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Article Dans Une Revue EBioMedicine Année : 2017

Combined Mass Spectrometry Imaging and Top-down Microproteomics Reveals Evidence of a Hidden Proteome in Ovarian Cancer

Résumé

Background: Recently, it was demonstrated that proteins can be translated from alternative open reading frames (altORFs), increasing the size of the actual proteome. Top-down mass spectrometry-based proteomics allows the identification of intact proteins containing post-translational modifications (PTMs) as well as truncated forms translated from reference ORFs or altORFs. Methods: Top-down tissue microproteomics was applied on benign, tumor and necrotic-fibrotic regions of serous ovarian cancer biopsies, identifying proteins exhibiting region-specific cellular localization and PTMs. The regions of interest (ROIs) were determined by MALDI mass spectrometry imaging and spatial segmentation. Findings: Analysis with a customized protein sequence database containing reference and alternative proteins (altprots) identified 15 altprots, including alternative G protein nucleolar 1 (AltGNL1) found in the tumor, and translated from an altORF nested within the GNL1 canonical coding sequence. Co-expression of GNL1 and altGNL1 was validated by transfection in HEK293 and HeLa cells with an expression plasmid containing a GNL1-FLAG(V5) construct. Western blot and immunofluorescence experiments confirmed constitutive co-expression of altGNL1-V5 with GNL1-FLAG. Conclusions: Taken together, our approach provides means to evaluate protein changes in the case of serous ovarian cancer, allowing the detection of potential markers that have never been considered.

Dates et versions

inserm-02940659 , version 1 (16-09-2020)

Identifiants

Citer

Vivian Delcourt, Julien Franck, Eric Leblanc, Fabrice Narducci, Yves-Marie Robin, et al.. Combined Mass Spectrometry Imaging and Top-down Microproteomics Reveals Evidence of a Hidden Proteome in Ovarian Cancer. EBioMedicine, 2017, 21, pp.55-64. ⟨10.1016/j.ebiom.2017.06.001⟩. ⟨inserm-02940659⟩

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