| 1. |
- Alvez, Maria Bueno, et al.
(författare)
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Next generation pan-cancer blood proteome profiling using proximity extension assay
- 2023
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Ingår i: Nature Communications. - : Springer Nature. - 2041-1723. ; 14:1
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Tidskriftsartikel (refereegranskat)abstract
- A comprehensive characterization of blood proteome profiles in cancer patients can contribute to a better understanding of the disease etiology, resulting in earlier diagnosis, risk stratification and better monitoring of the different cancer subtypes. Here, we describe the use of next generation protein profiling to explore the proteome signature in blood across patients representing many of the major cancer types. Plasma profiles of 1463 proteins from more than 1400 cancer patients are measured in minute amounts of blood collected at the time of diagnosis and before treatment. An open access Disease Blood Atlas resource allows the exploration of the individual protein profiles in blood collected from the individual cancer patients. We also present studies in which classification models based on machine learning have been used for the identification of a set of proteins associated with each of the analyzed cancers. The implication for cancer precision medicine of next generation plasma profiling is discussed.
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| 2. |
- Thul, Peter J., et al.
(författare)
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A subcellular map of the human proteome
- 2017
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Ingår i: Science. - : American Association for the Advancement of Science. - 0036-8075 .- 1095-9203. ; 356:6340
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Tidskriftsartikel (refereegranskat)abstract
- Resolving the spatial distribution of the human proteome at a subcellular level can greatly increase our understanding of human biology and disease. Here we present a comprehensive image-based map of subcellular protein distribution, the Cell Atlas, built by integrating transcriptomics and antibody-based immunofluorescence microscopy with validation by mass spectrometry. Mapping the in situ localization of 12,003 human proteins at a single-cell level to 30 subcellular structures enabled the definition of the proteomes of 13 major organelles. Exploration of the proteomes revealed single-cell variations in abundance or spatial distribution and localization of about half of the proteins to multiple compartments. This subcellular map can be used to refine existing protein-protein interaction networks and provides an important resource to deconvolute the highly complex architecture of the human cell.
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| 3. |
- Fagerberg, Linn, et al.
(författare)
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Analysis of the human tissue-specific expression by genome-wide integration of transcriptomics and antibody-based proteomics
- 2014
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Ingår i: Molecular & Cellular Proteomics. - 1535-9476 .- 1535-9484. ; 13:2, s. 397-406
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Tidskriftsartikel (refereegranskat)abstract
- Global classification of the human proteins with regards to spatial expression patterns across organs and tissues is important for studies of human biology and disease. Here, we used a quantitative transcriptomics analysis (RNA-Seq) to classify the tissue-specific expression of genes across a representative set of all major human organs and tissues and combined this analysis with antibody- based profiling of the same tissues. To present the data, we launch a new version of the Human Protein Atlas that integrates RNA and protein expression data corresponding to 80% of the human protein-coding genes with access to the primary data for both the RNA and the protein analysis on an individual gene level. We present a classification of all human protein-coding genes with regards to tissue-specificity and spatial expression pattern. The integrative human expression map can be used as a starting point to explore the molecular constituents of the human body.
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| 4. |
- Fagerberg, Linn, et al.
(författare)
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Contribution of antibody-based protein profiling to the human chromosome-centric proteome project (C-HPP)
- 2013
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Ingår i: Journal of Proteome Research. - : American Chemical Society (ACS). - 1535-3893 .- 1535-3907. ; 12:6, s. 2439-2448
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Tidskriftsartikel (refereegranskat)abstract
- A gene-centric Human Proteome Project has been proposed to characterize the human protein-coding genes in a chromosome-centered manner to understand human biology and disease. Here, we report on the protein evidence for all genes predicted from the genome sequence based on manual annotation from literature (UniProt), antibody-based profiling in cells, tissues and organs and analysis of the transcript profiles using next generation sequencing in human cell lines of different origins. We estimate that there is good evidence for protein existence for 69% (n = 13985) of the human protein-coding genes, while 23% have only evidence on the RNA level and 7% still lack experimental evidence. Analysis of the expression patterns shows few tissue-specific proteins and approximately half of the genes expressed in all the analyzed cells. The status for each gene with regards to protein evidence is visualized in a chromosome-centric manner as part of a new version of the Human Protein Atlas (www.proteinatlas.org).
