Biology subjects

Hughes, S. E.

Publications and source records attributed to Hughes, S. E..

3 recordsLinked to original sources

bioRxiv2025

A proximity labeling approach to identify proteins that associate with synaptonemal complex components in Drosophila melanogaster females

Organisms use a specialized cell division called meiosis for the creation of haploid gametes. Multiple carefully orchestrated steps must occur at specific times and places for meiosis to be successful, including chromosome pairing, meiotic entry, recombination, synapsis, and two rounds of chromosome segregation. The regulation and molecular mechanisms for many of the steps of meiosis have not been fully elucidated. During synapsis, the synaptonemal complex (SC) builds along the entire lengths of the homologs to maintain the pairing of the homologs and promote the formation of the crossovers that help ensure proper segregation of homologs at the meiosis I division in many organisms. The SC is a large tripartite structure that is believed to function as a biomolecular condensate. To attempt to identify proteins that interact with SC components during female meiosis in Drosophila melanogaster, a protein of the lateral element, C(2)M, and a protein of the central element, Cona, were tagged with the APEX2 enzyme, which can biotinylate nearby proteins under the appropriate conditions. Under biotinylating promoting conditions, biotin labeled proteins were observed to be associated with the SC by immunofluorescence. Biotinylated proteins were isolated for mass spectrometry analysis, and multiple proteins were found to be enriched compared to control samples. RNAi knockdown lines targeting a subset of enriched proteins were examined for phenotypes in early Drosophila female meiosis. RNAi knockdown of Cpsf5, an mRNA cleavage factor, caused delayed or defective SC formation, as well as additional meiotic defects, indicating a role for maturation of mRNA in regulating processes of female meiosis. These results support proximity labeling as a strategy for identifying additional meiotic proteins.

genetics↗

bioRxiv2025

The recombination landscape of Drosophila melanogaster can be repatterned by a single gene

Meiotic recombination plays an important role in ensuring proper chromosome segregation during meiosis I through the creation of chiasmata that connect homologous chromosomes. Recombination plays an additional role in evolution by creating new allelic combinations. Organisms display species-specific crossover patterns, but how these patterns are established is poorly understood. Drosophila mauritana displays a differing recombination pattern compared to Drosophila melanogaster, with D. mauritiana experiencing a reduced centromere effect, the suppression of recombination emanating from the centromeres. To evaluate the contribution of the synaptonemal complex (SC) C(3)G protein to these recombination rate differences, the D. melanogaster allele was replaced with D. mauritiana c(3)G coding sequence. We found that the D. mauritiana C(3)G could interact with the D. melanogaster SC machinery to build full length tripartite SC and chromosomes segregated accurately, indicating sufficient crossovers were generated. However, the placement of crossovers was altered, displaying an increase in frequency of the centromere-proximal euchromatin indicating a decrease in the centromere effect similar to that observed in D. mauritiana. Recovery of chromatids with more than one crossover was also increased, likely due to the larger chromosome span now available for crossovers. As replacement of a single gene mediated a strong shift of one species crossover pattern towards another species, it indicates a small number of discrete factors may have major influence on species-specific crossover patterning. Additionally, it demonstrates the SC, a structure known to be required for crossover formation in many species, is likely one of these discrete factors. Lay AbstractMeiotic crossovers are important for ensuring proper chromosome segregation and generating genetic diversity. Different species display unique crossover patterns but the mechanisms that establish these patterns are poorly understood. The synaptonemal complex (SC) is built between meiotic chromosomes and promotes crossover formation. Replacement of the SC gene c(3)G in the fruit fly Drosophila melanogaster with Drosophila mauritiana c(3)G resulted in full-length SC assembly and proper chromosome segregation, but the D. melanogaster crossover pattern was shifted to appear more similar to D. mauritiana. This demonstrates that crossover patterning can be largely influenced by minor changes in the makeup of the SC.

genetics↗

bioRxiv2024

Patterns of crossover distribution in Drosophila mauritiana necessitate a re-thinking of the centromere effect on crossing over

We present a SNP-based crossover map for Drosophila mauritiana. Using females derived by crossing two different strains of D. mauritiana, we analyzed crossing over on all five major chromosome arms. Analysis of 105 male progeny allowed us to identify 327 crossover chromatids bearing single, double, or triple crossover events, representing 398 separate crossover events. We mapped these crossover sites along these five chromosome arms using a genome sequence map that includes the euchromatin-heterochromatin boundary. Confirming previous studies, we show that the overall crossover frequency in D. mauritiana is higher than is seen in D. melanogaster. Much of the increase in exchange frequency in D. mauritiana is due to a greatly diminished centromere effect. Using larval neuroblast metaphases from D. mauritiana -D. melanogaster hybrids we show that the lengths of the pericentromeric heterochromatin do not differ substantially between the two species, and thus cannot explain the observed differences in crossover distribution. Using a new and robust maximum likelihood estimation tool for obtaining Weinstein tetrad distributions, we observed an increase in bivalents with two or more crossovers when compared to D. melanogaster. This increase in crossing over along the arms of D. mauritiana likely reflects an expansion of the crossover-available euchromatin caused by the reduction in the centromere effect. The pattern of crossing over in D. mauritiana conflicts with the commonly accepted view of centromeres as polar suppressors of exchange (whose intensity is buffered by sequence non-specific heterochromatin) and demonstrates the importance of expanding such studies into other species of Drosophila. Article SummaryIn meiosis, homolog segregation is usually ensured by crossovers. The number and distribution of crossovers is in part regulated by cis-acting factors such as the cis- acting centromere effect, a polar suppression of exchange emanating from the vicinity of the centromere. We use SNP-based crossover mapping to show that in Drosophila mauritiana, the centromere effect is greatly reduced on four of the five major chromosome arms. We conclude that the centromere effect differs between Drosophila mauritiana and Drosophila melanogaster and that the ability to attenuate the centromere effect is not a general property of heterochromatin.

genetics↗