Identifying Genes that Interact with Calcineurin During Egg Activation in Drosophila melanogaster

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When an egg is fertilized, before the new embryo can begin to develop, the egg must be “activated.” The process of activation triggers the egg to complete meiosis, to begin translating new proteins, to degrade proteins and RNAs that are no longer required, and to modify its membranes to form a harder outer covering. The universal signaling event is an increase in calcium levels that occurs in the egg at this time. In many cell types, calcium causes changes through a conserved signaling pathway by binding to a protein called calmodulin, which in turn regulates the activities of other proteins. Calcineurin, a phosphatase, is one of those other proteins. It has been proven experimentally that calcineurin plays a critical role in regulating egg activation in fruit flies and in frogs. For example, Drosophila females expressing a constitutively active calcineurin (cnA-act) in their germline are sterile. To identify the proteins in the pathway by which calcineurin regulates egg activation, the Toshiro Aigaki lab tested whether there were Drosophila genes that interact genetically with calcineurin mutations. To do this, they searched for chromosomal regions whose deletion suppresses the sterility of cnA-act female flies. Fourteen large chromosomal regions were identified. To define the precise gene(s) within each region that suppresses the sterility of cnA-act females, I have tested smaller deletions, and then individual mutations, within the five regions I am examining for interaction with cnA-act. For three of the deletions, I have identified a gene or genes whose mutants significantly restore fertility of cnA-act females, and thus is a candidate to act within the calcineurin pathway. The first is CG6927, a gene that is expressed in the ovary but currently has no known function. The second is eIF3-S8, a transcription initiation factor. The third is RpS24, a ribosomal protein. The fourth is CG42565, which like CG6927, has no known function. In the other two regions, I have narrowed down the cnA-act interacting genes to a small number of candidates, instead of the hundreds of genes within the original large region. I am currently testing the remaining candidates.
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egg activation; calcineurin; Drosophila; fertilization; calmodulin
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