Spatial-proteomics reveal in-vivo phospho-signaling dynamics at subcellular resolution

Ana Martínez‐Val; Dorte B. Bekker‐Jensen; Sophia Steigerwald; Claire Koenig; Ole Østergaard; Adi Mehta; Trung The Tran; Krzysztof Sikorski; Estefanía Torres-Vega; Ewa Kwasniewicz; Sólveig Hlín Brynjólfsdóttir; Lisa B. Frankel; Rasmus Kjøbsted; Nicolai Krogh; Alicia Lundby; Simon Bekker‐Jensen; Fridtjof Lund‐Johansen; Jesper  Velgaard Olsen

doi:10.1038/s41467-021-27398-y

Systems BiologyNature CommunicationsDec 7, 2021

Peer-reviewed journal article

Spatial-proteomics reveal in-vivo phospho-signaling dynamics at subcellular resolution

Ana Martínez‐Val iD, Dorte B. Bekker‐Jensen iD, Sophia Steigerwald iD, Claire Koenig iD, Ole Østergaard iD, Adi Mehta, Trung The Tran iD, Krzysztof Sikorski, Estefanía Torres-Vega iD, Ewa Kwasniewicz, Sólveig Hlín Brynjólfsdóttir iD, Lisa B. Frankel iD, Rasmus Kjøbsted iD, Nicolai Krogh iD, Alicia Lundby iD, Simon Bekker‐Jensen iD, Fridtjof Lund‐Johansen iD, Jesper Velgaard Olsen iD

Abstract

Dynamic change in subcellular localization of signaling proteins is a general concept that eukaryotic cells evolved for eliciting a coordinated response to stimuli. Mass spectrometry (MS)-based proteomics in combination with subcellular fractionation can provide comprehensive maps of spatio-temporal regulation of cells, but involves laborious workflows that does not cover the phospho-proteome level. Here we present a high-throughput workflow based on sequential cell fractionation to profile the global proteome and phospho-proteome dynamics across six distinct subcellular fractions. We benchmarked the workflow by studying spatio-temporal EGFR phospho-signaling dynamics in-vitro in HeLa cells and in-vivo in mouse tissues. Finally, we investigated the spatio-temporal stress signaling, revealing cellular relocation of ribosomal proteins in response to hypertonicity and muscle contraction. Proteomics data generated in this study can be explored through https://SpatialProteoDynamics.github.io.

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