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| 5. |
- Grapotte, Mathys, et al.
(författare)
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Discovery of widespread transcription initiation at microsatellites predictable by sequence-based deep neural network
- 2021
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Ingår i: Nature Communications. - : Springer Science and Business Media LLC. - 2041-1723.
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Tidskriftsartikel (refereegranskat)abstract
- Using the Cap Analysis of Gene Expression (CAGE) technology, the FANTOM5 consortium provided one of the most comprehensive maps of transcription start sites (TSSs) in several species. Strikingly, ~72% of them could not be assigned to a specific gene and initiate at unconventional regions, outside promoters or enhancers. Here, we probe these unassigned TSSs and show that, in all species studied, a significant fraction of CAGE peaks initiate at microsatellites, also called short tandem repeats (STRs). To confirm this transcription, we develop Cap Trap RNA-seq, a technology which combines cap trapping and long read MinION sequencing. We train sequence-based deep learning models able to predict CAGE signal at STRs with high accuracy. These models unveil the importance of STR surrounding sequences not only to distinguish STR classes, but also to predict the level of transcription initiation. Importantly, genetic variants linked to human diseases are preferentially found at STRs with high transcription initiation level, supporting the biological and clinical relevance of transcription initiation at STRs. Together, our results extend the repertoire of non-coding transcription associated with DNA tandem repeats and complexify STR polymorphism.
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| 6. |
- Grapotte, M, et al.
(författare)
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Discovery of widespread transcription initiation at microsatellites predictable by sequence-based deep neural network
- 2021
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Ingår i: Nature communications. - : Springer Science and Business Media LLC. - 2041-1723. ; 12:1, s. 3297-
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Tidskriftsartikel (refereegranskat)abstract
- Using the Cap Analysis of Gene Expression (CAGE) technology, the FANTOM5 consortium provided one of the most comprehensive maps of transcription start sites (TSSs) in several species. Strikingly, ~72% of them could not be assigned to a specific gene and initiate at unconventional regions, outside promoters or enhancers. Here, we probe these unassigned TSSs and show that, in all species studied, a significant fraction of CAGE peaks initiate at microsatellites, also called short tandem repeats (STRs). To confirm this transcription, we develop Cap Trap RNA-seq, a technology which combines cap trapping and long read MinION sequencing. We train sequence-based deep learning models able to predict CAGE signal at STRs with high accuracy. These models unveil the importance of STR surrounding sequences not only to distinguish STR classes, but also to predict the level of transcription initiation. Importantly, genetic variants linked to human diseases are preferentially found at STRs with high transcription initiation level, supporting the biological and clinical relevance of transcription initiation at STRs. Together, our results extend the repertoire of non-coding transcription associated with DNA tandem repeats and complexify STR polymorphism.
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| 7. |
- Hober, Andreas, 1992-, et al.
(författare)
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Evaluation of an enhanced antibody-validation strategy for Western blot applications based on migration pattern recognition
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Annan publikation (övrigt vetenskapligt/konstnärligt)abstract
- The use of affinity reagents, such as antibodies, for studying specific molecules in complex backgrounds are some of the most powerful tools for researchers in molecular biology. However, all experiments performed using affinity reagents are directly affected by each reagent’s context-dependent ability to bind specifically to a target of interest. A growing issue with non-validated, or poorly validated affinity reagents, has been highlighted by the International Working Group for Antibody Validation (IWGAV). It has been suggested that antibodies should be evaluated in an application-specific manner since they can perform well in one application but fail to deliver reproducible results in another. One of the most commonly used antibody-based applications is the Western blot (WB) technology. When evaluating the result from a WB experiment, the initial measure used for determining whether or not the antibody binds the protein of interest is to determine the molecular weight of the protein detected by the antibody compared to a set of reference proteins. As WB relies on the SDS-PAGE for separating differently sized proteins, the comparison is actually based on protein migration during electrophoresis. It is, however, well known that the migration of a protein can differ significantly from how the reference proteins migrate. Here, we suggest a method for determining the actual migration patterns of proteins instead of relying on the theoretical molecular weight of the protein. Using this approach, called migration capture mass spectrometry (MS), a dataset containing the migration patterns of more than 39,000 protein products from more than 10,500 genes across eleven cell lines and tissues has been created. This migration capture MS approach has been validated using k-fold cross validation against 249 siRNA knockdown WBs showing that the method has a sensitivity of 96.4%, specificity of 87.4% and accuracy of 91.9%, which makes the dataset a useful resource that can facilitate antibody validation strategies in a fit-for-purpose manner. The data set has allowed the automatic evaluation of more than 12,000 antibodies in the Human Protein Atlas using the method.
