Cooperative interactions between pairs of homologous chromatids during meiosis in Saccharomyces cerevisiae

Abstract

We report a novel instance of negative interference during Saccharomyces cerevisiae meiosis, where Cre-mediated recombination between pairs of allelic loxP sites is more frequent than expected. We suggest that endogenous crossover recombination mediates cooperative pairing interactions between all four chromatids of a meiotic bivalent.

Figure 1.—

Fluorescent tetrad analysis of allelic loxP collisions. (a) A schematic is shown for the Cre-mediated allelic loxP recombination system, where the frequency of reporter activation (“collision”) is proportional to the physical proximity of the two interacting loxP sites (Hildebrandt and Cozzarelli 1995; Burgess and Kleckner 1999). The loxP site has been incorporated into the ACT1 intron to allow for sufficient expression of the GFP protein to visualize the Gfp⁺ phenotype by fluorescence microscopy (plasmid and strain construction details available upon request). Presumably, expression from the GPD1 promoter in spores and the removal of the inverted repeat loxP site from the 5′-UTR of GFP mRNA by splicing allow for robust expression of GFP (K. Komachi and O. Hughes, unpublished data). Two paths are shown for generating 2:2 Gfp⁺ (N) tetrads, with Cre-mediated recombination occurring either before or after meiotic S-phase. (b) Two fields of asci sporulated with 0.03% galactose (which activates expression of Cre recombinase) added at t = 1 hr are shown. On the left, 1:3 Gfp⁺ (T) and 2:2 Gfp⁺ (N) four-spore asci are visible. On the right, a single tetrad showing 1:3 Gfp⁺ (T) segregation is shown at higher magnification.

Figure 2.—

Nonrandom segregation of Gfp⁺ in tetrads. The observed and expected frequencies of (a) 1:3 Gfp⁺ and (b) 2:2 Gfp⁺ are shown (indicating fT and fN, respectively). For expected values, the assumptions of Papazian (1952) and the method described at the Stahl Lab Online Tools were used: and , where and R = fN_obs + fT_obs/2. (a) The observed and expected frequency of 1:3 Gfp⁺ tetrads (fT_obs and fT_exp) for different Cre induction times is illustrated. (b) The observed and expected frequency of 2:2 Gfp⁺ tetrads (fN_obs and fN_exp) is illustrated. Estimated standard error was used to size the error bars.