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Alpha-E-catenin binds to dynamitin and regulates dynactin-mediated intracellular traffic.

Lien WH, Gelfand VI, Vasioukhin V - J. Cell Biol. (2008)

Bottom Line: Dynactin-mediated organelle trafficking is increased in alpha-E-catenin(-/-) keratinocytes, an effect that is reversed by expression of exogenous alpha-E-catenin.Although neither the integrity of dynactin-dynein complexes nor their association with vesicles is affected by alpha-E-catenin, alpha-E-catenin is necessary for the attenuation of microtubule-dependent trafficking by the actin cytoskeleton.Because the actin-binding domain of alpha-E-catenin is necessary for this regulation, we hypothesize that alpha-E-catenin functions as a dynamic link between the dynactin complex and actin and, thus, integrates the microtubule and actin cytoskeleton during intracellular trafficking.

View Article: PubMed Central - PubMed

Affiliation: Division of Human Biology, Fred Hutchinson Cancer Research Center, Seattle, WA 98109, USA.

ABSTRACT
Alpha-epithelial catenin (E-catenin) is an important cell-cell adhesion protein. In this study, we show that alpha-E-catenin also regulates intracellular traffic by binding to the dynactin complex component dynamitin. Dynactin-mediated organelle trafficking is increased in alpha-E-catenin(-/-) keratinocytes, an effect that is reversed by expression of exogenous alpha-E-catenin. Disruption of adherens junctions in low-calcium media does not affect dynactin-mediated traffic, indicating that alpha-E-catenin regulates traffic independently from its function in cell-cell adhesion. Although neither the integrity of dynactin-dynein complexes nor their association with vesicles is affected by alpha-E-catenin, alpha-E-catenin is necessary for the attenuation of microtubule-dependent trafficking by the actin cytoskeleton. Because the actin-binding domain of alpha-E-catenin is necessary for this regulation, we hypothesize that alpha-E-catenin functions as a dynamic link between the dynactin complex and actin and, thus, integrates the microtubule and actin cytoskeleton during intracellular trafficking.

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α–E-catenin is necessary to couple dynactin-mediated organelle traffic and the actin cytoskeleton. (A–D) Prominent actin cytoskeletons in wild-type (WT; A) and α–E-catenin−/− (KO; B) keratinocytes and their disruption by latrunculin A treatment (C and D). Immunofluorescence staining with phalloidin. (E and F) Quantitation of lysosome motility in wild-type (E) and α–E-catenin−/− (F) keratinocytes treated with latrunculin A (+LatruA) or DMSO control. Note that the disruption of the actin cytoskeleton significantly accelerates lysosome motility in wild-type cells (E) but has only a minor impact in α–E-catenin−/− cells (F). n = 336 for WT + DMSO, n = 337 for WT + LatruA, n = 319 for KO + DMSO, and n = 347 for KO + LatruA. (G) Quantitation of lysosome motility in α–E-catenin−/− cells expressing the HBT tag, full-length (HBT–α-cat), or VH1–VH2 fragment (HBT–α-cat VH1–2) of α–E-catenin. n = 326 for KO + HBT, n = 323 for KO + HBT–α-cat, and n = 327 for KO + HBT–α-cat VH1–2. Note the decrease in lysosome motility in cells expressing full-length but not truncated α–E-catenin. Quantitation was performed as described in Fig. 4 A. Bar, 23 μm.
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fig5: α–E-catenin is necessary to couple dynactin-mediated organelle traffic and the actin cytoskeleton. (A–D) Prominent actin cytoskeletons in wild-type (WT; A) and α–E-catenin−/− (KO; B) keratinocytes and their disruption by latrunculin A treatment (C and D). Immunofluorescence staining with phalloidin. (E and F) Quantitation of lysosome motility in wild-type (E) and α–E-catenin−/− (F) keratinocytes treated with latrunculin A (+LatruA) or DMSO control. Note that the disruption of the actin cytoskeleton significantly accelerates lysosome motility in wild-type cells (E) but has only a minor impact in α–E-catenin−/− cells (F). n = 336 for WT + DMSO, n = 337 for WT + LatruA, n = 319 for KO + DMSO, and n = 347 for KO + LatruA. (G) Quantitation of lysosome motility in α–E-catenin−/− cells expressing the HBT tag, full-length (HBT–α-cat), or VH1–VH2 fragment (HBT–α-cat VH1–2) of α–E-catenin. n = 326 for KO + HBT, n = 323 for KO + HBT–α-cat, and n = 327 for KO + HBT–α-cat VH1–2. Note the decrease in lysosome motility in cells expressing full-length but not truncated α–E-catenin. Quantitation was performed as described in Fig. 4 A. Bar, 23 μm.

Mentions: Cellular organelles and vesicles can use both actin filaments and microtubules for intracellular transport. Usually, microtubules are used for rapid long-range movements, and actin is used for short-range movements. Because vesicles can constantly switch between actin filaments and microtubules, the disruption of actin filaments often results in a significant increase in the mobility of vesicles and organelles (Cordonnier et al., 2001). We analyzed whether the integrity of the actin cytoskeleton is disrupted in α–E-catenin−/− cells. Staining for filamentous actin (F-actin) revealed the presence of well-organized F-actin bundles in α–E-catenin−/− cells (Fig. 5, A and B). We conclude that the organization of actin filaments is not disrupted in α–E-catenin−/− cells, and, thus, this cannot explain stimulation in dynactin-dependent traffic in α–E-catenin−/− keratinocytes.


