UCSF

Receptor tyrosine kinase (RTK) fusions are recently described drivers in lung adenocarcinoma that arise from chromosomal rearrangements, resulting in the C-terminal kinase domain of an RTK attached to a variety of N-terminal fusion partners. Here, we elucidate the importance of N-terminal partners of ROS1 and ALK fusions in driving oncoprotein localization and downstream signaling pathway activation. ROS1 gene fusions involve rearrangements with several 5’ genes, and we have discovered that the resultant N-terminal partner mediates the intracellular localization of the fusion. This localization, in turn, dictates which downstream pathways these fusions are able to engage, which determines signaling pathway dependency in ROS1 fusion-positive cells. We find that some ROS1 fusions are present on endosomes and are able to activate the RAS/MAPK pathway, while the most common fusion, CD74-ROS1, is found on the endoplasmic reticulum (ER), where it is unable to engage this pathway. Mislocalization of CD74-ROS1 from the ER to the endosome results in activation of the MAPK pathway by this fusion, demonstrating that localization is critical in driving downstream signaling pathway activation. These findings have clinical significance, as we reveal that ROS1 fusions that can activate MAPK form more aggressive tumors and are less responsive to the targeted ROS1 inhibitor crizotinib in vivo. The most common ALK fusion seen in lung cancer is EML4-ALK variant 1 (v1), and here we demonstrate that EML4-ALKv1 exists in intracellular non-membrane-associated clusters, and that this localization and cluster formation is dependent on oligimerization domains present within EML4. Additionally, we reveal that EML4-ALKv1 is able to engage the RAS/MAPK pathway despite the absence of a membrane, which runs counter to the canonical idea that RAS requires a membrane scaffold for downstream pathway activation. In summary, my findings demonstrate both the importance of N-terminal RTK fusion partners in driving downstream oncogenic signaling pathways, which may inform therapeutic approaches for treatment of ROS1 and ALK fusion-positive patients, and uncovers a novel signaling role for cytoplasmic RAS, representing a shift in how we understand a central node of signaling biology.