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Spatial organization of the nucleolar proteome during mitosis
KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science, Cellular and Clinical Proteomics. KTH, Centres, Science for Life Laboratory, SciLifeLab.ORCID iD: 0000-0002-2387-3491
KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science, Cellular and Clinical Proteomics.ORCID iD: 0000-0003-0750-1070
KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science, Cellular and Clinical Proteomics. KTH, Centres, Science for Life Laboratory, SciLifeLab.ORCID iD: 0000-0001-6566-3559
KTH, Centres, Science for Life Laboratory, SciLifeLab. KTH, School of Engineering Sciences in Chemistry, Biotechnology and Health (CBH), Protein Science, Cellular and Clinical Proteomics.ORCID iD: 0000-0002-6107-1465
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(English)Manuscript (preprint) (Other academic)
Abstract [en]

In the interphase cell, the membrane-less nucleoli are the sites of ribosome biogenesis. As part of the Human Protein Atlas we created an image catalogue comprising 1,314 nucleolar proteins using antibody-based proteomics. We show experimental evidence for 1,027 proteins localizing to the whole nucleoli and 287 to the fibrillar center or dense fibrillar component. We also propose a new sub-compartment located in the nucleoplasmic border denoted as nucleoli rim, comprising at least 131 proteins. As a step toward better understanding of nucleolar protein function during cell division, we additionally generated confocal images of 68 nucleolar proteins being recruited to the chromosomal periphery in mitosis. Thanks to the single cell resolution we were able to define three expression phenotypes among the mitotic chromosome proteins; early, intermediate and late recruitment suggesting phase specific functions. We also for the first time provide a proteome-wide confirmation that the nucleoli in general, but mitotic chromosome proteins in particular have a higher predicted intrinsic disorder level compared to cytoplasmic proteins, indicating that the perichromosomal layer indeed is a liquid-like layer.

Keywords [en]
Spatial proteomics, Human Protein Atlas, Immunofluorescence, Nucleolar proteome, Intrinsic disorder, chromosomal periphery
National Category
Cell and Molecular Biology Natural Sciences
Identifiers
URN: urn:nbn:se:kth:diva-261238OAI: oai:DiVA.org:kth-261238DiVA, id: diva2:1357413
Funder
Science for Life Laboratory - a national resource center for high-throughput molecular bioscience
Note

QC 20191007

Available from: 2019-10-03 Created: 2019-10-03 Last updated: 2019-10-07Bibliographically approved
In thesis
1. Spatiotemporal characterization of the human proteome
Open this publication in new window or tab >>Spatiotemporal characterization of the human proteome
2019 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Characterizing the molecular components of the basic unit of life; the cell, is crucial for a complete understanding of human biology. The cell is divided into compartments to create a suitable environment for the resident proteins to fulfill their functions. Therefore, spatial mapping of the human proteome is essential to understand protein function in health and disease.

 

Spatial proteomics is most commonly investigated using mass spectrometry or imaging, combined with machine learning for the data analysis. Until now, studies have been limited to high abundant proteins and relied on the purification of organelle fractions from a bulk of cells. Within the scope of this thesis, we were able to systematically localize proteins in their native cellular environment using antibody-based imaging techniques, and to investigate protein subcellular localization and dynamics on a single cell level, introducing a major advance within the field of spatial proteomics.

 

Paper I of this thesis presents a subcellular map of the human proteome, where the spatial distribution of 12,003 human proteins was mapped into 30 subcellular structures, half of which were not previously localized. Besides providing a valuable dataset for cell biology, this study is the first to reveal the spatial complexity of human cells with proteins localizing to multiple compartments and pronounced single cell variations. Paper II reports on the systematic temporal dissection of these single cell variations and the identification of cell cycle correlated variations. We identified 258 novel cell cycle regulated proteins and showed that several of these proteins may be connected to proliferative diseases. A key finding of Paper II is that proteins showing non-cell cycle dependent variations are significantly enriched in mitochondria, whereas cell cycle dependent proteins are enriched in nucleoli. In Paper III and IV, we spatiotemporally characterized the proteomes of these two organelles, mitochondria and nucleoli, in greater detail.

In Paper III, we expanded the mitochondrial proteome with 560 novel proteins. As many as 20% of the mitochondrial proteome showed variations in their expression pattern at the single cell level, most often independent of the cell cycle. Paper IV provides a complete characterization of the nucleolar proteome. Nucleoli are not only important for ribosome synthesis and assembly, but are also crucial for cell cycle regulation through the recruitment of its proteins to the chromosomal periphery during cell division. Here, we presented the first proteome-wide spatiotemporal analysis of the nucleolus with its sub-compartments, and identified 69 nucleolar proteins that relocated to the chromosomes periphery during mitosis.

 

In conclusion, this thesis unravels the spatiotemporal proteome organization of the human cell over the course of a cell cycle and offers a valuable starting point for a better understanding of human cell biology in health and disease.

Place, publisher, year, edition, pages
Stockholm, Sweden: KTH Royal Institute of Technology, 2019. p. 65
Series
TRITA-CBH-FOU ; 2019:48
Keywords
Spatial proteomics, Spatiotemporal proteomics, Immunofluorescence, Human Protein Atlas, Cell compartments, Single cell proteomics, Cell cycle, Cancer
National Category
Cell and Molecular Biology Biological Sciences
Identifiers
urn:nbn:se:kth:diva-261245 (URN)978-91-7873-302-6 (ISBN)
Public defence
2019-10-25, Atrium, Nobels väg 12B, Wargentinhuset, solna, 09:00 (English)
Opponent
Supervisors
Funder
Science for Life Laboratory - a national resource center for high-throughput molecular bioscience
Note

QC 2019-10-04

Available from: 2019-10-04 Created: 2019-10-03 Last updated: 2019-10-04Bibliographically approved

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Åkesson, LovisaMahdessian, DianaGnann, ChristianThul, PeterLundberg, Emma

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