Dyakov, B. J. A. | bioRxiv Science

bioRxiv2024

Spatial proteomic mapping of human nuclear bodies reveals new functional insights into RNA regulation

Nuclear bodies are diverse membraneless suborganelles with emerging links to development and disease. Explaining their structure, function, regulation, and implications in human health will require understanding their protein composition; however, isolating nuclear bodies for proteomic analysis remains challenging. We present the first comprehensive proximity proteomics-based map of nuclear bodies, featuring 140 bait proteins (encoded by 119 genes) and 1,816 unique prey proteins. We identified 641 potential nuclear body components, including 131 paraspeckle proteins and 147 nuclear speckle proteins. After validating 31 novel paraspeckle and nuclear speckle components, we discovered regulatory functions for the poorly characterised nuclear speckle- and RNA export-associated proteins PAXBP1, PPIL4, and C19ORF47, and revealed that QKI regulates paraspeckle size. This work provides a systematic framework of nuclear body composition in live cells that will accelerate future research into their organisation and roles in human health and disease.

cell biology↗

bioRxiv2022

Comprehensive Interactome Mapping of the DNA Repair Scaffold SLX4 using Proximity Labeling and Affinity Purification

The DNA repair scaffold SLX4 has pivotal roles in cellular processes that maintain genome stability, most notably homologous recombination. Germline mutations in SLX4 are associated with Fanconi anemia, a disease characterized by chromosome instability and cancer susceptibility. The role of mammalian SLX4 in homologous recombination depends critically on binding and activating structure-selective endonucleases, namely SLX1, MUS81-EME1, and XPF-ERCC1. Increasing evidence indicates that cells rely on distinct SLX4-dependent complexes to remove DNA lesions in specific regions of the genome. Despite our understanding of SLX4 as a scaffold for DNA repair proteins, a detailed repertoire of SLX4 interactors has never been reported. Here, we provide the first comprehensive map of the human SLX4 interactome using proximity-dependent biotin identification (BioID) and affinity purification coupled to mass spectrometry (AP-MS). We identified 237 high-confidence interactors, of which the vast majority represent novel SLX4 binding proteins. Network analysis of these hits revealed pathways with known involvement of SLX4, such as DNA repair, and novel or emerging pathways of interest, including RNA metabolism and chromatin remodeling. In summary, the comprehensive SLX4 interactome we report here provides a deeper understanding of how SLX4 functions in DNA repair while revealing new cellular processes that may involve SLX4.

biochemistry↗