recombination of unduplicated chromosomes (i.e., during GI) or the unduplicated regions of chromosomes during the S phase of mitosis. The associated heteroallelic recombination frequently reflects the formation of symmetric Holliday structures, is not strongly polarized with respect to conversion at the heteroallelic trp5 sites studied, occasionally results in simultaneous conversion of widely separated genetic markers, and is positively correlated with recombination of flanking markers.The recent compelling evidence that genetic recombination involves the formation of Holliday structures (1) prompted a reexamination of the classical G2 model (Fig. 1) (2) of mitotic recombination. Fig. 2 illustrates that sectored colonies can result from Holliday structures formed at the two-strand stage (e.g., Gi) when the Holliday structure is not cleaved and is replicated during the subsequent round of DNA synthesis (S phase). This mechanism, like that of exchange at the four-strand stage, generates the chromosomal genotypes resulting in daughter cells homozygous for markers distal to the site of exchange.The purpose of the experiment described below was to determine whether spontaneous mitotic recombination in yeast occurs at the four-strand stage as in Fig. 1 or at the two-strand stage as in Fig. 2. The experimental data lead to the conclusion that the spontaneous mitotic recombination observed is due primarily to recombination between either unreplicated homologous chromosomes (during Gl) or unreplicated regions of homologous chromosomes (during S) and therefore occurs at the two-strand stage for the chromosomal regions undergoing exchange. Media and Genetic Techniques. The compositions of the glucose nutrient medium (YPD), synthetic media, and sporulation medium and the relevant genetic techniques have been reported (4). MATERIALS AND METHODSMeasurement of Recombination Rates. Liquid YPD cultures of OR1, each consisting of 6 ml, were inoculated at ca 103 cells + buds per ml, grown aerobically at 30 , harvested during the exponential phase of growth (ca 2 X 107 cells + buds per ml), washed twice with sterile distilled water, and plated on complete synthetic medium and tryptophanless synthetic medium. The recombination rates resulting in tryptophanindependent prototrophic colonies were calculated by the method of the median as described by Lea and Coulson (5). 4436The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U. S. C. §1734 solely to indicate this fact.

Recombination in diploid vegetative cells of Saccharomyces cerevisiae

“…The cells that were used in this study were in stationary phase, namely, in GI. EseOmTO (2) and FABRE (3) have demonstrated conclusively Table III. The classes of prototrophs induced by x-rays in 2-allele combinations. The genotypes are given as noncrossovers (nco) and crossovers (co) in regions I, II, III, or a combination of these.…”

Homologous Recombination Conserves DNA Sequence Integrity Throughout the Cell Cycle in Embryonic Stem Cells

“…Thus, patterns of LOH might be directly related to organismal development and cell differentiation. An intriguing point of possible similarity to yeast is that 70% of yeast MR initiates in G1 [60,61].…”

Yeast Mph1 helicase dissociates Rad51-made D-loops: implications for crossover control in mitotic recombination

“…Crossovers are potentially dangerous, and occur much less frequently in mitotic than meiotic cells (Esposito 1978;Nassif et al 1994;Johnson and Jasin 2000;Virgin et al 2001;Stark and Jasin 2003). Specifically, crossovers may lead to loss of heterozygosity (Beumer et al 1998) and can result in chromosomal aberrations such as translocations, inversions, or deletions.…”

“…In meiotic S. cerevisiae cells the correct number of crossovers are required for proper chromosome segregation; the proportion of gene conversion accompanied by crossing over varies between 25% and 50% depending on the locus (Roeder 1995). In mitotic cells, the proportion of crossovers is much lower, ranging between <1% and 15%, depending on the recombination assay and organism (Esposito 1978;Nassif et al 1994;Johnson and Jasin 2000;Virgin et al 2001;Stark and Jasin 2003). These differences are best explained if recombination proceeds most often according to the dHJ model in meiotic cells and via SDSA and other noncrossover means in mitotic cells.…”