Phospholipase C⑀ (PLC⑀) is a novel class of phosphoinositide-specific PLC characterized by possession of CDC25 homology and Ras/Rap1-associating domains. We and others have shown that human PLC⑀ is translocated from the cytoplasm to the plasma membrane and activated by direct association with Ras at its Ras/Rap1-associating domain. In addition, translocation to the perinuclear region was induced upon association with Rap1⅐GTP. However, the function of the CDC25 homology domain remains to be clarified. Here we show that the CDC25 homology domain of PLC⑀ functions as a guanine nucleotide exchange factor for Rap1 but not for any other Ras family GTPases examined including Rap2 and Ha-Ras. Consistent with this, coexpression of fulllength PLC⑀ or its N-terminal fragment carrying the CDC25 homology domain causes an increase of the intracellular level of Rap1⅐GTP. Concurrently, stimulation of the downstream kinases B-Raf and extracellular signalregulated kinase is observed, whereas the intracellular level of Ras⅐GTP and Raf-1 kinase activity are unaffected. In wild-type Rap1-overexpressing cells, epidermal growth factor induces translocation of PLC⑀ to the perinuclear compartments such as the Golgi apparatus, which is sustained for at least 20 min. In contrast, PLC⑀ lacking the CDC25 domain translocates to the perinuclear compartments only transiently. Further, the formation of Rap1⅐GTP upon epidermal growth factor stimulation exhibits a prolonged time course in cells expressing fulllength PLC⑀ compared with those expressing PLC⑀ lacking the CDC25 homology domain. These results suggest a pivotal role of the CDC25 homology domain in amplifying Rap1-dependent signal transduction, including the activation of PLC⑀ itself, at specific subcellular locations such as the Golgi apparatus.

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