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| Open AccessMimicked synthetic ribosomal protein complex for benchmarking crosslinking mass spectrometry workflows
Cross-linking mass spectrometry is widely used to elucidate protein structures and interactions. Here, the authors generate an extensive peptide library to benchmark the most common cross-link search engines with frequently used cross-linking reagents in low and high complex sample systems.
- Manuel Matzinger
- , Adrian Vasiu
- & Karl Mechtler
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Article
| Open AccessEndosomal LC3C-pathway selectively targets plasma membrane cargo for autophagic degradation
Autophagy can selectively target cargo for degradation. Here the authors map the proximal interactome of ATG8-paralogs LC3B and LC3C uncovering an LC3C-Endocytic-Associated-Pathway that selectively recruits internalized plasma membrane cargo, Met and transferrin receptors, to nascent autophagosomes.
- Paula P. Coelho
- , Geoffrey G. Hesketh
- & Morag Park
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Article
| Open AccessProteogenomic characterization of 2002 human cancers reveals pan-cancer molecular subtypes and associated pathways
Pan-cancer proteomics analysis enables the analysis of protein expression across multiple cancer types. Here, the authors compare proteomics from 14 cancer types and show 11 distinct subtypes across multiple cancer types. Proteome data could link higher pathway activity levels with somatic alteration of specific genes in the pathway.
- Yiqun Zhang
- , Fengju Chen
- & Chad J. Creighton
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Article
| Open AccessPhase separation and zinc-induced transition modulate synaptic distribution and association of autism-linked CTTNBP2 and SHANK3
Autism impacts synapses. This study reports that autism-linked mutations of CTTNBP2 regulate phase separation to control synaptic enrichment of that protein. A zinc-induced liquid-to-gel transition improves synaptic retention of CTTNBP2 and SHANK3.
- Pu-Yun Shih
- , Yu-Lun Fang
- & Yi-Ping Hsueh
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Article
| Open AccessHuman transcription factor protein interaction networks
Transcription factors (TFs) interact with several other proteins in the process of transcriptional regulation. Here the authors identify 6703 and 1536 protein–protein interactions for 109 different human TFs through BioID and AP-MS analyses, respectively.
- Helka Göös
- , Matias Kinnunen
- & Markku Varjosalo
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Article
| Open AccessLight-mediated discovery of surfaceome nanoscale organization and intercellular receptor interaction networks
The spatial organization of cell surface receptors is critical for cell signaling and drug action. Here, the authors develop an optoproteomic method for mapping surface protein interactions, revealing cellular responses to antibodies, drugs and viral particles as well as immunosynapse signaling events.
- Maik Müller
- , Fabienne Gräbnitz
- & Bernd Wollscheid
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Article
| Open AccessLarge scale discovery of coronavirus-host factor protein interaction motifs reveals SARS-CoV-2 specific mechanisms and vulnerabilities
Many interactions between viral and host proteins are mediated by short peptide motifs. Here, using a phage-based viral peptide library, the authors identify 269 peptide-based interactions for 18 coronaviruses, including an interaction between SARS-CoV-2 N and G3BP1/2 that affects stress granules.
- Thomas Kruse
- , Caroline Benz
- & Ylva Ivarsson
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Article
| Open AccessIdentification of proximal SUMO-dependent interactors using SUMO-ID
Several proteomic approaches allow the analysis of covalent protein SUMOylation, but it remains challenging to systematically study the consequences of a substrate being modified. Here, the authors combine proximity biotinylation and protein-fragment complementation to identify SUMO-dependent protein interactors.
- Orhi Barroso-Gomila
- , Fredrik Trulsson
- & James D. Sutherland
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Article
| Open AccessOff-the-shelf proximity biotinylation for interaction proteomics
Proximity biotinylation is a powerful tool to profile interactomes, but it requires genetic engineering of the target protein. Here, the authors develop a proximity biotinylation enzyme that can be directed to the target using antibodies, enabling interactome profiling of endogenous proteins or PTMs.
- Irene Santos-Barriopedro
- , Guido van Mierlo
- & Michiel Vermeulen
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Article
| Open AccessPERM1 interacts with the MICOS-MIB complex to connect the mitochondria and sarcolemma via ankyrin B
Mitochondria in skeletal muscle have distinct localization and properties through unclear mechanisms. Here, the authors use complexome profiling and immunoprecipitations to identify PERM1 as a MICOS-MIB complex interactor that also binds ankyrin B, suggesting PERM1 directs the mitochondria to the membrane.
