Rare recombination events generate sequence diversity among balancer chromosomes in Drosophila melanogaster

Abstract

Multiply inverted balancer chromosomes that suppress exchange with their homologs are an essential part of the Drosophila melanogaster genetic toolkit. Despite their widespread use, the organization of balancer chromosomes has not been characterized at the molecular level, and the degree of sequence variation among copies of balancer chromosomes is unknown. To map inversion breakpoints and study potential diversity in descendants of a structurally identical balancer chromosome, we sequenced a panel of laboratory stocks containing the most widely used X chromosome balancer, First Multiple 7 (FM7). We mapped the locations of FM7 breakpoints to precise euchromatic coordinates and identified the flanking sequence of breakpoints in heterochromatic regions. Analysis of SNP variation revealed megabase-scale blocks of sequence divergence among currently used FM7 stocks. We present evidence that this divergence arose through rare double-crossover events that replaced a female-sterile allele of the singed gene (sn X2 ) on FM7c with a sequence from balanced chromosomes. We propose that although double-crossover events are rare in individual crosses, many FM7c chromosomes in the Bloomington Drosophila Stock Center have lost sn X2 by this mechanism on a historical timescale. Finally, we characterize the original allele of the Bar gene (B 1 ) that is carried on FM7, and validate the hypothesis that the origin and subsequent reversion of the B 1 duplication are mediated by unequal exchange. Our results reject a simple nonrecombining, clonal mode for the laboratory evolution of balancer chromosomes and have implications for how balancer chromosomes should be used in the design and interpretation of genetic experiments in Drosophila.B alancer chromosomes are genetically engineered chromosomes that suppress crossing over with their homologs and are used for many purposes in genetics, including construction of complex genotypes, maintenance of stocks, and estimation of mutation rates. Balancers typically carry multiple inversions that suppress genetic exchange or result in the formation of abnormal meiotic products if crossing over does occur (Fig. 1A); for example, single crossovers inside the inverted segment create acentric or dicentric chromosomes that will fail to segregate properly during meiosis or large deletions or duplications that will likely result in inviable gametes (1, 2). Balancers also often carry recessive lethal or sterile mutations to prevent their propagation as homozygotes as well as dominant markers for easy identification. First developed for use in Drosophila melanogaster, balancer chromosomes remain some of the most powerful tools for genetic analysis in this species (3).Despite their widespread use, very little is known about the organization of Drosophila balancer chromosomes at the molecular level. Since their original syntheses decades ago, balancers have undergone many manipulations, including the addition or removal of genetic markers. Moreover, rare recombination events can cause spontaneous loss o...

Genomics of Natural Populations: Evolutionary Forces that Establish and Maintain Gene Arrangements inDrosophila pseudoosbscura

“…We hypothesize that gene conversion and double crossovers can break up associations across an arrangement, but purifying selection removes any maladaptive combinations, consistent with inversion establishment models (Charlesworth & Charlesworth, 1973). These results should be interpreted with caution because some of the shared polymorphism we observe may be due to gene conversion events from balancer chromosomes used to create the isogenic strains (Miller et al, 2016).…”

“…Shared mutations either predate two or more inversion mutations or are alleles transferred among arrangements via gene conversion or double cross overs (Arcadio Navarro, Betrán, Barbadilla, & Ruiz, 1997). Shared polymorphisms could also result from the balancer crosses that were used to generate the isochromosomal strains (Miller et al, 2016). See the Supporting Information for more details on how polymorphic sites were classified.…”

Unc-4 acts to promote neuronal identity and development of the take-off circuit in the Drosophila CNS

“…These flies lacked 3XP3-dsRED expression, which marks edited unc-4 locus and behaved like wild-type. Wild type unc-4 locus in the FM7 chromosome is located close to the Bar locus which is known to exhibit rare chromosomal arrangements (MILLER et al 2016).…”

Drosophila Chromosomal Polymorphism: From Population Aspects to Origin Mechanisms of Inversions

“…The inversion also influences the events of recombination of regions outside their limits. All these findings have implications for the analyses that use balancer chromosomes [32][33][34].…”

“…Despite the high acceptance and diffusion of the co-adaptation model of the genes contained in the inversions in Drosophila [15], alternative hypotheses point to different scenarios for the propagation and distribution of chromosomal inversions in populations of living beings, as a result of the increasing acquisition of knowledge and domain of improved analysis techniques [33,34,36].…”