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| 8. |
- Hober, Sophia, Professor, 1965-, et al.
(författare)
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Systematic evaluation of SARS-CoV-2 antigens enables a highly specific and sensitive multiplex serological COVID-19 assay
- 2021
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Ingår i: Clinical & Translational Immunology. - : Wiley. - 2050-0068. ; 10:7
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Tidskriftsartikel (refereegranskat)abstract
- Objective. The COVID-19 pandemic poses an immense need for accurate, sensitive and high-throughput clinical tests, and serological assays are needed for both overarching epidemiological studies and evaluating vaccines. Here, we present the development and validation of a high-throughput multiplex bead-based serological assay. Methods. More than 100 representations of SARS-CoV-2 proteins were included for initial evaluation, including antigens produced in bacterial and mammalian hosts as well as synthetic peptides. The five best-performing antigens, three representing the spike glycoprotein and two representing the nucleocapsid protein, were further evaluated for detection of IgG antibodies in samples from 331 COVID-19 patients and convalescents, and in 2090 negative controls sampled before 2020. Results. Three antigens were finally selected, represented by a soluble trimeric form and the S1-domain of the spike glycoprotein as well as by the C-terminal domain of the nucleocapsid. The sensitivity for these three antigens individually was found to be 99.7%, 99.1% and 99.7%, and the specificity was found to be 98.1%, 98.7% and 95.7%. The best assay performance was although achieved when utilising two antigens in combination, enabling a sensitivity of up to 99.7% combined with a specificity of 100%. Requiring any two of the three antigens resulted in a sensitivity of 99.7% and a specificity of 99.4%. Conclusion. These observations demonstrate that a serological test based on a combination of several SARS-CoV-2 antigens enables a highly specific and sensitive multiplex serological COVID-19 assay.
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| 9. |
- Häggmark, Anna, et al.
(författare)
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Classification of protein profiles from antibody microarrays using heat and detergent treatment.
- 2011
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Ingår i: New Biotechnology. - : Elsevier BV. - 1871-6784 .- 1876-4347. ; 29:5, s. 564-570
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Tidskriftsartikel (refereegranskat)abstract
- Antibody microarrays offer new opportunities for exploring the proteome and to identify biomarker candidates in human serum and plasma. Here, we have investigated the effect of heat and detergents on an antibody-based suspension bead array (SBA) assay using polyclonal antibodies and biotinylated plasma samples. With protein profiles from more than 2300 antibodies generated in 384-plex antibody SBAs, three major classes of heat and detergent susceptibility could be described. The results show that washing of the beads with SDS (rather than Tween) after target binding lowered intensity levels of basically all profiles and that about 50% of the profiles appeared to be lowered to a similar extent by heating of the sample. About 33% of the profiles appeared to be insensitive to heat treatment while another 17% showed a positive influence of heat to yield elevated profiles. The results suggest that the classification of antibodies is driven by the molecular properties of the antibody-antigen interaction and can generally not be predicted based on protein class or Western blot data. The experimental scheme presented here can be used to systematically categorize antibodies and thereby combine antibodies with similar properties into targeted arrays for analysis of plasma and serum.
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| 10. |
- Jin, Han, et al.