- Theresa Bock
- , Clara Türk
- & Marcus Krüger
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Article
| Open AccessLsm12 is an NAADP receptor and a two-pore channel regulatory protein required for calcium mobilization from acidic organelles
Nicotinic acid adenine dinucleotide phosphate (NAADP) potent Ca2+ mobilizing second messenger which uniquely triggers Ca2+ release from acidic endolysosomal organelles. Here the authors identify Lsm12 as an NAADP receptor essential for NAADP-evoked Ca2+ release from lysosomes via NAADP binding on its Lsm domain.
- Jiyuan Zhang
- , Xin Guan
- & Jiusheng Yan
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Article
| Open AccessSystematic detection of functional proteoform groups from bottom-up proteomic datasets
Many proteins exist in various proteoforms but detecting these variants by bottom-up proteomics remains difficult. Here, the authors present a computational approach based on peptide correlation analysis to identify and characterize proteoforms from bottom-up proteomics data.
- Isabell Bludau
- , Max Frank
- & Ruedi Aebersold
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Article
| Open AccessReliable identification of protein-protein interactions by crosslinking mass spectrometry
Cross-linking mass spectrometry (MS) can identify protein-protein interaction (PPI) networks but assessing the reliability of these data remains challenging. To address this issue, the authors develop and validate a method to determine the false-discovery rate of PPIs identified by cross-linking MS.
- Swantje Lenz
- , Ludwig R. Sinn
- & Juri Rappsilber
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Article
| Open AccessSite-specific ubiquitylation acts as a regulator of linker histone H1
While the role of specific posttranslational modifications (PTMs) is increasingly well understood for core histones, this is not the case for linker histone H1. Here the authors show that site-specific ubiquitylation of H1 results in distinct interactomes, regulates phase separation, and modulates assembly of chromatosomes.
- Eva Höllmüller
- , Simon Geigges
- & Florian Stengel
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Article
| Open AccessRetention time prediction using neural networks increases identifications in crosslinking mass spectrometry
Predicting chromatographic retention times (RTs) has proven beneficial in proteomics but has not yet been achieved for crosslinked peptides. Here, the authors develop an RT prediction tool for crosslinked peptides and leverage predicted RTs to increase identifications in crosslinking mass spectrometry studies.
- Sven H. Giese
- , Ludwig R. Sinn
- & Juri Rappsilber
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Article
| Open AccessAnalysis of Zika virus capsid-Aedes aegypti mosquito interactome reveals pro-viral host factors critical for establishing infection
Here the authors develop mosquito cell lines expressing Zika virus (ZIKV) capsid and perform proteomics experiments retrieving 157 protein interactors, of which they show 8 to act as pro-viral factors, showing the transitional endoplasmic reticulum 94 (TER94) and its human ortholog VCP target ZIKV capsid to proteasomal degradation to facilitate infection.
- Rommel J. Gestuveo
- , Jamie Royle
- & Margus Varjak
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Article
| Open AccessGenoppi is an open-source software for robust and standardized integration of proteomic and genetic data
Genetic variation can impact protein complexes and interaction networks, but reconciling genetic and proteomic information remains challenging. To address this need, the authors develop Genoppi —a computational tool for integrating genetics and cell-type-specific proteomics data.
- Greta Pintacuda
- , Frederik H. Lassen
- & Kasper Lage
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Article
| Open AccessA CSB-PAF1C axis restores processive transcription elongation after DNA damage repair
The transcription-coupled repair pathway removes transcription-blocking DNA lesions, but how transcription is restored following DNA repair is not clear. Here the authors reveal that the PAF1 complex, while dispensable for the repair process, restores transcription after DNA damage.
- Diana van den Heuvel
- , Cornelia G. Spruijt
- & Martijn S. Luijsterburg
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Article
| Open AccessCross-linking mass spectrometry uncovers protein interactions and functional assemblies in synaptic vesicle membranes
Synaptic vesicles store neurotransmitters and fuse with the pre-synaptic membrane when an action potential arrives at the nerve terminal. Here authors apply cross-linking mass spectrometry to study interactions of synaptic vesicle proteins and describe a protein network of vesicle sub-populations and functional assemblies.
- Sabine Wittig
- , Marcelo Ganzella
- & Carla Schmidt
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Article
| Open AccessSpatiotemporal profiling of cytosolic signaling complexes in living cells by selective proximity proteomics
APEX-based proximity labeling allows capturing protein interaction dynamics but its high labeling background limits its utility for cytosolic proteins. Here, the authors develop more selective proximity labeling probes, enabling the APEX-based characterization of time-resolved cytosolic protein interactomes.
