Faculty Opinions recommendation of Off-the-shelf proximity biotinylation for interaction proteomics.

Rab11 GTPase proteins are required for cytokinesis, ciliogenesis, and lumenogenesis. Rab11a is critical for apical delivery of podocalyxin (PODXL) during lumen formation in epithelial cells. SH3BP5 and SH3BP5L are guanine nucleotide exchange factors (GEFs) for Rab11. We show that SH3BP5 and SH3BP5L are required for activation of Rab11a and cyst lumen formation. Using proximity-dependent biotin identification (BioID) interaction proteomics, we have identified SH3BP5 and its paralogue SH3BP5L as new substrates of the poly-ADP-ribose polymerase Tankyrase and the E3 ligase RNF146. We provide data demonstrating that epithelial polarity via cyst lumen formation is governed by Tankyrase, which inhibits Rab11a activation through the suppression of SH3BP5 and SH3BP5L. RNF146 reduces Tankyrase protein abundance and restores Rab11a activation and lumen formation. Thus, Rab11a activation is controlled by a signaling pathway composed of the sequential inhibition of SH3BP5 paralogues by Tankyrase, which is itself suppressed by RNF146.

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New Insights on Estrogen Receptor Actions in Hormone-Responsive Breast Cancer Cells by Interaction Proteomics

Cell and Molecular Biology of Breast Cancer ◽

10.1007/978-1-62703-634-4_8 ◽

2013 ◽

pp. 149-174 ◽

Cited By ~ 1

Author(s):

Concetta Ambrosino ◽

Roberta Tarallo ◽

Giovanni Nassa ◽

Francesca Cirillo ◽

Alessandro Weisz

Keyword(s):

Breast Cancer ◽

Estrogen Receptor ◽

Cancer Cells ◽

Breast Cancer Cells ◽

Interaction Proteomics

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Interaction Proteomics Identifies ERbeta Association with Chromatin Repressive Complexes to Inhibit Cholesterol Biosynthesis and Exert An Oncosuppressive Role in Triple-negative Breast Cancer

Molecular & Cellular Proteomics ◽

10.1074/mcp.ra119.001817 ◽

2019 ◽

Vol 19 (2) ◽

pp. 245-260 ◽

Cited By ~ 6

Author(s):

Elena Alexandrova ◽

Giorgio Giurato ◽

Pasquale Saggese ◽

Giovanni Pecoraro ◽

Jessica Lamberti ◽

...

Keyword(s):

Breast Cancer ◽

Gene Regulation ◽

Chromatin Remodeling ◽

Triple Negative Breast Cancer ◽

Triple Negative ◽

Cholesterol Biosynthesis ◽

Poor Response ◽

Response To Therapy ◽

Protein Partners ◽

Interaction Proteomics

Triple-negative breast cancer (TNBC) is characterized by poor response to therapy and low overall patient survival. Recently, Estrogen Receptor beta (ERβ) has been found to be expressed in a fraction of TNBCs where, because of its oncosuppressive actions on the genome, it represents a potential therapeutic target, provided a better understanding of its actions in these tumors becomes available. To this end, the cell lines Hs 578T, MDA-MB-468 and HCC1806, representing the claudin-low, basal-like 1 and 2 TNBC molecular subtypes respectively, were engineered to express ERβ under the control of a Tetracycline-inducible promoter and used to investigate the effects of this transcription factor on gene activity. The antiproliferative effects of ERβ in these cells were confirmed by multiple functional approaches, including transcriptome profiling and global mapping of receptor binding sites in the genome, that revealed direct negative regulation by ERβ of genes, encoding for key components of cellular pathways associated to TNBC aggressiveness representing novel therapeutic targets such as angiogenesis, invasion, metastasis and cholesterol biosynthesis. Supporting these results, interaction proteomics by immunoprecipitation coupled to nano LC-MS/MS mass spectrometry revealed ERβ association with several potential nuclear protein partners, including key components of regulatory complexes known to control chromatin remodeling, transcriptional and post-transcriptional gene regulation and RNA splicing. Among these, ERβ association with the Polycomb Repressor Complexes 1 and 2 (PRC1/2), known for their central role in gene regulation in cancer cells, was confirmed in all three TNBC subtypes investigated, suggesting its occurrence independently from the cellular context. These results demonstrate a significant impact of ERβ in TNBC genome activity mediated by its cooperation with regulatory multiprotein chromatin remodeling complexes, providing novel ground to devise new strategies for the treatment of these diseases based on ligands affecting the activity of this nuclear receptor or some of its protein partners.

