Limits...
FAK signaling is critical for ErbB-2/ErbB-3 receptor cooperation for oncogenic transformation and invasion.

Benlimame N, He Q, Jie S, Xiao D, Xu YJ, Loignon M, Schlaepfer DD, Alaoui-Jamali MA - J. Cell Biol. (2005)

Bottom Line: The overexpression of members of the ErbB tyrosine kinase receptor family has been associated with cancer progression.This colocalization requires intact FAK.In summary, distinct FAK signaling has an essential function in ErbB-induced oncogenesis and invasiveness.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine, Lady Davis Institute of the Sir Mortimer B. Davis Jewish General Hospital, McGill University, Montreal, Quebec H3T 1E2, Canada.

ABSTRACT
The overexpression of members of the ErbB tyrosine kinase receptor family has been associated with cancer progression. We demonstrate that focal adhesion kinase (FAK) is essential for oncogenic transformation and cell invasion that is induced by ErbB-2 and -3 receptor signaling. ErbB-2/3 overexpression in FAK-deficient cells fails to promote cell transformation and rescue chemotaxis deficiency. Restoration of FAK rescues both oncogenic transformation and invasion that is induced by ErbB-2/3 in vitro and in vivo. In contrast, the inhibition of FAK in FAK-proficient invasive cancer cells prevented cell invasion and metastasis formation. The activation of ErbB-2/3 regulates FAK phosphorylation at Tyr-397, -861, and -925. ErbB-induced oncogenic transformation correlates with the ability of FAK to restore ErbB-2/3-induced mitogen-activated protein kinase (MAPK) activation; the inhibition of MAPK prevented oncogenic transformation. In contrast, the inhibition of Src but not MAPK prevented ErbB-FAK-induced chemotaxis. In migratory cells, activated ErbB-2/3 receptors colocalize with activated FAK at cell protrusions. This colocalization requires intact FAK. In summary, distinct FAK signaling has an essential function in ErbB-induced oncogenesis and invasiveness.

Show MeSH

Related in: MedlinePlus

ErbB-2 colocalizes with FAK at focal adhesions in motile cells. (A) ErbB-2 and FAK colocalize in motile FAK+/+-2/3 cells. Confluent cells were scratch wounded and allowed to heal for the indicated time points before fixation. Cells were then coimmunostained with anti–ErbB-2 and anti-FAK antibodies followed by appropriate secondary antibodies as described in Materials and methods. Note that both ErbB-2 and FAK are recruited into newly formed lamellipodia near the leading edge of the wounded cells during cell migration to the acellular area (30 min). Typical ventral focal contacts that were stained for FAK become detectable 6 h after wound healing and become more pronounced after 24 h. Arrowheads indicate the newly formed protrusions. Stars indicate the folded cell layer at the wounded area. Bar, 30 μm. (B) Colocalization of ErbB-2 and FAK at the cell protrusion. Cells were fixed, permeabilized, and double immunostained with anti–ErbB-2 and anti-FAK antibodies followed by appropriate secondary antibodies conjugated either to aminomethylcoumarin (AMCA) or Texas red to detect ErbB-2 and FAK, respectively. The figure shows that FAK−/− control cells were negatively stained for both ErbB-2 and FAK, whereas FAK−/−-2/3 cells were strongly labeled for ErbB-2 receptors, which were homogeneously distributed around the cell periphery. In contrast, FAK+/+ control cells exhibit strong labeling of FAK, which was localized to cell extensions and ventral focal contact sites within the cells, whereas FAK+/+-2/3 cells exhibit strong labeling of FAK at cell protrusions, which colocalize partially with ErbB-2 receptors as revealed by dual-color merged confocal images. (C) Tyrosine-phosphorylated FAK colocalized with ErbB-2. FAK+/+ control cells and FAK+/+-2/3 were grown in complete medium, fixed, and immunostained with ErbB-2 in combination with either FAK Tyr-397, -861, or -925 antibodies. Confocal microscopy reveals partial colocalization of phospho-FAK with ErbB-2 receptor at the cell protrusions (arrows). In contrast, control cells that do not express any ErbB exhibit homogeneous distribution of phospho-FAK at the focal contacts. (B and C) Bars, 50 μm.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2171271&req=5