- Mi Ke
- , Xiao Yuan
- & Ruijun Tian
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Perspective
| Open AccessTowards a unified open access dataset of molecular interactions
The IMEx consortium provides one of the largest resources of curated, experimentally verified molecular interaction data. Here, the authors review how IMEx evolved into a fundamental resource for life scientists and describe how IMEx data can support biomedical research.
- Pablo Porras
- , Elisabet Barrera
- & Sandra Orchard
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Article
| Open AccessSurfaceome dynamics reveal proteostasis-independent reorganization of neuronal surface proteins during development and synaptic plasticity
Cell surface proteins contribute to neuronal development and activity-dependent synaptic plasticity. Here, the authors perform a time-resolved surfaceome analysis of developing primary neurons and in response to homeostatic synaptic scaling and chemical long-term potentiation (cLTP), revealing surface proteome remodeling largely independent of global proteostasis.
- Marc van Oostrum
- , Benjamin Campbell
- & Bernd Wollscheid
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Article
| Open AccessCross-regulation of viral kinases with cyclin A secures shutoff of host DNA synthesis
Herpesviruses code for conserved protein kinases (CHPKs) that exert several regulatory functions by interacting with cellular factors. Here, the authors use affinity purification mass spectrometry (AP–MS) to identify differential interaction partners of CHPKs from seven different human herpesviruses, finding Cyclin A and associated factors as a specific signature of β-herpesvirus kinases.
- Boris Bogdanow
- , Max Schmidt
- & Lüder Wiebusch
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Article
| Open AccessMulti-layered proteomic analyses decode compositional and functional effects of cancer mutations on kinase complexes
Diseases can be associated with various mutations of the same gene, but the molecular consequences of specific mutations remain incompletely understood. Here, the authors present an integrated proteomic workflow to determine the molecular response of cells to different cancer-associated mutations of the kinase Dyrk2.
- Martin Mehnert
- , Rodolfo Ciuffa
- & Ruedi Aebersold
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Article
| Open AccessThe ataxin-1 interactome reveals direct connection with multiple disrupted nuclear transport pathways
Patients with spinocerebellar ataxia type 1 express ataxin-1 with an extended polyglutamine (polyQ) tract that forms distinctive nuclear bodies. Here, the authors characterize the cellular pathways affected by polyQ-ataxin-1, showing that it disrupts multiple nuclear transport processes.
- Sunyuan Zhang
- , Nicholas A. Williamson
- & Marie A. Bogoyevitch
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Article
| Open AccessMass spectrometry reveals the chemistry of formaldehyde cross-linking in structured proteins
Formaldehyde (FA) is a popular cross-linking reagent, but applying it for cross-linking mass spectrometry (XLMS) has been largely unsuccessful. Here, the authors show that cross-links in structured proteins are the product of two FA molecules and identify hundreds of FA cross-links by XLMS in vitro and in situ.
- Tamar Tayri-Wilk
- , Moriya Slavin
- & Nir Kalisman
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Article
| Open AccessDeconstructing sarcomeric structure–function relations in titin-BioID knock-in mice
Titin determines the elasticity of the sarcomere and integrates into both the Z-disc and the M-band. Here, the authors generate a BioID mouse to study the titin interactome at the Z-disc region in neonatal and adult heart and skeletal muscle.
- Franziska Rudolph
- , Claudia Fink
- & Michael Gotthardt
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Article
| Open AccessSplit Intein-Mediated Protein Ligation for detecting protein-protein interactions and their inhibition
Protein-protein interactions are fundamental to the regulation of protein activity and cellular phyisology. Here the authors present Split Intein-Mediated Protein Ligation, which uses bait and prey proteins fused to intein fragments to generate single intact proteins upon interaction.
- Zhong Yao
- , Farzaneh Aboualizadeh
- & Igor Stagljar
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Article
| Open AccessProteasome inhibitor-induced modulation reveals the spliceosome as a specific therapeutic vulnerability in multiple myeloma
The mechanisms of action of proteasome inhibitors (PI) in multiple myeloma (MM) treatment are not fully elucidated. Here, the authors use unbiased phosphoproteomics in PI-treated MM and show increased phosphorylation of splicing-associated proteins, ultimately revealing splicing interference as a mode of PI action as well as demonstrating the spliceosome as a specific therapeutic vulnerability in this disease.
