Proteomic analysis of fibroblast exosomes and characterization of the effects of TGF-beta treatment on the protein composition

Authors

• LM Jenkins
• VC Sanchez
• SH Yuspa

Abstract

Extracellular vesicles, such as exosomes, are a heterogeneous population of nano-sized vesicles with important regulatory roles. For example, they are involved in intercellular communications, stem cell maintenance, tissue repair, and immune surveillance, and play a part in the pathology underlying several diseases. Secretion of exosomes by tumor cells has been shown to induce tumor growth and promote metastatic phenotypes. To better understand the function of exosomes, mass spectrometry is used to characterize the proteomes of exosomes from different cell types and following cell treatment. Here, we have analyzed the effect of TGF-beta on exosomes released from primary murine fibroblasts. The exosomes were purified by differential centrifugation and ultracentrifugation, followed by density gradient fractionation. Mass spectrometry analysis of the protein content, coupled with label-free quantitation of changes in protein level, was performed to both characterize the basal composition of the exosomes and understand the effects of TGF-beta treatment. The proteomic results identified several important changes in the exosome proteome, including TGF-beta-dependent increases in the level of chloride intracellular channel protein 4 (CLIC4). These studies provide new insights into the role of exosomes.

Scientific Focus Area: Cancer Biology

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