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Control of axon elongation via an SDF-1alpha/Rho/mDia pathway in cultured cerebellar granule neurons.

Arakawa Y, Bito H, Furuyashiki T, Tsuji T, Takemoto-Kimura S, Kimura K, Nozaki K, Hashimoto N, Narumiya S - J. Cell Biol. (2003)

Bottom Line: SDF-1alpha-induced axon elongating activity under ROCK inhibition was replicated by the dominant-active form of the mammalian homologue of the Drosophila gene Diaphanous (mDia)1 and counteracted by its dominant-negative form.Furthermore, RNAi knockdown of mDia1 abolished SDF-1alpha-induced axon elongation.Together, our results support a critical role for an SDF-1alpha/Rho/mDia1 pathway in mediating axon elongation.

View Article: PubMed Central - PubMed

Affiliation: Dept. of Pharmacology, Kyoto University Faculty of Medicine, Yoshida, Sakyo-ku, Kyoto 606-8315, Japan.

ABSTRACT
Rho-GTPase has been implicated in axon outgrowth. However, not all of the critical steps controlled by Rho have been well characterized. Using cultured cerebellar granule neurons, we show here that stromal cell-derived factor (SDF)-1alpha, a neural chemokine, is a physiological ligand that can turn on two distinct Rho-dependent pathways with opposite consequences. A low concentration of the ligand stimulated a Rho-dependent pathway that mediated facilitation of axon elongation. In contrast, Rho/ROCK activation achieved by a higher concentration of SDF-1alpha caused repression of axon formation and induced no more increase in axon length. However, even at this higher concentration a Rho-dependent axon elongating activity could be recovered upon removal of ROCK activity using Y-27632. SDF-1alpha-induced axon elongating activity under ROCK inhibition was replicated by the dominant-active form of the mammalian homologue of the Drosophila gene Diaphanous (mDia)1 and counteracted by its dominant-negative form. Furthermore, RNAi knockdown of mDia1 abolished SDF-1alpha-induced axon elongation. Together, our results support a critical role for an SDF-1alpha/Rho/mDia1 pathway in mediating axon elongation.

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A putative role for mDia1 in facilitated process outgrowth. (A) Domain structure of wild-type and dominant mutant constructs of mDia1. (B) Coupling of elevated mDia1 activity (by overexpression of mDia1-ΔN3) with lower ROCK activity (in the presence of Y-27632) is sufficient to induce elongated axon-like processes in Swiss3T3 cells. Note the high amount of F-actin stained with phalloidin in the thin processes (arrowheads) of the GFP-mDia1-ΔN3–expressing cells. Bars, 50 μm.
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fig4: A putative role for mDia1 in facilitated process outgrowth. (A) Domain structure of wild-type and dominant mutant constructs of mDia1. (B) Coupling of elevated mDia1 activity (by overexpression of mDia1-ΔN3) with lower ROCK activity (in the presence of Y-27632) is sufficient to induce elongated axon-like processes in Swiss3T3 cells. Note the high amount of F-actin stained with phalloidin in the thin processes (arrowheads) of the GFP-mDia1-ΔN3–expressing cells. Bars, 50 μm.

Mentions: Recently, mDia was found to be a critical Rho effector that may predominantly act as an inducer/regulator of actin polymerization in HeLa cells and may be required for establishment of cell polarity and directed growth (Watanabe et al., 1997; Ishizaki et al., 2001; Ozaki-Kuroda et al., 2001; Tsuji et al., 2002). Indeed, we found that Swiss3T3 cells can elongate prolonged neurite-like processes best when higher mDia activity was achieved by overexpression a DA form of mDia1 (mDia1-ΔN3; Fig. 4 A; Ishizaki et al., 2001) and was coupled with lower ROCK activity in the presence of 30 μM Y-27632 (Fig. 4 B), consistent with previous work from our laboratory (Tsuji et al., 2002).


