Limits...
Dishevelled-1 regulates microtubule stability: a new function mediated by glycogen synthase kinase-3beta.

Krylova O, Messenger MJ, Salinas PC - J. Cell Biol. (2000)

Bottom Line: The microtubule stabilizing function of DVL-1 is mimicked by lithium-mediated inhibition of glycogen synthase kinase-3beta (GSK-3beta) and blocked by expression of GSK-3beta.These findings suggest that DVL-1, through GSK-3beta, can regulate microtubule dynamics.This new function of DVL-1 in controlling microtubule stability may have important implications for Dishevelled proteins in regulating cell polarity.

View Article: PubMed Central - PubMed

Affiliation: The Randall Institute, King's College London, London, United Kingdom.

ABSTRACT
Dishevelled has been implicated in the regulation of cell fate decisions, cell polarity, and neuronal function. However, the mechanism of Dishevelled action remains poorly understood. Here we examine the cellular localization and function of the mouse Dishevelled protein, DVL-1. Endogenous DVL-1 colocalizes with axonal microtubules and sediments with brain microtubules. Expression of DVL-1 protects stable microtubules from depolymerization by nocodazole in both dividing cells and differentiated neuroblastoma cells. Deletion analyses reveal that the PDZ domain, but not the DEP domain, of DVL-1 is required for microtubule stabilization. The microtubule stabilizing function of DVL-1 is mimicked by lithium-mediated inhibition of glycogen synthase kinase-3beta (GSK-3beta) and blocked by expression of GSK-3beta. These findings suggest that DVL-1, through GSK-3beta, can regulate microtubule dynamics. This new function of DVL-1 in controlling microtubule stability may have important implications for Dishevelled proteins in regulating cell polarity.

Show MeSH

Related in: MedlinePlus

The subcellular distribution of DVL-1 in maturing cerebellar granule cell neurons. Granule cell cultures were grown for 2 d and stained with antibodies to DVL-1 (A, D, G, and J), GAP-43 (B and E), and acetylated tubulin (H and K). DVL-1 has a punctate distribution in the neuronal cell bodies and along the axon shaft (A–C). High levels of DVL-1 were detected in the central domain of the growth cone (D and F). DVL-1 immunostaining colocalizes with acetylated tubulin along the axon shaft (G–I). In neurons fixed in the presence of detergent, a pool of DVL-1 remains colocalized with acetylated tubulin (J–L). CB, cell body; AS, axon shaft; GC, growth cone. Bar: A–C and G–I, 20 μM; D–F and J–L, 10 μM.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2189803&req=5

Figure 3: The subcellular distribution of DVL-1 in maturing cerebellar granule cell neurons. Granule cell cultures were grown for 2 d and stained with antibodies to DVL-1 (A, D, G, and J), GAP-43 (B and E), and acetylated tubulin (H and K). DVL-1 has a punctate distribution in the neuronal cell bodies and along the axon shaft (A–C). High levels of DVL-1 were detected in the central domain of the growth cone (D and F). DVL-1 immunostaining colocalizes with acetylated tubulin along the axon shaft (G–I). In neurons fixed in the presence of detergent, a pool of DVL-1 remains colocalized with acetylated tubulin (J–L). CB, cell body; AS, axon shaft; GC, growth cone. Bar: A–C and G–I, 20 μM; D–F and J–L, 10 μM.

Mentions: To determine the localization of DVL-1 in neurons, we examined DVL-1 in cerebellar granule cell cultures. Double labeling for GAP-43, a protein that reveals neuronal morphology, shows that DVL-1 protein has a punctate distribution in the cell body, along the axon shaft and at the growth cone (Fig. 3, A–F). At the growth cone, the highest levels of DVL-1 were found in the central domain that contains numerous MTs (Fig. 3, D–F). In the axon shaft, DVL-1 immunoreactivity seems to localize to regions of the axon where MTs are present (Fig. 3 A). To visualize MTs, we used acetylated tubulin antibody that labels stable MTs (Bulinski et al. 1988). Double labeling for acetylated MTs and DVL-1 reveals that high levels of DVL-1 appear to colocalize with axonal MTs (Fig. 3, G–L). To test whether DVL-1 interacts with MTs, cerebellar granule cells were detergent-extracted during fixation, a procedure that leaves the cytoskeleton and its associated proteins intact yet removes cytoplasmic and membrane-associated proteins. DVL-1 immunoreactivity was retained after detergent extraction, although at a lower level, suggesting that a fraction of DVL-1 is associated with the cytoskeleton (Fig. 3, J–L). In contrast, the membrane-bound protein, GAP-43, was removed by this treatment (data not shown). Most of the DVL-1 immunoreactivity colocalizes with acetylated MTs (Fig. 3, J–L). However, at the growth cone, low levels of DVL-1 were found in areas lacking acetylated MTs, suggesting that DVL-1 may also be associated with other cytoskeleton components (Fig. 3, J–L). These results suggest that a pool of DVL-1 colocalizes with axonal MTs.


