Deletion or insertion in the first immunoglobulin-plexin-transcription (IPT) domain differentially regulates expression and tumorigenic activities of RON receptor Tyrosine Kinase

BACKGROUND: Activation of the RON receptor tyrosine kinase, a member of the c-MET family, regulates tumorigenic phenotypes. The RON extracellular domains are critical in regulating these activities. The objective of this study was to determine the role of the first IPT domain in regulating RON-mediated tumorigenic activities and the underlying mechanisms. RESULTS: Two RON variants, RON160 and RON(E5/6in )with deletion and insertion in the first IPT domain, respectively, were molecularly cloned. RON160 was a splicing variant generated by deletion of 109 amino acids encoded by exons 5 and 6. In contrast, RON(E5/6in )was derived from a transcript with an insertion of 20 amino acids between exons 5 and 6. Both RON160 and RON(E5/6in )were proteolytically matured into two-chain receptor and expressed on the cell surface. RON160 was constitutively active with tyrosine phosphorylation. However, activation of RON(E5/6in )required ligand stimulation. Deletion resulted in the resistance of RON160 to proteolytic digestion by cell associated trypsin-like enzymes. RON160 also resisted anti-RON antibody-induced receptor internalization. These features contributed to sustained intracellular signaling cascades. On the other hand, RON(E5/6in )was highly susceptible to protease digestion, which led to formation of a truncated variant known as RONp110. RON(E5/6in )also underwent rapid internalization upon anti-RON antibody treatment, which led to signaling attenuation. Although ligand-induced activation of RON(E5/6in )partially caused epithelial to mesenchymal transition (EMT), it was RON160 that showed cell-transforming activities in cell focus formation and anchorage-independent growth. RON160-mediated EMT is also associated with increased motile/invasive activity. CONCLUSIONS: Alterations in the first IPT domain in extracellular region differentially regulate RON mediated tumorigenic activities. Deletion of the first IPT results in formation of oncogenic variant RON160. Enhanced degradation and internalization with attenuated signaling cascades could be the mechanisms underlying non-tumorigenic features of RON(E5/6in).