Combining laser capture microdissection and proteomics reveals an active translation machinery controlling invadosome formation

• Published in: Nature communications Vol. 9; no. 1; pp. 2031 - 11
• Main Authors: Ezzoukhry, Zakaria, Henriet, Elodie, Cordelières, Fabrice P., Dupuy, Jean-William, Maître, Marlène, Gay, Nathan, Di-Tommaso, Sylvaine, Mercier, Luc, Goetz, Jacky G., Peter, Marion, Bard, Frédéric, Moreau, Violaine, Raymond, Anne-Aurélie, Saltel, Frédéric
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group UK 23.05.2018; Nature Publishing Group; Nature Portfolio
• Subjects: 14/63; 631/1647/2230; 631/337/475; 631/80/84/2339; 631/80/84/2340; 82/58; Actin; Actins - metabolism; Active control; Animals; Automation; Biomarkers, Tumor - analysis; Biomarkers, Tumor - metabolism; Cell Line, Tumor; Chromatography, High Pressure Liquid - methods; Combinatorial analysis; Extracellular matrix; Extracellular Matrix - metabolism; Humanities and Social Sciences; Humans; Laser Capture Microdissection - methods; Lasers; Life Sciences; Machinery and equipment; Mass spectrometry; Mass spectroscopy; Metastases; Mice; Molecular chains; multidisciplinary; Neoplasms - diagnosis; Neoplasms - pathology; NIH 3T3 Cells; Podosomes - metabolism; Podosomes - pathology; Protein Biosynthesis; Proteins; Proteomics; Proteomics - methods; RNA, Messenger - metabolism; Science; Science (multidisciplinary); Scientific imaging; Tandem Mass Spectrometry - methods; Translation
• ISSN: 2041-1723; 2041-1723
• DOI: 10.1038/s41467-018-04461-9

Invadosomes are F-actin-based structures involved in extracellular matrix degradation, cell invasion, and metastasis formation. Analyzing their proteome is crucial to decipher their molecular composition, to understand their mechanisms, and to find specific elements to target them. However, the specific analysis of invadosomes is challenging, because it is difficult to maintain their integrity during isolation. In addition, classical purification methods often suffer from contaminations, which may impair data validation. To ensure the specific identification of invadosome components, we here develop a method that combines laser microdissection and mass spectrometry, enabling the analysis of subcellular structures in their native state based on low amounts of input material. Using this combinatorial method, we show that invadosomes contain specific components of the translational machinery, in addition to known marker proteins. Moreover, functional validation reveals that protein translation activity is an inherent property of invadosomes, which is required to maintain invadosome structure and activity. Invadosomes degrade extracellular matrix and facilitate cell invasion but their molecular composition is not fully understood. Here, the authors combine laser capture and mass spectrometry to map the proteome of invadosomes, showing that they rely on internal translational activity to maintain their structure.