Regulatory mechanisms for Ras family and Rho family GTP-binding protein networks

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Dbl family guanine nucleotide exchange factors (GEFs), which target the Rho family, direct various cellular functions, such as cell motility, cell adhesion, and cell polarity, through cytoskeletal rearrangements. Here, we investigated the role of the Sec14-like domain of the Dbl family proteins Dbl and Ost, which is localized at the N terminal portion.Splice variants containing this domain, which are localized in the perinuclear region, did not induce change of cell shape, whereas those without the Sec14-like domain were localized in the plasma membrane, and induced filopodia formation. By employing a novel method to visualize the activation of the Rho family in situ, we demonstrated that Cdc42 colocalized with Dbl in the plasma membrane is in fact activated. Therefore, the Sec14-like domain has an important role in the regulation of subcellular localization and activation of Dbl family GEFs. We also analyzed insulin-dependent activation of Rac and Cdc42 in the muscle cell line L6 to examine the involvement of the Rho family in insulin-stimulated glucose uptake in the muscle. We clarified that the activation of Rac was partially dependent on phosphoinositide 3-kinase (PI3K), and PI3K-dependent activation of Rac occurred in the ruffling membrane areas, whereas PI3K-independent activation was observed in endomembranes.