Work

The differential roles of rad9 alternatively spliced forms in double- strand DNA break repair during Drosophila meiosis

Year: 2025

Type: article

Abstract: The 9–1–1 complex, comprising the Rad9, Hus1 and Rad1 proteins, is believed to operate as a component of a DNA damage checkpoint pathway. Our initial analysis of the Drosophila hus1 gene showed that Hus1 plays a dual role in meiosis, regulating both meiotic DNA damage checkpoint and homologous recombination repair. In this study, we further analyzed the meiotic roles of another protein in the complex, Rad9, which has two alternatively spliced forms, Rad9A and Rad9B. Using CRISPR/Cas9, we generated flies mutant for both rad9 isoforms. We found that, similarly to hus1 , mutations in rad9 lead to female sterility. Also, double-strand DNA breaks (DSBs) that form during meiosis are not processed efficiently, and the DNA within the oocyte nucleus fails to form its characteristic shape in rad9 mutants. On the other hand, the hus1 mutation completely disrupts checkpoint activation in DSB repair enzyme mutants, whereas the rad9 mutation only partially impairs checkpoint activation in this context. Moreover, spatial rescue experiments revealed that Rad9B is efficient in repairing meiotic DSBs, while Rad9A is not. Furthermore, we found that female fertility in rad9 mutants depends on early efficient meiotic DSB repair but not on karyosome formation. In summary, our results demonstrate a differential role of Rad9 alternatively spliced forms during Drosophila meiosis in oogenesis, and while former studies showed that Hus1 is sufficient for the effective activation of the meiotic recombination checkpoint, our results revealed that this is not true for Rad9. • Drosophila Rad9A is necessary but not sufficient for the efficient repair of meiotic DSBs. • Drosophila Rad9B is both necessary and sufficient for the efficient repair of meiotic DSBs. • Early and efficient repair of meiotic DSBs is necessary for Drosophila female fertility. • Mutation in rad9 partially activates the Drosophila meiotic recombination checkpoint via Checkpoint kinase 2.

Source: DNA repair

DOI: 10.1016/j.dnarep.2025.103833

Authors: Bareket Goldstein, Suad Sheikh-Suliman, Anna Bakhrat, Uri Abdu

Institution: Ben-Gurion University of the Negev

Language: English

Topic: DNA Repair Mechanisms

Subfield: Molecular Biology

Field: Biochemistry, Genetics and Molecular Biology

Domain: Life Sciences

Open Access status: hybrid

Estimated APC paid: $3,390

Funder: Israel Science Foundation