(författare)
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Systematic transcriptional analysis of human cell lines for gene expression landscape and tumor representation
- 2023
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Ingår i: Nature Communications. - : Springer Nature. - 2041-1723. ; 14:1, s. 5417-
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Tidskriftsartikel (refereegranskat)abstract
- Cell lines are valuable resources as model for human biology and translational medicine. It is thus important to explore the concordance between the expression in various cell lines vis-à-vis human native and disease tissues. In this study, we investigate the expression of all human protein-coding genes in more than 1,000 human cell lines representing 27 cancer types by a genome-wide transcriptomics analysis. The cell line gene expression is compared with the corresponding profiles in various tissues, organs, single-cell types and cancers. Here, we present the expression for each cell line and give guidance for the most appropriate cell line for a given experimental study. In addition, we explore the cancer-related pathway and cytokine activity of the cell lines to aid human biology studies and drug development projects. All data are presented in an open access cell line section of the Human Protein Atlas to facilitate the exploration of all human protein-coding genes across these cell lines.
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| 11. |
- Karlsson, Max, et al.
(författare)
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A single-cell type transcriptomics map of human tissues
- 2021
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Ingår i: Science Advances. - : American Association for the Advancement of Science (AAAS). - 2375-2548. ; 7:31
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Tidskriftsartikel (refereegranskat)abstract
- Advances in molecular profiling have opened up the possibility to map the expression of genes in cells, tissues, and organs in the human body. Here, we combined single-cell transcriptomics analysis with spatial antibody-based protein profiling to create a high-resolution single-cell type map of human tissues. An open access atlas has been launched to allow researchers to explore the expression of human protein-coding genes in 192 individual cell type clusters. An expression specificity classification was performed to determine the number of genes elevated in each cell type, allowing comparisons with bulk transcriptomics data. The analysis highlights distinct expression clusters corresponding to cell types sharing similar functions, both within the same organs and between organs.
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| 12. |
- Karlsson, Max, et al.
(författare)
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Genome-wide single cell annotation of the human protein-coding genes
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Annan publikation (övrigt vetenskapligt/konstnärligt)abstract
- An important quest for the life science community is to deliver a complete annotation of the human building-blocks of life, the genes and the proteins. Here, we report on a genome-wide effort to annotate all protein-coding genes based on single cell transcriptomics data representing all major tissues and organs in the human body, integrated with data from bulk transcriptomics and antibody-based tissue profiling. Altogether, 25 tissues have been analyzed with single cell transcriptomics resulting in genome-wide expression in 444 single cell types using a strategy involving pooling data from individual cells to obtain genome-wide expression profiles of individual cell type. We introduce a new genome-wide classification tool based on clustering of similar expression profiles across single cell types, which can be visualized using dimensional reduction maps (UMAP). The clustering classification is integrated with a new “tau” score classification for all protein-coding genes, resulting in a measure of single cell specificity across all cell types for all individual genes. The analysis has allowed us to annotate all human protein-coding genes with regards to function and spatial distribution across individual cell types across all major tissues and organs in the human body. A new version of the open access Human Protein Atlas (www.proteinatlas.org) has been launched to enable researchers to explore the new genome-wide annotation on an individual gene level.
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| 13. |
- Mank, Judith E, et al.
(författare)
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Pleiotropic constraint hampers the resolution of sexual antagonism in vertebrate gene expression
- 2008
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Ingår i: American Naturalist. - : University of Chicago Press. - 0003-0147 .- 1537-5323. ; 171:1, s. 35-43
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Tidskriftsartikel (refereegranskat)abstract
- The numerous physiological and phenotypic differences between the sexes, as well as the disparity between male and female reproductive interests, result in sexual conflicts, which are often manifested at the genomic level. Sexually antagonistic genes benefit one sex at the expense of the other and experience strong pressure to evolve male- and female-specific expression patterns to resolve sexual conflicts and maximize fitness for both sexes. Sex-biased gene expression has recently been demonstrated for much of the metazoan transcriptome, suggesting that many loci are sexually antagonistic. However, many coding regions function in multiple processes throughout the organism. This pleiotropy increases the complexity of selection for any given gene, which in turn may obscure sex-specific selective pressures and hamper the evolution of sex-biased gene expression. Here we use microarray gene expression data, in conjunction with data on transcript abundance from expressed sequence tag libraries, to demonstrate that loci with sex-biased gene expression are significantly less pleiotropic than unbiased genes. This relationship was independent of sex chromosome gene dosage effects, and the results were concordant across two study organisms, chicken and mouse. These results suggest that the resolution of sexually antagonistic gene expression is determined by the evolutionary constraints acting on any given antagonistic locus.
