P190B RhoGAP has pro-tumorigenic functions during MMTV-Neu mammary tumorigenesis and metastasis.
BACKGROUND
Rho GTPases are overexpressed and hyperactivated in human breast cancers. Deficiency of p190B RhoGAP, a major inhibitor of the Rho GTPases, inhibits mouse mammary tumor virus long terminal repeat (MMTV)-Neu/ErbB2 mammary tumor formation and progression in part through effects within the stromal environment, suggesting that p190B function is pro-tumorigenic. To further investigate the potential pro-tumorigenic actions of p190B, we examined the effects of exogenous p190B expression within the mammary epithelium on MMTV-Neu tumor formation and progression.


METHODS
Tetracycline-regulatable p190B transgenic mice were bred to MMTV-Neu mice, and the effects of exogenous p190B expression on tumor latency, multiplicity, growth rates, angiogenesis, and metastasis were examined. The effects of exogenous p190B expression on cell-matrix adhesion and invasion were tested using non-transformed primary mammary epithelial cells (MECs). Rho GTPase activity, oxidative stress as an indicator of reactive oxygen species (ROS) production, and downstream signaling pathways were analyzed.


RESULTS
Altered p190B expression resulted in a 2-fold increase in tumor multiplicity and a 3-fold increase in metastases compared to control mice indicating that exogenous p190B expression in the mammary epithelium promotes MMTV-Neu mammary tumor formation and progression. Interestingly, non-transformed primary MECs expressing exogenous p190B displayed increased adhesion to laminin and type IV collagen and formed invasive structures in a three-dimensional culture assay. Ras related C3 botulinum toxin 1 (Rac1)-GTP levels were elevated in p190B transgenic tumors whereas Ras homologous A (RhoA) and cell division cycle 42 (Cdc42)-GTP levels were not significantly altered. Rac1 activity affects production of ROS, which regulate transformation, metastasis, and oxidative stress. Protein carbonylation, which is indicative of oxidative stress, was elevated 1.75-fold in p190B transgenic tumors as compared to control tumors suggesting that exogenous p190B expression may affect Rac1-dependent ROS production.


CONCLUSIONS
These studies indicate that paradoxically, p190B RhoGAP, a major inhibitor of the Rho GTPases in vitro, has pro-tumorigenic functions that enhance MMTV-Neu induced mammary tumor formation and metastasis. Furthermore, exogenous p190B expression enhances cell adhesion and invasion, which may facilitate metastasis. Rac1 activity and oxidative stress are elevated in tumors expressing exogenous p190B suggesting that p190B may promote tumorigenesis through a Rac1/ROS dependent mechanism.
McHenry
Peter R
PR
Department of Biochemistry and Molecular Biology, Indiana University School of Medicine, South Bend, 46617, USA.
Sears
James C
JC
Herrick
Matthew P
MP
Chang
Peggy
P
Heckman-Stoddard
Brandy M
BM
Rybarczyk
Megan
M
Chodosh
Lewis A
LA
Gunther
Edward J
EJ
Hilsenbeck
Susan G
SG
Rosen
Jeffrey M
JM
Vargo-Gogola
Tracy
T
D000818:Animals; D002448:Cell Adhesion; D002471:Cell Transformation, Neoplastic / Q000235:genetics / Q000378:metabolism*; D021982:Cell-Matrix Junctions; D002478:Cells, Cultured; D004847:Epithelial Cells / Q000378:metabolism / Q000473:pathology; D005260:Female; D020690:GTPase-Activating Proteins / Q000235:genetics / Q000378:metabolism*; D008321:Mammary Glands, Animal / Q000378:metabolism / Q000473:pathology; D015674:Mammary Neoplasms, Animal / Q000378:metabolism* / Q000473:pathology; D008324:Mammary Tumor Virus, Mouse / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D009362:Neoplasm Metastasis; D009389:Neovascularization, Pathologic; D018384:Oxidative Stress; D017382:Reactive Oxygen Species; D020764:cdc42 GTP-Binding Protein / Q000096:biosynthesis; D020830:rac1 GTP-Binding Protein / Q000096:biosynthesis; D020742:rhoA GTP-Binding Protein / Q000096:biosynthesis / Q000378:metabolism
C513042:Arhgap5 protein, mouse; D020690:GTPase-Activating Proteins; D017382:Reactive Oxygen Species; C085379:rho GTPase-activating protein; D020764:cdc42 GTP-Binding Protein; D020830:rac1 GTP-Binding Protein; D020742:rhoA GTP-Binding Protein