Dataset Information

The FUS-DDIT3 Interactome in Myxoid Liposarcoma.

ABSTRACT: Myxoid liposarcoma is a malignant lipogenic tumor that develops in deep soft tissues. While local control rates are good, current chemotherapy options remain ineffective against metastatic disease. Myxoid liposarcoma is characterized by the FUS-DDIT3 fusion oncoprotein that is proposed to function as an aberrant transcription factor, but its exact mechanism of action has remained unclear. To identify the key functional interacting partners of FUS-DDIT3, this study utilized immunoprecipitation-mass spectrometry (IP-MS) to identify the FUS-DDIT3 interactome in whole cell lysates of myxoid liposarcoma cells, and results showed an enrichment of RNA processing proteins. Further quantitative MS analyses of FUS-DDIT3 complexes isolated from nuclear lysates showed that members of several chromatin regulatory complexes were present in the FUS-DDIT3 interactome, including NuRD, SWI/SNF, PRC1, PRC2, and MLL1 COMPASS-like complexes. Co-immunoprecipitation validated the associations of FUS-DDIT3 with BRG1/SMARCA4, BAF155/SMARCC1, BAF57/SMARCE1, and KDM1A. Data from this study provides candidates for functional validation as potential therapeutic targets, particularly for emerging epigenetic drugs.

SUBMITTER: Yu JSE

PROVIDER: S-EPMC6584455 | biostudies-literature | 2019 Aug

REPOSITORIES: biostudies-literature

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The FUS-DDIT3 Interactome in Myxoid Liposarcoma.

Yu Jamie S E JSE Colborne Shane S Hughes Christopher S CS Morin Gregg B GB Nielsen Torsten O TO

Neoplasia (New York, N.Y.) 20190617 8

Myxoid liposarcoma is a malignant lipogenic tumor that develops in deep soft tissues. While local control rates are good, current chemotherapy options remain ineffective against metastatic disease. Myxoid liposarcoma is characterized by the FUS-DDIT3 fusion oncoprotein that is proposed to function as an aberrant transcription factor, but its exact mechanism of action has remained unclear. To identify the key functional interacting partners of FUS-DDIT3, this study utilized immunoprecipitation-ma ...[more]

PMID: 31220736

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Project description:Myxoid liposarcoma has the pathognomonic fusion oncogene FUS-DDIT3 encoding a chimeric transcription factor. Metastatic risk is higher with an increased round cell component and has been linked to aberrations involving the IGFR/PI3K/AKT pathway. These molecular insights have yet to translate to targeted therapies, and the lack of experimental models is a major hindrance. We describe the initial in-depth characterization of a new cell line (DL-221) and establishment of a mouse xenograft model. The cell line DL-221 was derived from a metastatic pleural lesion showing myxoid and round cell histology. This newly established cell line was characterized for phenotypic properties and molecular cytogenetic profile, using PCR, COBRA-FISH, and western blot. Next-generation whole-exome sequencing was performed to further characterize the cell line and the parent tumor. NOD-SCID-IL2R gamma knockout mice were xenograft hosts. DL-221 cells grew an adhering monolayer and COBRA-FISH showed an aneuploid karyotype with t(12;16)(q13;p11) and several other rearrangements; RT-PCR demonstrated a FUS-DDIT3 fusion transcript type 1. Both the cell line and the original tumor harbored a TP53 compound heterozygous mutation in exon 4 and 7, and were wild-type for PIK3CA. Moreover, among the 1254 variants called by whole-exome sequencing, there was 77% concordance between the cell line and parent tumor. The recently described hotspot mutation in the TERT promoter region in myxoid liposarcomas was also found at C228T in DL-221. Xenografts suitable for additional preclinical studies were successfully established in mice after subcutaneous injection. The established DL-221 cell line is the only published available myxoid liposarcoma cell line that underwent spontaneous immortalization, without requiring SV40 transformation. The cell line and its xenograft model are unique and helpful tools to study the biology and novel potential-targeted treatment approaches for myxoid liposarcoma.

Project description:BackgroundLiposarcomas are the most common class of soft tissue sarcomas, and myxoid liposarcoma is the second most common liposarcoma. EWSR1-DDIT3 is a chimeric fusion protein generated by the myxoid liposarcoma-specific chromosomal translocation t(12;22)(q13;q12). Current studies indicate that multipotent mesenchymal cells are the origin of sarcomas. The mechanism whereby EWSR1-DDIT3 contributes to the phenotypic selection of target cells during oncogenic transformation remains to be elucidated.Methodology/principal findingsReporter assays showed that the EWSR1-DDIT3 myxoid liposarcoma fusion protein, but not its wild-type counterparts EWSR1 and DDIT3, selectively repressed the transcriptional activity of cell lineage-specific marker genes in multipotent mesenchymal C3H10T1/2 cells. Specifically, the osteoblastic marker Opn promoter and chondrocytic marker Col11a2 promoter were repressed, while the adipocytic marker Ppar-?2 promoter was not affected. Mutation analyses, transient ChIP assays, and treatment of cells with trichostatin A (a potent inhibitor of histone deacetylases) or 5-Aza-2'-deoxycytidine (a methylation-resistant cytosine homolog) revealed the possible molecular mechanisms underlying the above-mentioned selective transcriptional repression. The first is a genetic action of the EWSR1-DDIT3 fusion protein, which results in binding to the functional C/EBP site within Opn and Col11a2 promoters through interaction of its DNA-binding domain and subsequent interference with endogenous C/EBP? function. Another possible mechanism is an epigenetic action of EWSR1-DDIT3, which enhances histone deacetylation, DNA methylation, and histone H3K9 trimethylation at the transcriptional repression site. We hypothesize that EWSR1-DDIT3-mediated transcriptional regulation may modulate the target cell lineage through target gene-specific genetic and epigenetic conversions.Conclusions/significanceThis study elucidates the molecular mechanisms underlying EWSR1-DDIT3 fusion protein-mediated phenotypic selection of putative target multipotent mesenchymal cells during myxoid liposarcoma development. A better understanding of this process is fundamental to the elucidation of possible direct lineage reprogramming in oncogenic sarcoma transformation mediated by fusion proteins.

Dataset Information

The FUS-DDIT3 Interactome in Myxoid Liposarcoma.

Publications

The FUS-DDIT3 Interactome in Myxoid Liposarcoma.

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