Extensive interallelic polymorphisms drive meiotic recombination into a crossover pathway

Abstract

Recombinants isolated from most meiotic intragenic recombination experiments in maize, but not in yeast, are borne principally on crossover chromosomes. This excess of crossovers is not explained readily by the canonical double-strand break repair model of recombination, proposed to account for a large body of yeast data, which predicts that crossovers (COs) and noncrossovers (NCOs) should be recovered equally. An attempt has been made here to identify general rules governing the recovery of the CO and NCO classes of intragenic recombinants in maize. Recombination was analyzed in bz heterozygotes between a variety of mutations derived from the same or different progenitor alleles. The mutations include point mutations, transposon insertions, and transposon excision footprints. Consequently, the differences between the bz heteroalleles ranged from just two nucleotides to many nucleotides, indels, and insertions. In this article, allelic pairs differing at only two positions are referred to as dimorphic to distinguish them from polymorphic pairs, which differ at multiple positions. The present study has revealed the following effects at these bz heteroalleles: (1) recombination between polymorphic heteroalleles produces mostly CO chromosomes; (2) recombination between dimorphic heteroalleles produces both CO and NCO chromosomes, in ratios apparently dependent on the nature of the heteroalleles; and (3) in dimorphic heterozygotes, the two NCO classes are recovered in approximately equal numbers when the two mutations are point mutations but not when one or both mutations are insertions. These observations are discussed in light of a recent version of the double-strand break repair model of recombination that postulates separate pathways for the formation of CO and NCO products.

Figure 1.

Locations of Mutations and Heterologies in Different bz Alleles. The locations of the mutations and heterologies in all of the bz alleles used in this study are shown in a composite map of the two progenitor wild-type alleles, Bz-W22 and Bz-McC. Mutations derived from Bz-W22 are shown above the line, and those derived from Bz-McC are shown below the line. Numbers refer to the published Bz-W22 genomic DNA sequence, with 1 corresponding to the translation start site (Ralston et al., 1988). The designations E2 to E9 and EMc1 refer to the mutations bz-E2 to bz-E9 and bz-EMc1 induced by EMS in the Bz-W22 and Bz-McC progenitor alleles, respectively. The nucleotide difference between a bz-E mutation and the contrasting wild-type site is shown immediately below (E2 to E9) or above (EMc1) each mutation. The Ds insertion mutations bz-m1 and bz-m2(DI) arose in the Bz-McC progenitor allele and are represented by triangles. The number inside each triangle refers to the size of the insertion in kilobases. The location of the 8-bp direct repeat generated by the insertion is indicated in the box above each insertion. The locations of the Ac excision footprint in the bz-s30, bz-s2.1, and bz-s2.2 mutations are indicated by similar boxes. The nucleotide polymorphisms between Bz-W22 and Bz-McC are shown, respectively, above and below the lines representing the two alleles. The single intron (105 bp in Bz-W22 and 100 bp in Bz-McC) is shown by stippling. Oligonucleotides used as primers in polymerase chain reaction and sequencing are identified by arrows that indicate their 5′ to 3′ strand polarity.

Figure 2.

Modified DSBR Model of Recombination with Separate Pathways for the Formation of CO and NCO Products. The diagram shows the two interacting DNA duplexes (chromatids) as red and blue double strands. A double-strand break (DSB) formed in the blue duplex is resected by a 5′ to 3′ exonuclease to generate long 3′ single-strand overhangs. One single strand invades the homologous duplex, forming a heteroduplex and displacing a D loop, which is extended by DNA synthesis. The resulting D loop has two possible fates. (A) In the DSBR pathway, the D loop anneals to the second 3′ end overhang, serving as a template for additional DNA synthesis. Ligation of the newly synthesized strands to the recessed ends leads to the formation of a DHJ with heteroduplex DNA flanking the double-strand break site. This recombination intermediate can be resolved by cutting the outside strands or the inside strands of each junction. The resolution always involves opposite sense cutting (two outside strands and two inside strands); only one of the two alternatives is shown here. This modified version of the DSBR pathway (Szostak et al., 1983) would generate only CO products. (B) In the SDSA pathway (Pâques and Haber, 1999), the D loop intermediate is disassembled by displacement of the newly synthesized DNA strand, which then anneals with the second single-strand overhang. Repair of the break is completed by DNA synthesis and ligation. The SDSA pathway generates only NCO products. (Figure adapted from Allers and Lichten, 2001.)