Osteopontin Is an Oncogenic Vav1– but not Wild-type Vav1–Responsive Gene: Implications for Fibroblast Transformation

Mammalian wild-type Vav1 (wtVav1) encodes a specific GDP/GTP nucleotide exchange factor that is exclusively expressed in the hematopoietic system. Despite numerous studies, the mechanism underlying transformation of fibroblasts by oncogenic Vav1 (oncVav1) is not well defined. We identified osteopontin, a marker for tumor aggressiveness, as an oncVav1-inducible gene. Osteopontin is highly expressed in oncVav1-transformed NIH3T3 cells (NIH/oncVav1) but is barely detected in NIH3T3 expressing wtVav1 (NIH/wtVav1) even following epidermal growth factor stimulation, which normally induces osteopontin. Depleting oncVav1 in NIH/oncVav1 using small interfering RNA led to a considerable decrease in osteopontin, whereas reducing osteopontin expression did not affect oncVav1 expression, suggesting that oncVav1 operates upstream of osteopontin. Vav1-depleted NIH/oncVav1 cells, but not osteopontin-depleted NIH/oncVav1 cells, exhibited impaired extracellular signal-regulated kinase (ERK) and c-Jun NH 2 -terminal kinase phosphorylation. Inhibition of ERK phosphorylation in NIH/oncVav1 cells led to a decrease in osteopontin expression, implying that the elevated osteopontin expression in these cells is dependent on ERK phosphorylation. Vav1-depleted or osteopontin-depleted NIH/oncVav1 cells lost their tumorigenic properties as judged by the soft agar and invasion assays, although loss of osteopontin expression had a less dramatic effect. Suppression of Vav1 expression in NIH/oncVav1 cells led to reversion to “normal” morphology, whereas when only osteopontin expression was diminished cells retained their transformed morphology. This work strongly supports a role for oncVav1 as a master oncogene and provides clues to the molecular mechanism underlying oncVav1 transformation. (Cancer Res 2006; 66(12): 6183-91)