YAP1 is a key transcriptional regulator that promotes cell proliferation, and is negatively regulated by several members of the Hippo pathway by nuclear exclusion and degradation of the YAP1 protein. Fusions between the N’-terminal regions of YAP1 and other downstream genes, such as YAP1-MAMLD1, YAP1-TFE3, and YAP1-FAM118B, have been identified in several cancers, and might be the driving force in these tumors. However, it remains unclear how these fusion genes drive tumorigenesis and if different YAP1 fusion genes function in similar ways.<br/>We used overlap extension PCR to generate several of the known YAP1 fusion products–using the wild type sequences as templates–and subsequently sub-cloned these constructs into RCAS plasmids. After transfection into DF1 chicken cells a western blot was run to confirm the presence of the fusion constructs. Luciferase assays were performed with YAP1 responsive promoters to characterize the YAP1 transcriptional activity of the fusion constructs and the wild type counter parts. Immunofluorescence stainings were used to assess the cellular localization of the fusion proteins. The YAP1-TFE3/FAM118B constructs did not recapitulate the enhanced YAP1 activity of the YAP1-MAMLD1 gene in the luciferase assays. However, all three fusion constructs showed a similar constitutive nuclear localization, which might circumvent negative regulation by the Hippo pathway and enable constitutive activity. Future experiments, such as RNA sequencing and ChIP sequencing will be required to further elucidate the exact mechanism of these fusion genes. Animal experiments will be performed to analyze which of these constructs are able to cause tumor formation <i>in vivo</i>.

Overexpression of YAP1-TFE3/FAM118B fusion genes

255th National Meeting (2018)

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