Dissecting the interplay between deregulated signaling networks and aberrant YAP/TAZ activity in myxoid liposarcoma

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Dissecting the interplay between deregulated signaling networks and aberrant YAP/TAZ activity in myxoid liposarcomaMyxoid liposarcoma (MLS) are malignant adipocytic tumors driven by the FUS-DDIT3 fusion gene encoding an aberrant transcription factor. The mechanisms whereby FUS-DDIT3 mediates sarcomagenesis are incompletely understood, and strategies to selectively target MLS cells remain elusive. We have previously shown that MLS cells are dependent on YAP, the central effector of the Hippo pathway, that FUS-DDIT3 promotes YAP expression and transcriptional activity, and that both proteins physically interact. Furthermore, we provided evidence of recurrent PI3K/AKT signaling pathway activation in MLS through mutations in PIK3CA, PTEN loss, or FUS-DDIT3-dependent transcriptional induction of a cell-autonomous IGF-II/IGF-IR feed-forward mechanism. Recent data from others documented physical interactions between the Hippo effectors YAP/TAZ and members of the epigenetic regulatory SWI/SNF complex, which modulate the formation YAP/TAZ-TEAD complexes. In addition, the FUS-DDIT3 protein itself was shown to associate with SWI/SNF subunits. The functional interplay between FUS-DDIT3, activated kinase signaling, the Hippo-YAP/TAZ cascade, and the SWI/SNF complex has not been investigated in MLS.The first part of the project will include immunohistochemical investigations and analyses of whole-exome/genome and RNA sequencing data in a large cohort of MLS tissues to identify informative patterns of YAP/TAZ, PI3K/AKT, and MEK/ERK activation.The second part will address regulatory dependencies between FUS-DDIT3, specific kinase pathways, and YAP/TAZ activation in vitro through (i) interrogation of kinase pathways known to be of relevance in MLS and (ii) an unbiased CRISPR interference screen to identify signaling intermediates specifically acting between kinases and phosphatases and the Hippo pathway.The third part will be dedicated to elucidating physical associations between FUS-DDIT3, YAP/TAZ, TEAD, and selected SWI/SNF complex proteins by systematic co-immunoprecipitation experiments, focusing on interactions that modulate YAP/TAZ signals in MLS.Employing a combined antibody-guided chromatin tagmentation sequencing and RNA sequencing approach, the fourth part will approach the transcriptional interplay between YAP/TAZ, FUS-DDIT3, and the SWI/SNF complex through identification of genomic loci/chromatin sites regulated in a specific/overlapping manner. Against this background, it will comparatively analyze the distribution of functional chromatin marks and gene expression profiles upon modulation of YAP/TAZ, FUS-DDIT3, and selected SWI/SNF complex members.The results of this project will contribute to the understanding of MLS biology and will lay the groundwork for the development of future molecularly targeted therapies.