Dishevelled-1 regulates microtubule stability: a new function mediated by glycogen synthase kinase-3beta.

Krylova O, Messenger MJ, Salinas PC - J. Cell Biol. (2000)

The subcellular distribution of DVL-1 in maturing cerebellar granule cell neurons. Granule cell cultures were grown for 2 d and stained with antibodies to DVL-1 (A, D, G, and J), GAP-43 (B and E), and acetylated tubulin (H and K). DVL-1 has a punctate distribution in the neuronal cell bodies and along the axon shaft (A–C). High levels of DVL-1 were detected in the central domain of the growth cone (D and F). DVL-1 immunostaining colocalizes with acetylated tubulin along the axon shaft (G–I). In neurons fixed in the presence of detergent, a pool of DVL-1 remains colocalized with acetylated tubulin (J–L). CB, cell body; AS, axon shaft; GC, growth cone. Bar: A–C and G–I, 20 μM; D–F and J–L, 10 μM.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2189803&req=5

Figure 3: The subcellular distribution of DVL-1 in maturing cerebellar granule cell neurons. Granule cell cultures were grown for 2 d and stained with antibodies to DVL-1 (A, D, G, and J), GAP-43 (B and E), and acetylated tubulin (H and K). DVL-1 has a punctate distribution in the neuronal cell bodies and along the axon shaft (A–C). High levels of DVL-1 were detected in the central domain of the growth cone (D and F). DVL-1 immunostaining colocalizes with acetylated tubulin along the axon shaft (G–I). In neurons fixed in the presence of detergent, a pool of DVL-1 remains colocalized with acetylated tubulin (J–L). CB, cell body; AS, axon shaft; GC, growth cone. Bar: A–C and G–I, 20 μM; D–F and J–L, 10 μM.
Mentions: To determine the localization of DVL-1 in neurons, we examined DVL-1 in cerebellar granule cell cultures. Double labeling for GAP-43, a protein that reveals neuronal morphology, shows that DVL-1 protein has a punctate distribution in the cell body, along the axon shaft and at the growth cone (Fig. 3, A–F). At the growth cone, the highest levels of DVL-1 were found in the central domain that contains numerous MTs (Fig. 3, D–F). In the axon shaft, DVL-1 immunoreactivity seems to localize to regions of the axon where MTs are present (Fig. 3 A). To visualize MTs, we used acetylated tubulin antibody that labels stable MTs (Bulinski et al. 1988). Double labeling for acetylated MTs and DVL-1 reveals that high levels of DVL-1 appear to colocalize with axonal MTs (Fig. 3, G–L). To test whether DVL-1 interacts with MTs, cerebellar granule cells were detergent-extracted during fixation, a procedure that leaves the cytoskeleton and its associated proteins intact yet removes cytoplasmic and membrane-associated proteins. DVL-1 immunoreactivity was retained after detergent extraction, although at a lower level, suggesting that a fraction of DVL-1 is associated with the cytoskeleton (Fig. 3, J–L). In contrast, the membrane-bound protein, GAP-43, was removed by this treatment (data not shown). Most of the DVL-1 immunoreactivity colocalizes with acetylated MTs (Fig. 3, J–L). However, at the growth cone, low levels of DVL-1 were found in areas lacking acetylated MTs, suggesting that DVL-1 may also be associated with other cytoskeleton components (Fig. 3, J–L). These results suggest that a pool of DVL-1 colocalizes with axonal MTs.

Bottom Line: The microtubule stabilizing function of DVL-1 is mimicked by lithium-mediated inhibition of glycogen synthase kinase-3beta (GSK-3beta) and blocked by expression of GSK-3beta.These findings suggest that DVL-1, through GSK-3beta, can regulate microtubule dynamics.This new function of DVL-1 in controlling microtubule stability may have important implications for Dishevelled proteins in regulating cell polarity.

View Article: PubMed Central - PubMed

Affiliation: The Randall Institute, King's College London, London, United Kingdom.

ABSTRACT
Dishevelled has been implicated in the regulation of cell fate decisions, cell polarity, and neuronal function. However, the mechanism of Dishevelled action remains poorly understood. Here we examine the cellular localization and function of the mouse Dishevelled protein, DVL-1. Endogenous DVL-1 colocalizes with axonal microtubules and sediments with brain microtubules. Expression of DVL-1 protects stable microtubules from depolymerization by nocodazole in both dividing cells and differentiated neuroblastoma cells. Deletion analyses reveal that the PDZ domain, but not the DEP domain, of DVL-1 is required for microtubule stabilization. The microtubule stabilizing function of DVL-1 is mimicked by lithium-mediated inhibition of glycogen synthase kinase-3beta (GSK-3beta) and blocked by expression of GSK-3beta. These findings suggest that DVL-1, through GSK-3beta, can regulate microtubule dynamics. This new function of DVL-1 in controlling microtubule stability may have important implications for Dishevelled proteins in regulating cell polarity.

Show MeSH
Related in: MedlinePlus