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Peptide-based Interaction Proteomics

Molecular & Cellular Proteomics ◽

10.1074/mcp.r120.002034 ◽

2020 ◽

Vol 19 (7) ◽

pp. 1070-1075 ◽

Cited By ~ 3

Author(s):

Katrina Meyer ◽

Matthias Selbach

Keyword(s):

Protein Interactions ◽

Synthetic Peptides ◽

Protein Protein Interactions ◽

Intrinsically Disordered ◽

Intrinsically Disordered Regions ◽

Interaction Partners ◽

Linear Motifs ◽

Cellulose Membranes ◽

Interaction Proteomics ◽

Proteins Interactions

Protein-protein interactions are often mediated by short linear motifs (SLiMs) that are located in intrinsically disordered regions (IDRs) of proteins. Interactions mediated by SLiMs are notoriously difficult to study, and many functionally relevant interactions likely remain to be uncovered. Recently, pull-downs with synthetic peptides in combination with quantitative mass spectrometry emerged as a powerful screening approach to study protein-protein interactions mediated by SLiMs. Specifically, arrays of synthetic peptides immobilized on cellulose membranes provide a scalable means to identify the interaction partners of many peptides in parallel. In this minireview we briefly highlight the relevance of SLiMs for protein-protein interactions, outline existing screening technologies, discuss unique advantages of peptide-based interaction screens and provide practical suggestions for setting up such peptide-based screens.

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Epigenetic reader complexes of the human malaria parasite, Plasmodium falciparum

Nucleic Acids Research ◽

10.1093/nar/gkz1044 ◽

2019 ◽

Vol 47 (22) ◽

pp. 11574-11588 ◽

Cited By ~ 9

Author(s):

Wieteke Anna Maria Hoeijmakers ◽

Jun Miao ◽

Sabine Schmidt ◽

Christa Geeke Toenhake ◽

Sony Shrestha ◽

...

Keyword(s):

Plasmodium Falciparum ◽

Protein Complexes ◽

Human Malaria Parasite ◽

Phd Finger ◽

Finger Protein ◽

Binding Preference ◽

Parasite Plasmodium ◽

Parasite Plasmodium Falciparum ◽

Eukaryotic Organisms ◽

Interaction Proteomics

Abstract Epigenetic regulatory mechanisms are central to the development and survival of all eukaryotic organisms. These mechanisms critically depend on the marking of chromatin domains with distinctive histone tail modifications (PTMs) and their recognition by effector protein complexes. Here we used quantitative proteomic approaches to unveil interactions between PTMs and associated reader protein complexes of Plasmodium falciparum, a unicellular parasite causing malaria. Histone peptide pull-downs with the most prominent and/or parasite-specific PTMs revealed the binding preference for 14 putative and novel reader proteins. Amongst others, they highlighted the acetylation-level-dependent recruitment of the BDP1/BDP2 complex and identified an PhD-finger protein (PHD 1, PF3D7_1008100) that could mediate a cross-talk between H3K4me2/3 and H3K9ac marks. Tagging and interaction proteomics of 12 identified proteins unveiled the composition of 5 major epigenetic complexes, including the elusive TBP-associated-factor complex as well as two distinct GCN5/ADA2 complexes. Furthermore, it has highlighted a remarkable degree of interaction between these five (sub)complexes. Collectively, this study provides an extensive inventory of PTM-reader interactions and composition of epigenetic complexes. It will not only fuel further explorations of gene regulation amongst ancient eukaryotes, but also provides a stepping stone for exploration of PTM-reader interactions for antimalarial drug development.

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