Investigating X chromosome non-disjunction in Drosophila melanogaster su(var)3-9 mutants

Meiotic recombination is a highly regulated process necessary for promoting proper chromosome disjunction during the first meiotic division. Notably, reduced levels of meiotic recombination are observed in heterochromatic regions of the chromosome. This study seeks to investigate the molecular mechanisms underlying this observation by examining the effects of reduced heterochromatin on non-disjunction rates in Drosophila melanogaster. To accomplish this, we measured non-disjunction in wild-type and reduced heterochromatin mutant su(var)3-9 flies. To begin, we confirmed the presence of a mutation within su(var)3-9 via Sanger sequencing. Next, we created allele-specific primers and designed a PCR protocol to more accurately identify mutant flies at the molecular level. Finally, we assayed non-disjunction in wild-type and su(var)3-9 mutant flies and discovered that su(var)3-9 mutants have significantly higher levels of non-disjunction than wild-type flies. We also uncovered a striking sex bias in the non-disjunction progeny of su(var)3-9 mutants. INTRODUCTION Chromosomes are thread-like structures of DNA wrapped around histone proteins. This genetic information can be transferred across the generations through meiosis, a type of cellular division resulting in the production of four genetically diverse haploid gametes. During prophase I of meiosis, replicated paternal and maternal homologs pair up and exchange genetic material in a process called meiotic recombination, or crossing over. Notably, this exchange appears to show high levels of regulation. For example, crossover assurance states that every chromosome pair will engage in crossover events during cellular division (Shinohara et al. 2008), resulting in increased genetic variability among offspring. Moreover, crossover events are essential for proper chromosome segregation because they serve as a tether for homologs as they align at the metaphase plate (Youds and Boulton 2011). During normal cellular activity, chromosomes are also subject post-translational modifications to histones. These modifications — referred to as “marks” — regulate gene expression by organizing the genome into two distinct regions: euchromatin and heterochromatin. These distinct regions are characterized by their varying levels of DNA compaction and gene expression. Euchromatin, which comprises a large portion of the genome, functions as the most active region of gene expression. Alternatively, heterochromatin — found in regions proximal to the centromere and telomere — is characterized by tight DNA compaction, low gene density, transcriptional inertness, and strongly reduced levels of meiotic recombination. Studies have also shown that crossovers that occur in heterochromatic regions show strong inhibition of proper chromosome segregation (McKim et al. 2002). Within this study, we seek to examine the role of heterochromatin in the suppression of meiotic recombination and its effect on chromosome segregation. For this study, we used Drosophila melanogaster as a model system. Drosophila is an ideal organism for genetic studies because of their fast generation times, large brood sizes, and fully sequenced genome. Drosophila is also particularly useful for meiotic studies because flies are able to survive some forms of aneuploidy and moreover, meiotic crossing over was first observed in Drosophila. The Drosophila genome consists of 4 chromosome pairs including its sex chromosomes. With respect to its composition, chromosome 4 is >70% heterochromatic compared to the other