Gab2 regulates cytoskeletal organization and migration of mammary epithelial cells by modulating RhoA activation.
The docking protein Gab2 is overexpressed in several human malignancies, including breast cancer, and is associated with increased metastatic potential. Here we report that Gab2 overexpression in MCF-10A mammary epithelial cells led to delayed cell spreading, a decrease in stress fibers and mature focal adhesions, and enhanced cell migration. Expression of a Gab2 mutant uncoupled from 14-3-3-mediated negative feedback (Gab2(2xA)) led to a more mesenchymal morphology and acquisition of invasive potential. Expression of either Gab2 or Gab2(2xA) led to decreased activation of RhoA, but only the latter increased levels of Rac-GTP. Expression of constitutively active RhoA in MCF-10A/Gab2 cells restored stress fibers and focal adhesions, indicating that Gab2 signals upstream of RhoA to suppress these structures. Mutation of the two Shp2-binding sites to phenylalanine (Gab2(ΔShp2)) markedly reduced the effects of Gab2 on cellular phenotype and RhoA activation. Expression of Gab2 or Gab2(2xA), but not Gab2(ΔShp2), promoted Vav2 phosphorylation and plasma membrane recruitment of p190A RhoGAP. Knockdown of p190A RhoGAP reversed Gab2-mediated effects on stress fibers and focal adhesions. The identification of a novel pathway downstream of Gab2 involving negative regulation of RhoA by p190A RhoGAP sheds new light on the role of Gab2 in cancer progression.
Cancer Research Program and Phospholipid Biology Group, Garvan Institute of Medical Research, Sydney, NSW 2010, Australia.
Maria Teresa
MT
Herrera Abreu
William E
WE
Hughes
Katarina
K
Mele
Ruth J
RJ
Lyons
Danny
D
Rickwood
Brigid C
BC
Browne
Haley L
HL
Bennett
Pascal
P
Vallotton
Tilman
T
Brummer
Roger J
RJ
Daly
D048868:Adaptor Proteins, Signal Transducing / Q000235:genetics* / Q000378:metabolism*; D001940:Breast / Q000166:cytology* / Q000648:ultrastructure; D001943:Breast Neoplasms / Q000473:pathology; D002460:Cell Line; D002465:Cell Movement; D049109:Cell Proliferation; D003599:Cytoskeleton / Q000378:metabolism / Q000648:ultrastructure; D004847:Epithelial Cells / Q000502:physiology / Q000648:ultrastructure; D005260:Female; D022001:Focal Adhesions; D020690:GTPase-Activating Proteins / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D006801:Humans; D015151:Immunoblotting; D008856:Microscopy, Fluorescence; D009154:Mutation; D009361:Neoplasm Invasiveness; D009362:Neoplasm Metastasis; D010766:Phosphorylation; D054592:Protein Tyrosine Phosphatase, Non-Receptor Type 11 / Q000378:metabolism; D051567:Proto-Oncogene Proteins c-vav / Q000378:metabolism; D034741:RNA, Small Interfering; D022502:Stress Fibers / Q000378:metabolism / Q000648:ultrastructure; D020742:rhoA GTP-Binding Protein / Q000235:genetics / Q000378:metabolism*
C513043:ARHGAP5 protein, human; D048868:Adaptor Proteins, Signal Transducing; C490895:GAB2 protein, human; D020690:GTPase-Activating Proteins; D051567:Proto-Oncogene Proteins c-vav; D034741:RNA, Small Interfering; C093651:VAV2 protein, human; D054592:Protein Tyrosine Phosphatase, Non-Receptor Type 11; D020742:rhoA GTP-Binding Protein