SUMMARY Fertility requires that a mature oocyte become competent to transform into an embryo. This “egg activation” initiates with increases of Ca2+ in the oocyte. In mammals, the fertilizing sperm triggers a local rise in cytoplasmic Ca2+ that induces a wave of Ca2+ release from internal stores that moves across the egg. Oscillations in Ca2+ then occur via Ca2+ uptake from the external environment through TRPM7 channels. Drosophila egg activation also begins with a local rise in Ca2+ that induces a wave via Ca2+ release from internal stores. The initial Ca2+ rise is due to uptake of Ca2+ from the external environment. Recently, we found that TRPM, the Drosophila ortholog of TRPM7, mediates this Ca2+ uptake. Thus Ca2+ influx through a TRPM- channel is fundamental to fertility from flies to mammals, and dissecting its mechanism and effects is of relevance to human infertility diagnosis and its alleviation. However, how TRPM channels operate in oocytes, and how the Ca2+ rise relates to the macromolecular changes that make the oocyte competent to develop, is unknown in any organism. Drosophila’s genetic advantages, and large eggs, make it the ideal model system with which to determine such mechanisms and identify molecules critical for this transition in all animals. We will do so via two aims: First, we will investigate why TRPM channels cause a local rather than global calcium rise: are the channels localized at the oocyte poles or are they uniformly distributed around the oocyte plasma membrane and only activated at the poles? In addition to revealing how TRPM channels activate during egg activation, our results are relevant to how these channels act locally in several additional medically-relevant biological processes. Depending on which hypothesis we find to be correct, our future studies will determine how TRPM is localized to the poles during oogenesis or, alternatively how a non-localized channel is activated to act only locally. Second, we will follow up our surprising discovery that, similar to the situation in mouse, despite egg activation abnormalities TRPM-deficient Drosophila oocytes can initiate early embryogenesis, although development later becomes abnormal. We will investigate the relationship of TRPM-mediated Ca2+ influx to the phospho- modulation of essential maternal proteins that occurs during egg activation. We and others have shown that the Ca2+-activated phosphatase calcineurin is essential for egg activation in both flies and frogs. Exploiting genetic and biochemical advantages unique to Drosophila, we then showed that calcineurin activity regulates changes in the phospho-state of cell cycle regulators and translation initiation factors during egg activation. We propose here to determine whether these phosphoproteome changes depend on TRPM- mediated Ca2+ influx. If they do, our future studies will aim to identify how development can initiate in the absence of this phosphomodulation. If the calcineurin-based changes to th...