1. Academic Validation
  2. Unraveling mitotic protein networks by 3D multiplexed epitope drug screening

Unraveling mitotic protein networks by 3D multiplexed epitope drug screening

  • Mol Syst Biol. 2018 Aug 13;14(8):e8238. doi: 10.15252/msb.20188238.
Lorenz J Maier 1 2 3 Stefan M Kallenberger 1 2 3 Katharina Jechow 2 4 Marcel Waschow 2 Roland Eils 5 2 4 Christian Conrad 5 2
Affiliations

Affiliations

  • 1 Center for Quantitative Analysis of Molecular and Cellular Biosystems (BioQuant), Heidelberg University, Heidelberg, Germany.
  • 2 Division of Theoretical Bioinformatics, German Cancer Research Center (DKFZ), Heidelberg, Germany.
  • 3 Department for Bioinformatics and Functional Genomics, Institute for Pharmacy and Molecular Biotechnology (IPMB), Heidelberg University, Heidelberg, Germany.
  • 4 BIH Center for Digital Health, Charité Universitätsmedizin Berlin and Berlin Institute of Health, Berlin, Germany.
  • 5 Center for Quantitative Analysis of Molecular and Cellular Biosystems (BioQuant), Heidelberg University, Heidelberg, Germany r.eils@dkfz.de c.conrad@dkfz.de.
Abstract

Three-dimensional protein localization intricately determines the functional coordination of cellular processes. The complex spatial context of protein landscape has been assessed by multiplexed immunofluorescent staining or mass spectrometry, applied to 2D Cell Culture with limited physiological relevance or tissue sections. Here, we present 3D SPECS, an automated technology for 3D Spatial characterization of Protein Expression Changes by microscopic Screening. This workflow comprises iterative antibody staining, high-content 3D imaging, and machine learning for detection of mitoses. This is followed by mapping of spatial protein localization into a spherical, cellular coordinate system, a basis for model-based prediction of spatially resolved affinities of proteins. As a proof-of-concept, we mapped twelve epitopes in 3D-cultured spheroids and investigated the network effects of twelve mitotic Cancer drugs. Our approach reveals novel insights into spindle fragility and chromatin stress, and predicts unknown interactions between proteins in specific mitotic pathways. 3D SPECS's ability to map potential drug targets by multiplexed immunofluorescence in 3D Cell Culture combined with our automated high-content assay will inspire future functional protein expression and drug assays.

Keywords

cell profiling; mitosis modeling; multiplexed immunostaining; protein–protein interactions.

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