- Hector H. Huang
- , Ian D. Ferguson
- & Arun P. Wiita
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Article
| Open AccessA common MET polymorphism harnesses HER2 signaling to drive aggressive squamous cell carcinoma
The MET receptor is frequently activated in cancer. Here, the authors show that in head and neck and lung squamous carcinoma, a polymorphic MET variant enhances binding to HER2, resulting in activation of HER2 signalling and progression of the cancers.
- Li Ren Kong
- , Nur Afiqah Binte Mohamed Salleh
- & Boon Cher Goh
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Article
| Open AccessMulti-level and lineage-specific interactomes of the Hox transcription factor Ubx contribute to its functional specificity
Many transcription factors regulate gene expression in a lineage- and process-specific manner, despite being expressed in several cell types. Here, the authors show that the Hox transcription factor Ubx has lineage-specific interactomes, which contribute to its cell context-dependent functions.
- Julie Carnesecchi
- , Gianluca Sigismondo
- & Ingrid Lohmann
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Article
| Open AccessA synthetic peptide library for benchmarking crosslinking-mass spectrometry search engines for proteins and protein complexes
Validating crosslinking-mass spectrometry workflows is hampered by the lack of a ground truth to assess the robustness of the crosslink identifications. Here, the authors present a synthetic library of crosslinked peptides, enabling unambiguous discrimination of correct and incorrect crosslink identifications.
- Rebecca Beveridge
- , Johannes Stadlmann
- & Karl Mechtler
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Article
| Open AccessExtensive rewiring of the EGFR network in colorectal cancer cells expressing transforming levels of KRASG13D
Kras is often mutated in colorectal cancer but how this oncogenic mutation alters signalling pathways globally is undetermined. Here, the authors analyse how this mutation affects protein interaction networks and signal flow showing an extensive re‐wiring of signalling in response to KRas mutation
- Susan A. Kennedy
- , Mohamed-Ali Jarboui
- & Walter Kolch
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Article
| Open AccessThe RepID–CRL4 ubiquitin ligase complex regulates metaphase to anaphase transition via BUB3 degradation
The spindle assembly checkpoint (SAC) safeguards chromosome segregation by regulating the anaphase promoting complex/cyclosome (APC/C), allowing chromosomes to correctly attach to mitotic spindles. Here the authors reveal a role for Cullin–RING ubiquitin ligase complex 4 (CRL4) in regulating metaphase to anaphase transition via BUB3 degradation.
- Sang-Min Jang
- , Jenny F. Nathans
- & Mirit I. Aladjem
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Article
| Open AccessRegulation of the ER stress response by a mitochondrial microprotein
Cells trigger an unfolded protein response (UPR) in the endoplasmic reticulum, but its regulation by mitochondria is unclear. Here, the authors report a 54-residue microprotein PIGBOS that participates in inter-organelle contact between the endoplasmic reticulum and the mitochondria and may regulate UPR.
- Qian Chu
- , Thomas F. Martinez
- & Alan Saghatelian
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Article
| Open AccessThe midbody interactome reveals unexpected roles for PP1 phosphatases in cytokinesis
The midbody is an organelle present at the bridge connecting two cells at the end of cell division. Here, the authors use mass spectrometry to define the midbody interactome and uncover a role for PP1 phosphatases in microtubule dynamics and regulation of cytokinesis.
- Luisa Capalbo
- , Zuni I. Bassi
- & Pier Paolo D’Avino
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Article
| Open AccessExtensive disruption of protein interactions by genetic variants across the allele frequency spectrum in human populations
Low frequency coding single-nucleotide variants (SNVs) are predicted to disproportionately affect protein function. Here, the authors evaluate 2,009 missense SNVs across 2,185 protein-protein interactions using yeast two-hybrid and protein complementation assays and find that disruptive SNVs often occur in disease-associated genes.
- Robert Fragoza
- , Jishnu Das
- & Haiyuan Yu
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Article
| Open AccessImproving mass spectrometry analysis of protein structures with arginine-selective chemical cross-linkers
Cross-linking mass spectrometry can provide insights into protein structures and interactions but its scope depends on the reactivity of the cross-linker. Here, the authors develop Arg-Arg and Lys-Arg cross-linkers, which provide structural information elusive to the widely used Lys-Lys cross-linkers.
- Alexander X. Jones
- , Yong Cao
- & Meng-Qiu Dong
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Article
| Open AccessMaximizing binary interactome mapping with a minimal number of assays
Comprehensive mapping of binary protein-protein interactions requires to combine several complementary assays. Here, the authors show that complete coverage could be reached with a minimal number of assays as long as they explore various experimental conditions.
