Inositol 1,4,5-trisphosphate (IP 3 ) is an important second messenger in animal cells and is central to a wide range of cellular responses. The major intracellular activity of IP 3 is to regulate release of Ca 2ϩ from intracellular stores through IP 3 receptors (IP 3 Rs). We describe a system for the transient disruption of IP 3 signaling in the model organism Caenorhabditis elegans. The IP 3 binding domain of the C. elegans IP 3 R, ITR-1, was expressed from heat shock-induced promoters in live animals. This results in a dominant-negative effect caused by the overexpressed IP 3 binding domain acting as an IP 3 "sponge." Disruption of IP 3 signaling resulted in disrupted defecation, a phenotype predicted by previous genetic studies. This approach also identified two new IP 3 -mediated processes. First, the up-regulation of pharyngeal pumping in response to food is dependent on IP 3 signaling. RNA-mediated interference studies and analysis of itr-1 mutants show that this process is also IP 3 R dependent. Second, the tissue-specific expression of the dominant-negative construct enabled us to circumvent the sterility associated with loss of IP 3 signaling through the IP 3 R and thus determine that IP 3 -mediated signaling is required for multiple steps in embryogenesis, including cytokinesis and gastrulation. Inositol 1,4, ) is an important and widely used second messenger in animal cells. IP 3 is produced by the hydrolysis of phosphatidylinositol 4,5-bisphosphate after the activation of phospholipase C (PLC). The best understood routes of PLC activation are through the stimulation of G protein-coupled receptors or tyrosine kinase-linked receptors at the cell surface, although the presence of additional isoforms of PLC suggests other possible routes (Rhee and Bae, 1997;Katan, 1998). The only known action of IP 3 is to induce Ca 2ϩ release from intracellular stores through activation of IP 3 receptors (IP 3 Rs) (Berridge, 1993). This pathway and the Ca 2ϩ signals derived from it are central to a wide range of cellular responses (Berridge, 1993(Berridge, , 1997Clapham 1995).
INTRODUCTIONTwo challenges in the study of this signaling network are to unravel the different functions played by IP 3 signaling in animals and to develop methods for manipulating these processes and thus advance our understanding of how the specificity of IP 3 signaling is achieved. Recent genetic approaches have provided new insights into IP 3 function in animals. The central role played by IP 3 Rs in a wide range of cellular responses is reflected by the high level of lethality resulting from gene knockout observed in both Drosophila (Acharya et al., 1997;Venkatesh and Hasan, 1997) and mice (Matsumoto et al., 1996). Nevertheless, careful analysis has enabled, for example, the functions of IP 3 Rs in cerebellum in mice (Inoue et al., 1998) and phototransduction in Drosophila to be studied (Acharya et al., 1997;Raghu et al., 2000). Caenorhabditis elegans is also proving to be a powerful system for understanding IP 3 signaling function. IP 3 Rs i...