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| 14. |
- Norreen-Thorsen, Marthe, et al.
(författare)
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A human adipose tissue cell-type transcriptome atlas
- 2022
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Ingår i: Cell Reports. - : Elsevier BV. - 2211-1247. ; 40:2
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Tidskriftsartikel (refereegranskat)abstract
- The importance of defining cell-type-specific genes is well acknowledged. Technological advances facilitate high-resolution sequencing of single cells, but practical challenges remain. Adipose tissue is composed pri-marily of adipocytes, large buoyant cells requiring extensive, artefact-generating processing for separation and analysis. Thus, adipocyte data are frequently absent from single-cell RNA sequencing (scRNA-seq) data -sets, despite being the primary functional cell type. Here, we decipher cell-type-enriched transcriptomes from unfractionated human adipose tissue RNA-seq data. We profile all major constituent cell types, using 527 visceral adipose tissue (VAT) or 646 subcutaneous adipose tissue (SAT) samples, identifying over 2,300 cell-type-enriched transcripts. Sex-subset analysis uncovers a panel of male-only cell-type-enriched genes. By resolving expression profiles of genes differentially expressed between SAT and VAT, we identify mesothelial cells as the primary driver of this variation. This study provides an accessible method to profile cell-type-enriched transcriptomes using bulk RNA-seq, generating a roadmap for adipose tissue biology.
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| 15. |
- Stadler, Charlotte, et al.
(författare)
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RNA- and Antibody-Based Profiling of the Human Proteome with Focus on Chromosome 19
- 2014
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Ingår i: Journal of Proteome Research. - : American Chemical Society (ACS). - 1535-3893 .- 1535-3907. ; 13:4, s. 2019-2027
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Tidskriftsartikel (refereegranskat)abstract
- An important part of the Human Proteome Project is to characterize the protein complement of the genome with antibody-based profiling. Within the framework of this effort, a new version 12 of the Human Protein Atlas (www.proteinatlas.org) has been launched, including transcriptomics data for 27 tissues and 44 cell lines to complement the protein expression data from antibody-based profiling. Besides the extensive addition of transcriptomics data, the Human Protein Atlas now contains antibody-based protein profiles for 82% of the 20 329 putative protein-coding genes. The comprehensive data resulting from RNA-seq analysis and antibody-based profiling performed within the Human Protein Atlas as well as information from UniProt were used to generate evidence summary scores for each of the 20 329 genes, of which 94% now have experimental evidence at least at transcript level. The evidence scores for all individual genes are displayed with regards to both RNA- and antibody-based protein profiles, including chromosome-centric visualizations. An analysis of the human chromosome 19 shows that similar to 43% of the genes are expressed at the transcript level in all 27 tissues analyzed, suggesting a "house-keeping" function, while 12% of the genes show a more tissue-specific pattern with enriched expression in one of the analyzed tissues only.
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| 16. |
- Tegel, Hanna, et al.
(författare)
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High throughput generation of a resource of the human secretome in mammalian cells
- 2020
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Ingår i: New Biotechnology. - : Elsevier BV. - 1871-6784 .- 1876-4347. ; 58, s. 45-54
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Tidskriftsartikel (refereegranskat)abstract
- The proteins secreted by human tissues and blood cells, the secretome, are important both for the basic understanding of human biology and for identification of potential targets for future diagnosis and therapy. Here, a high-throughput mammalian cell factory is presented that was established to create a resource of recombinant full-length proteins covering the majority of those annotated as 'secreted' in humans. The full-length DNA sequences of each of the predicted secreted proteins were generated by gene synthesis, the constructs were transfected into Chinese hamster ovary (CHO) cells and the recombinant proteins were produced, purified and analyzed. Almost 1,300 proteins were successfully generated and proteins predicted to be secreted into the blood were produced with a success rate of 65%, while the success rates for the other categories of secreted proteins were somewhat lower giving an overall one-pass success rate of ca. 58%. The proteins were used to generate targeted proteomics assays and several of the proteins were shown to be active in a phenotypic assay involving pancreatic beta-cell dedifferentiation. Many of the proteins that failed during production in CHO cells could be rescued in human embryonic kidney (HEK 293) cells suggesting that a cell factory of human origin can be an attractive alternative for production in mammalian cells. In conclusion, a high-throughput protein production and purification system has been successfully established to create a unique resource of the human secretome.