- Soon Gang Choi
- , Julien Olivet
- & Yves Jacob
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Article
| Open AccessA quantitative Streptococcus pyogenes–human protein–protein interaction map reveals localization of opsonizing antibodies
Characterizing host-pathogen protein interactions can help elucidate the molecular basis of bacterial infections. Here, the authors use an integrative proteomics approach to generate a quantitative map of protein interactions between Streptococcus pyogenes and human saliva and plasma.
- Lotta Happonen
- , Simon Hauri
- & Johan Malmström
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Article
| Open AccessMiniaturised interaction proteomics on a microfluidic platform with ultra-low input requirements
Affinity purification-mass spectrometry (AP-MS) can identify endogenous protein interactions but the need for high amounts of input material still limits its applicability. Here, the authors present a microfluidic-based AP-MS workflow that can capture protein interactions from 50─100-fold less input material than conventional approaches.
- Cristina Furlan
- , René A. M. Dirks
- & Michiel Vermeulen
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Article
| Open AccessNetwork-based prediction of protein interactions
Computational protein-protein interaction (PPI) prediction has the potential to complement experimental efforts to map interactomes. Here, the authors show that proteins tend to interact if one is similar to the other’s partners and that PPI prediction based on this principle is highly accurate.
- István A. Kovács
- , Katja Luck
- & Albert-László Barabási
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Article
| Open AccessRapid determination of quaternary protein structures in complex biological samples
Protein structure determination in complex biological samples is still challenging. Here, the authors develop a computational modeling-guided cross-linking mass spectrometry method, obtaining a high-resolution model of a 1.8 MDa protein assembly from cross-links detected in a mixture of human plasma and bacteria.
- Simon Hauri
- , Hamed Khakzad
- & Lars Malmström
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Article
| Open AccessA FRET biosensor for necroptosis uncovers two different modes of the release of DAMPs
Necroptotic cells activate MLKL and release inflammatory DAMPs, although the underlying regulatory mechanisms of this process are poorly understood. Here, Murai et al. develop a necroptosis-specific FRET sensor (SMART) that monitors MLKL membrane translocation to identify two modes of DAMP release.
- Shin Murai
- , Yoshifumi Yamaguchi
- & Hiroyasu Nakano
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Article
| Open AccessSlc7a5 regulates Kv1.2 channels and modifies functional outcomes of epilepsy-linked channel mutations
Kv1.2 is a voltage-gated potassium channel that influences action potential generation and propagation in the central nervous system. Here authors use electrophysiology and find that Slc7a5, a neutral amino acid transporter, has a profound impact on Kv1.2.
- Victoria A. Baronas
- , Runying Y. Yang
- & Harley T. Kurata
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Article
| Open AccessInterrogating the protein interactomes of RAS isoforms identifies PIP5K1A as a KRAS-specific vulnerability
RAS isoforms are frequently mutated in cancer but their inhibition remains challenging. By comparing the protein interactomes of the highly similar isoforms HRAS, NRAS and KRAS, the authors here identify PIP5K1A as a KRAS-specific interactor and a target to inhibit KRAS-driven cell growth.
- Hema Adhikari
- & Christopher M. Counter
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Article
| Open AccessThe replication initiation determinant protein (RepID) modulates replication by recruiting CUL4 to chromatin
RepID has previously been shown to promote origin firing. Here the authors reveal that RepID regulates replication origins via the recruitment of the CRL4 complex, and prevents re-initiation and unscheduled DNA replication.
- Sang-Min Jang
- , Ya Zhang
- & Mirit I. Aladjem
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Article
| Open AccessA quantitative mass spectrometry-based approach to monitor the dynamics of endogenous chromatin-associated protein complexes
Chromatin-associated protein complexes play a critical role in the regulation of gene expression in health and disease. Here, the authors describe a sensitive mass spectrometry-based method to monitor the dynamic interactions of endogenous chromatin-associated protein complexes in clinical samples.
- Evangelia K. Papachristou
- , Kamal Kishore
- & Jason S. Carroll
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Article
| Open AccessAn AP-MS- and BioID-compatible MAC-tag enables comprehensive mapping of protein interactions and subcellular localizations
AP-MS and BioID provide complementary insights into cellular protein interaction networks. To facilitate their combined use, the authors here present an AP-MS- and BioID-compatible affinity tag, enabling efficient determination of cellular protein locations and interaction distances.
- Xiaonan Liu
- , Kari Salokas
- & Markku Varjosalo