fig5: ErbB-2 colocalizes with FAK at focal adhesions in motile cells. (A) ErbB-2 and FAK colocalize in motile FAK+/+-2/3 cells. Confluent cells were scratch wounded and allowed to heal for the indicated time points before fixation. Cells were then coimmunostained with anti–ErbB-2 and anti-FAK antibodies followed by appropriate secondary antibodies as described in Materials and methods. Note that both ErbB-2 and FAK are recruited into newly formed lamellipodia near the leading edge of the wounded cells during cell migration to the acellular area (30 min). Typical ventral focal contacts that were stained for FAK become detectable 6 h after wound healing and become more pronounced after 24 h. Arrowheads indicate the newly formed protrusions. Stars indicate the folded cell layer at the wounded area. Bar, 30 μm. (B) Colocalization of ErbB-2 and FAK at the cell protrusion. Cells were fixed, permeabilized, and double immunostained with anti–ErbB-2 and anti-FAK antibodies followed by appropriate secondary antibodies conjugated either to aminomethylcoumarin (AMCA) or Texas red to detect ErbB-2 and FAK, respectively. The figure shows that FAK−/− control cells were negatively stained for both ErbB-2 and FAK, whereas FAK−/−-2/3 cells were strongly labeled for ErbB-2 receptors, which were homogeneously distributed around the cell periphery. In contrast, FAK+/+ control cells exhibit strong labeling of FAK, which was localized to cell extensions and ventral focal contact sites within the cells, whereas FAK+/+-2/3 cells exhibit strong labeling of FAK at cell protrusions, which colocalize partially with ErbB-2 receptors as revealed by dual-color merged confocal images. (C) Tyrosine-phosphorylated FAK colocalized with ErbB-2. FAK+/+ control cells and FAK+/+-2/3 were grown in complete medium, fixed, and immunostained with ErbB-2 in combination with either FAK Tyr-397, -861, or -925 antibodies. Confocal microscopy reveals partial colocalization of phospho-FAK with ErbB-2 receptor at the cell protrusions (arrows). In contrast, control cells that do not express any ErbB exhibit homogeneous distribution of phospho-FAK at the focal contacts. (B and C) Bars, 50 μm.

Mentions: To further understand the impact of ErbB–FAK interaction on the formation of focal adhesions in migratory cells, we first used the scratch-wound assay to follow ErbB–FAK localization at the cell protrusion. Fig. 5 A reveals that both ErbB-2 and FAK are recruited into newly formed lamellipodia near the leading edge of wounded cells during cell migration to the acellular area (Fig. 5 A, 30 min). Typical ventral focal contacts that were stained for FAK became detectable 6 h after wound healing and become more pronounced after 24 h (Fig. 5 A, 24 h).


FAK signaling is critical for ErbB-2/ErbB-3 receptor cooperation for oncogenic transformation and invasion.

Benlimame N, He Q, Jie S, Xiao D, Xu YJ, Loignon M, Schlaepfer DD, Alaoui-Jamali MA - J. Cell Biol. (2005)

ErbB-2 colocalizes with FAK at focal adhesions in motile cells. (A) ErbB-2 and FAK colocalize in motile FAK+/+-2/3 cells. Confluent cells were scratch wounded and allowed to heal for the indicated time points before fixation. Cells were then coimmunostained with anti–ErbB-2 and anti-FAK antibodies followed by appropriate secondary antibodies as described in Materials and methods. Note that both ErbB-2 and FAK are recruited into newly formed lamellipodia near the leading edge of the wounded cells during cell migration to the acellular area (30 min). Typical ventral focal contacts that were stained for FAK become detectable 6 h after wound healing and become more pronounced after 24 h. Arrowheads indicate the newly formed protrusions. Stars indicate the folded cell layer at the wounded area. Bar, 30 μm. (B) Colocalization of ErbB-2 and FAK at the cell protrusion. Cells were fixed, permeabilized, and double immunostained with anti–ErbB-2 and anti-FAK antibodies followed by appropriate secondary antibodies conjugated either to aminomethylcoumarin (AMCA) or Texas red to detect ErbB-2 and FAK, respectively. The figure shows that FAK−/− control cells were negatively stained for both ErbB-2 and FAK, whereas FAK−/−-2/3 cells were strongly labeled for ErbB-2 receptors, which were homogeneously distributed around the cell periphery. In contrast, FAK+/+ control cells exhibit strong labeling of FAK, which was localized to cell extensions and ventral focal contact sites within the cells, whereas FAK+/+-2/3 cells exhibit strong labeling of FAK at cell protrusions, which colocalize partially with ErbB-2 receptors as revealed by dual-color merged confocal images. (C) Tyrosine-phosphorylated FAK colocalized with ErbB-2. FAK+/+ control cells and FAK+/+-2/3 were grown in complete medium, fixed, and immunostained with ErbB-2 in combination with either FAK Tyr-397, -861, or -925 antibodies. Confocal microscopy reveals partial colocalization of phospho-FAK with ErbB-2 receptor at the cell protrusions (arrows). In contrast, control cells that do not express any ErbB exhibit homogeneous distribution of phospho-FAK at the focal contacts. (B and C) Bars, 50 μm.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2171271&req=5

