RalA mediates v-Src, v-Ras, and v-Raf regulation of CD44 and fibronectin expression in NIH3T3 fibroblasts

Area of Science:

• Cell Biology
• Molecular Biology
• Oncology

Background:

• Oncogenic transformation of fibroblasts by v-Src and v-Ras often leads to decreased fibronectin (FN) and elevated CD44 expression.
• The small GTPase RalA is a key mediator in transformation and tumorigenesis, activated by v-Src, v-Ras, and v-Raf, and also activates phospholipase D.

Purpose of the Study:

• To investigate the role of RalA in the downregulation of fibronectin (FN) and overexpression of CD44 during oncogenic transformation.
• To determine if RalA mediates the signaling pathways activated by v-Src, v-Ras, and v-Raf that affect FN and CD44 levels.

Main Methods:

• Comparison of FN and CD44 levels in NIH3T3 cells transformed by v-Src, v-Ras, or v-Raf versus untransfected cells.
• Assessment of extracellular FN fibril formation in transformed and control cells.
• Evaluation of the effects of co-expressing a dominant-negative S28N-RalA mutant on FN and CD44 levels and FN fibril formation.

Main Results:

• NIH3T3 cells transformed by v-Src, v-Ras, or v-Raf exhibited reduced FN levels and increased CD44 expression compared to untransfected cells.
• The ability of transformed cells to form extracellular FN fibrils was significantly diminished.
• Co-expression of dominant-negative S28N-RalA restored normal FN and CD44 levels and the capacity for FN fibril formation in v-Src-, v-Ras-, and v-Raf-transformed cells.

Conclusions:

• RalA plays a novel regulatory role in mediating the signal transduction pathways activated by v-Src, v-Ras, and v-Raf.
• RalA is essential for the oncogene-induced downregulation of fibronectin and overexpression of CD44.
• RalA is required for tumor formation in v-Src-, v-Ras-, and v-Raf-transformed NIH3T3 cells.