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Scientific EMU: Acquisition of a platform for single cell proteomics analysis

Abstract

The study of single cells is essential to investigate the molecular mechanisms that support population heterogeneity and, consequently, the development of diseases, allowing accurate answers to the cell biology and biological processes associated with specific cell types. In this scenario, genomics and transcriptomics studies of single cells have revealed the composition of tissue microenvironments, disease stages, immune response activity, metabolism, prognosis, risk of disease development, and response to therapy, among others. However, this knowledge at the level of nucleic acids provides an incomplete view of the cellular state, motivating the study at protein level, which are the main workhorses, and regulators of cellular life. Single-cell proteomic studies can surpass, without precedent, the knowledge of biology, such as signaling mechanisms based on the modification, interaction, activity, degradation, and localization of proteins in a single cell. Mass spectrometry-based proteomics has been used to analyze complex proteomes, and only recently it has been applied to quantify proteins in single cells. Considering that there is still no method for amplification of the complete proteome available, a specific workflow is needed to advance this frontier of knowledge. For this, the development of an automated platform is envisaged, combining (1) equipment for single cell separation and sample preparation for proteomics, with or without isobaric tags, CellenOne 1X, with (2) mass spectrometer with high sensitivity and quantitative capacity, an Orbitrap Eclipse coupled to the multifunctional liquid chromatography system Vanquish Neo. This platform will be installed at the Mass Spectrometry Laboratory (MAS), at the National Center for Research in Energy and Materials (CNPEM), which has a unique infrastructure to serve the users. Since 2010, the MAS facility has attended 527 research proposals from users outside the CNPEM, with 457 projects supported by FAPESP. In addition, CNPEM held ten annual proteomics events for training and dissemination of knowledge in the field of proteomics.The single-cell proteomic platform will be led by associated projects of CNPEM, AC Camargo Cancer Center and University of São Paulo Medical School, which seek to understand the molecular mechanisms of cancer initiation and immune response in patients with gastric and head and neck cancers, thus opening perspectives for new therapies or treatments for different types of cancer. In addition, about 70 complementary projects funded by FAPESP and other funding agencies can benefit from this platform, which will also be opened to internal and external users by submitting proposals through the SAUonline.This single-cell proteomic analysis platform, which is unique in Brazil and one of the pioneers worldwide, can contribute to the nascent field of single-cell proteomics, and allow a deeper understanding of the state of the art in biology, giving new insights into microenvironment heterogeneity and therapeutic resistance, elucidate mechanisms of immune response and immunotherapy, and accelerating the drug discovery. Finally, this platform will increase the international scientific competitiveness and leverage the translational of the findings to clinical practice. (AU)

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