Control of axon elongation via an SDF-1alpha/Rho/mDia pathway in cultured cerebellar granule neurons.

Arakawa Y, Bito H, Furuyashiki T, Tsuji T, Takemoto-Kimura S, Kimura K, Nozaki K, Hashimoto N, Narumiya S - J. Cell Biol. (2003)

A putative role for mDia1 in facilitated process outgrowth. (A) Domain structure of wild-type and dominant mutant constructs of mDia1. (B) Coupling of elevated mDia1 activity (by overexpression of mDia1-ΔN3) with lower ROCK activity (in the presence of Y-27632) is sufficient to induce elongated axon-like processes in Swiss3T3 cells. Note the high amount of F-actin stained with phalloidin in the thin processes (arrowheads) of the GFP-mDia1-ΔN3–expressing cells. Bars, 50 μm.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2172896&req=5

fig4: A putative role for mDia1 in facilitated process outgrowth. (A) Domain structure of wild-type and dominant mutant constructs of mDia1. (B) Coupling of elevated mDia1 activity (by overexpression of mDia1-ΔN3) with lower ROCK activity (in the presence of Y-27632) is sufficient to induce elongated axon-like processes in Swiss3T3 cells. Note the high amount of F-actin stained with phalloidin in the thin processes (arrowheads) of the GFP-mDia1-ΔN3–expressing cells. Bars, 50 μm.
Mentions: Recently, mDia was found to be a critical Rho effector that may predominantly act as an inducer/regulator of actin polymerization in HeLa cells and may be required for establishment of cell polarity and directed growth (Watanabe et al., 1997; Ishizaki et al., 2001; Ozaki-Kuroda et al., 2001; Tsuji et al., 2002). Indeed, we found that Swiss3T3 cells can elongate prolonged neurite-like processes best when higher mDia activity was achieved by overexpression a DA form of mDia1 (mDia1-ΔN3; Fig. 4 A; Ishizaki et al., 2001) and was coupled with lower ROCK activity in the presence of 30 μM Y-27632 (Fig. 4 B), consistent with previous work from our laboratory (Tsuji et al., 2002).

Bottom Line: SDF-1alpha-induced axon elongating activity under ROCK inhibition was replicated by the dominant-active form of the mammalian homologue of the Drosophila gene Diaphanous (mDia)1 and counteracted by its dominant-negative form.Furthermore, RNAi knockdown of mDia1 abolished SDF-1alpha-induced axon elongation.Together, our results support a critical role for an SDF-1alpha/Rho/mDia1 pathway in mediating axon elongation.

View Article: PubMed Central - PubMed

Affiliation: Dept. of Pharmacology, Kyoto University Faculty of Medicine, Yoshida, Sakyo-ku, Kyoto 606-8315, Japan.

ABSTRACT
Rho-GTPase has been implicated in axon outgrowth. However, not all of the critical steps controlled by Rho have been well characterized. Using cultured cerebellar granule neurons, we show here that stromal cell-derived factor (SDF)-1alpha, a neural chemokine, is a physiological ligand that can turn on two distinct Rho-dependent pathways with opposite consequences. A low concentration of the ligand stimulated a Rho-dependent pathway that mediated facilitation of axon elongation. In contrast, Rho/ROCK activation achieved by a higher concentration of SDF-1alpha caused repression of axon formation and induced no more increase in axon length. However, even at this higher concentration a Rho-dependent axon elongating activity could be recovered upon removal of ROCK activity using Y-27632. SDF-1alpha-induced axon elongating activity under ROCK inhibition was replicated by the dominant-active form of the mammalian homologue of the Drosophila gene Diaphanous (mDia)1 and counteracted by its dominant-negative form. Furthermore, RNAi knockdown of mDia1 abolished SDF-1alpha-induced axon elongation. Together, our results support a critical role for an SDF-1alpha/Rho/mDia1 pathway in mediating axon elongation.

Show MeSH