Alpha-E-catenin binds to dynamitin and regulates dynactin-mediated intracellular traffic.

Lien WH, Gelfand VI, Vasioukhin V - J. Cell Biol. (2008)

α–E-catenin is necessary to couple dynactin-mediated organelle traffic and the actin cytoskeleton. (A–D) Prominent actin cytoskeletons in wild-type (WT; A) and α–E-catenin−/− (KO; B) keratinocytes and their disruption by latrunculin A treatment (C and D). Immunofluorescence staining with phalloidin. (E and F) Quantitation of lysosome motility in wild-type (E) and α–E-catenin−/− (F) keratinocytes treated with latrunculin A (+LatruA) or DMSO control. Note that the disruption of the actin cytoskeleton significantly accelerates lysosome motility in wild-type cells (E) but has only a minor impact in α–E-catenin−/− cells (F). n = 336 for WT + DMSO, n = 337 for WT + LatruA, n = 319 for KO + DMSO, and n = 347 for KO + LatruA. (G) Quantitation of lysosome motility in α–E-catenin−/− cells expressing the HBT tag, full-length (HBT–α-cat), or VH1–VH2 fragment (HBT–α-cat VH1–2) of α–E-catenin. n = 326 for KO + HBT, n = 323 for KO + HBT–α-cat, and n = 327 for KO + HBT–α-cat VH1–2. Note the decrease in lysosome motility in cells expressing full-length but not truncated α–E-catenin. Quantitation was performed as described in Fig. 4 A. Bar, 23 μm.
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fig5: α–E-catenin is necessary to couple dynactin-mediated organelle traffic and the actin cytoskeleton. (A–D) Prominent actin cytoskeletons in wild-type (WT; A) and α–E-catenin−/− (KO; B) keratinocytes and their disruption by latrunculin A treatment (C and D). Immunofluorescence staining with phalloidin. (E and F) Quantitation of lysosome motility in wild-type (E) and α–E-catenin−/− (F) keratinocytes treated with latrunculin A (+LatruA) or DMSO control. Note that the disruption of the actin cytoskeleton significantly accelerates lysosome motility in wild-type cells (E) but has only a minor impact in α–E-catenin−/− cells (F). n = 336 for WT + DMSO, n = 337 for WT + LatruA, n = 319 for KO + DMSO, and n = 347 for KO + LatruA. (G) Quantitation of lysosome motility in α–E-catenin−/− cells expressing the HBT tag, full-length (HBT–α-cat), or VH1–VH2 fragment (HBT–α-cat VH1–2) of α–E-catenin. n = 326 for KO + HBT, n = 323 for KO + HBT–α-cat, and n = 327 for KO + HBT–α-cat VH1–2. Note the decrease in lysosome motility in cells expressing full-length but not truncated α–E-catenin. Quantitation was performed as described in Fig. 4 A. Bar, 23 μm.
Mentions: Cellular organelles and vesicles can use both actin filaments and microtubules for intracellular transport. Usually, microtubules are used for rapid long-range movements, and actin is used for short-range movements. Because vesicles can constantly switch between actin filaments and microtubules, the disruption of actin filaments often results in a significant increase in the mobility of vesicles and organelles (Cordonnier et al., 2001). We analyzed whether the integrity of the actin cytoskeleton is disrupted in α–E-catenin−/− cells. Staining for filamentous actin (F-actin) revealed the presence of well-organized F-actin bundles in α–E-catenin−/− cells (Fig. 5, A and B). We conclude that the organization of actin filaments is not disrupted in α–E-catenin−/− cells, and, thus, this cannot explain stimulation in dynactin-dependent traffic in α–E-catenin−/− keratinocytes.

Bottom Line: Dynactin-mediated organelle trafficking is increased in alpha-E-catenin(-/-) keratinocytes, an effect that is reversed by expression of exogenous alpha-E-catenin.Although neither the integrity of dynactin-dynein complexes nor their association with vesicles is affected by alpha-E-catenin, alpha-E-catenin is necessary for the attenuation of microtubule-dependent trafficking by the actin cytoskeleton.Because the actin-binding domain of alpha-E-catenin is necessary for this regulation, we hypothesize that alpha-E-catenin functions as a dynamic link between the dynactin complex and actin and, thus, integrates the microtubule and actin cytoskeleton during intracellular trafficking.

View Article: PubMed Central - PubMed

Affiliation: Division of Human Biology, Fred Hutchinson Cancer Research Center, Seattle, WA 98109, USA.

ABSTRACT
Alpha-epithelial catenin (E-catenin) is an important cell-cell adhesion protein. In this study, we show that alpha-E-catenin also regulates intracellular traffic by binding to the dynactin complex component dynamitin. Dynactin-mediated organelle trafficking is increased in alpha-E-catenin(-/-) keratinocytes, an effect that is reversed by expression of exogenous alpha-E-catenin. Disruption of adherens junctions in low-calcium media does not affect dynactin-mediated traffic, indicating that alpha-E-catenin regulates traffic independently from its function in cell-cell adhesion. Although neither the integrity of dynactin-dynein complexes nor their association with vesicles is affected by alpha-E-catenin, alpha-E-catenin is necessary for the attenuation of microtubule-dependent trafficking by the actin cytoskeleton. Because the actin-binding domain of alpha-E-catenin is necessary for this regulation, we hypothesize that alpha-E-catenin functions as a dynamic link between the dynactin complex and actin and, thus, integrates the microtubule and actin cytoskeleton during intracellular trafficking.

Show MeSH
Related in: MedlinePlus