Functions of a rho-specific guanine nucleotide exchange factor in neurite retraction. Possible role of a proline-rich motif of KIAA0380 in localization.
The Rho/Rho kinase signaling pathway plays an essential role in neurite retraction and cell rounding in response to G(12/13)-coupled receptor activation in neuronal cells. The Rho guanine nucleotide exchange factor involved in these processes has not been identified. To monitor the activation state of Rho kinase, we developed a vimentin head/Rho kinase chimera, which is intramolecularly phosphorylated in a Rho-dependent manner at Ser(71) of the fused vimentin head. Using this system, we identified a clone termed KIAA0380, which contains the G alpha(12/13)-binding domain as well as a tandem of the Dbl homology/pleckstrin homology (DH/PH) domain, as an activator of Rho/Rho kinase signaling. Molecular dissection analyses revealed that a proline-rich motif C-terminally adjacent to DH/PH domain is essential for plasma membrane localization of KIAA0380 and cortical actin reorganization followed by cell rounding. In contrast, the DH/PH domain of KIAA0380 is localized in the cytoplasm, where it activates Rho/Rho kinase and induces stress fiber formation, consistent with results using p115 Rho guanine nucleotide exchange factor, which has a similar structure to KIAA0380 but lacks a proline-rich motif. These results suggest that upon stimulation, KIAA0380 translocates to the plasma membrane via the proline-rich motif and there activates Rho/Rho kinase signaling. In neuroblastoma Neuro2a cells, KIAA0380 was observed in the tips of neurites, a location where cortical actin reorganization is induced upon stimulation with lysophosphatidic acid. Ectopic expression of the N-terminal fragment inhibited lysophosphatidic acid-induced neurite retraction of Neuro2a cells. These results suggest that KIAA0380 plays an important role in neurite retraction through Rho-dependent signaling.
Togashi
H
H
Division of Biochemistry, Aichi Cancer Center Research Institute, 1-1 Chikusa-ku, Nagoya, Aichi 464-8681, Japan.
Nagata
K
K
Takagishi
M
M
Saitoh
N
N
Inagaki
M
M
D000818:Animals; D002460:Cell Line; D020439:Growth Cones / Q000502:physiology; D020662:Guanine Nucleotide Exchange Factors / Q000502:physiology*; D047908:Intracellular Signaling Peptides and Proteins; D051379:Mice; D016501:Neurites / Q000502:physiology*; D009474:Neurons / Q000166:cytology / Q000502:physiology; D017346:Protein Serine-Threonine Kinases / Q000502:physiology*; D015398:Signal Transduction / Q000502:physiology*; D020741:rho GTP-Binding Proteins / Q000502:physiology*; D054460:rho-Associated Kinases
D020662:Guanine Nucleotide Exchange Factors; D047908:Intracellular Signaling Peptides and Proteins; D017346:Protein Serine-Threonine Kinases; D054460:rho-Associated Kinases; D020741:rho GTP-Binding Proteins