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| 17. |
- Uhlén, Mathias, et al.
(författare)
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A genome-wide transcriptomic analysis of protein-coding genes in human blood cells
- 2019
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Ingår i: Science. - : American Association for the Advancement of Science. - 0036-8075 .- 1095-9203. ; 366:6472, s. 1471-
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Tidskriftsartikel (refereegranskat)abstract
- Blood is the predominant source for molecular analyses in humans, both in clinical and research settings. It is the target for many therapeutic strategies, emphasizing the need for comprehensive molecular maps of the cells constituting human blood. In this study, we performed a genome-wide transcriptomic analysis of protein-coding genes in sorted blood immune cell populations to characterize the expression levels of each individual gene across the blood cell types. All data are presented in an interactive, open-access Blood Atlas as part of the Human Protein Atlas and are integrated with expression profiles across all major tissues to provide spatial classification of all protein-coding genes. This allows for a genome-wide exploration of the expression profiles across human immune cell populations and all major human tissues and organs.
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| 18. |
- Uhlén, Mathias, et al.
(författare)
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Tissue-based map of the human proteome
- 2015
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Ingår i: Science. - : American Association for the Advancement of Science (AAAS). - 0036-8075 .- 1095-9203. ; 347:6220, s. 1260419-
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Tidskriftsartikel (refereegranskat)abstract
- Resolving the molecular details of proteome variation in the different tissues and organs of the human body will greatly increase our knowledge of human biology and disease. Here, we present a map of the human tissue proteome based on an integrated omics approach that involves quantitative transcriptomics at the tissue and organ level, combined with tissue microarray-based immunohistochemistry, to achieve spatial localization of proteins down to the single-cell level. Our tissue-based analysis detected more than 90% of the putative protein-coding genes. We used this approach to explore the human secretome, the membrane proteome, the druggable proteome, the cancer proteome, and the metabolic functions in 32 different tissues and organs. All the data are integrated in an interactive Web-based database that allows exploration of individual proteins, as well as navigation of global expression patterns, in all major tissues and organs in the human body.
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| 19. |
- Uhlén, Mathias, et al.
(författare)
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Towards a knowledge-based Human Protein Atlas
- 2010
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Ingår i: Nature Biotechnology. - : Springer Science and Business Media LLC. - 1087-0156 .- 1546-1696. ; 28:12, s. 1248-1250
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Tidskriftsartikel (refereegranskat)
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| 20. |
- Öling, S., et al.
(författare)
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A human stomach cell type transcriptome atlas
- 2024
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Ingår i: BMC Biology. - : Springer Nature. - 1741-7007. ; 22:1
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Tidskriftsartikel (refereegranskat)abstract
- Background: The identification of cell type-specific genes and their modification under different conditions is central to our understanding of human health and disease. The stomach, a hollow organ in the upper gastrointestinal tract, provides an acidic environment that contributes to microbial defence and facilitates the activity of secreted digestive enzymes to process food and nutrients into chyme. In contrast to other sections of the gastrointestinal tract, detailed descriptions of cell type gene enrichment profiles in the stomach are absent from the major single-cell sequencing-based atlases. Results: Here, we use an integrative correlation analysis method to predict human stomach cell type transcriptome signatures using unfractionated stomach RNAseq data from 359 individuals. We profile parietal, chief, gastric mucous, gastric enteroendocrine, mitotic, endothelial, fibroblast, macrophage, neutrophil, T-cell, and plasma cells, identifying over 1600 cell type-enriched genes. Conclusions: We uncover the cell type expression profile of several non-coding genes strongly associated with the progression of gastric cancer and, using a sex-based subset analysis, uncover a panel of male-only chief cell-enriched genes. This study provides a roadmap to further understand human stomach biology.
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