fig5: ErbB-2 colocalizes with FAK at focal adhesions in motile cells. (A) ErbB-2 and FAK colocalize in motile FAK+/+-2/3 cells. Confluent cells were scratch wounded and allowed to heal for the indicated time points before fixation. Cells were then coimmunostained with anti–ErbB-2 and anti-FAK antibodies followed by appropriate secondary antibodies as described in Materials and methods. Note that both ErbB-2 and FAK are recruited into newly formed lamellipodia near the leading edge of the wounded cells during cell migration to the acellular area (30 min). Typical ventral focal contacts that were stained for FAK become detectable 6 h after wound healing and become more pronounced after 24 h. Arrowheads indicate the newly formed protrusions. Stars indicate the folded cell layer at the wounded area. Bar, 30 μm. (B) Colocalization of ErbB-2 and FAK at the cell protrusion. Cells were fixed, permeabilized, and double immunostained with anti–ErbB-2 and anti-FAK antibodies followed by appropriate secondary antibodies conjugated either to aminomethylcoumarin (AMCA) or Texas red to detect ErbB-2 and FAK, respectively. The figure shows that FAK−/− control cells were negatively stained for both ErbB-2 and FAK, whereas FAK−/−-2/3 cells were strongly labeled for ErbB-2 receptors, which were homogeneously distributed around the cell periphery. In contrast, FAK+/+ control cells exhibit strong labeling of FAK, which was localized to cell extensions and ventral focal contact sites within the cells, whereas FAK+/+-2/3 cells exhibit strong labeling of FAK at cell protrusions, which colocalize partially with ErbB-2 receptors as revealed by dual-color merged confocal images. (C) Tyrosine-phosphorylated FAK colocalized with ErbB-2. FAK+/+ control cells and FAK+/+-2/3 were grown in complete medium, fixed, and immunostained with ErbB-2 in combination with either FAK Tyr-397, -861, or -925 antibodies. Confocal microscopy reveals partial colocalization of phospho-FAK with ErbB-2 receptor at the cell protrusions (arrows). In contrast, control cells that do not express any ErbB exhibit homogeneous distribution of phospho-FAK at the focal contacts. (B and C) Bars, 50 μm.
Mentions: To further understand the impact of ErbB–FAK interaction on the formation of focal adhesions in migratory cells, we first used the scratch-wound assay to follow ErbB–FAK localization at the cell protrusion. Fig. 5 A reveals that both ErbB-2 and FAK are recruited into newly formed lamellipodia near the leading edge of wounded cells during cell migration to the acellular area (Fig. 5 A, 30 min). Typical ventral focal contacts that were stained for FAK became detectable 6 h after wound healing and become more pronounced after 24 h (Fig. 5 A, 24 h).

Bottom Line: The overexpression of members of the ErbB tyrosine kinase receptor family has been associated with cancer progression.This colocalization requires intact FAK.In summary, distinct FAK signaling has an essential function in ErbB-induced oncogenesis and invasiveness.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine, Lady Davis Institute of the Sir Mortimer B. Davis Jewish General Hospital, McGill University, Montreal, Quebec H3T 1E2, Canada.

ABSTRACT
The overexpression of members of the ErbB tyrosine kinase receptor family has been associated with cancer progression. We demonstrate that focal adhesion kinase (FAK) is essential for oncogenic transformation and cell invasion that is induced by ErbB-2 and -3 receptor signaling. ErbB-2/3 overexpression in FAK-deficient cells fails to promote cell transformation and rescue chemotaxis deficiency. Restoration of FAK rescues both oncogenic transformation and invasion that is induced by ErbB-2/3 in vitro and in vivo. In contrast, the inhibition of FAK in FAK-proficient invasive cancer cells prevented cell invasion and metastasis formation. The activation of ErbB-2/3 regulates FAK phosphorylation at Tyr-397, -861, and -925. ErbB-induced oncogenic transformation correlates with the ability of FAK to restore ErbB-2/3-induced mitogen-activated protein kinase (MAPK) activation; the inhibition of MAPK prevented oncogenic transformation. In contrast, the inhibition of Src but not MAPK prevented ErbB-FAK-induced chemotaxis. In migratory cells, activated ErbB-2/3 receptors colocalize with activated FAK at cell protrusions. This colocalization requires intact FAK. In summary, distinct FAK signaling has an essential function in ErbB-induced oncogenesis and invasiveness.

Show MeSH
Related in: MedlinePlus