The human protein atlas: Implications for human biology, drug development, and precision medicine

The Human Protein Atlas (HPA) is a Swedish-based program with the aim to map of all the human proteins in cells, tissues and organs using integration of various omics technologies, including genomics, transcriptomics, antibody-based imaging, mass spectrometry-based proteomics and systems biology. A Tissue Atlas was launch in 2015 (1) followed by a Cell Atlas in 2016 (2) and a Pathology Atlas in 2017 (3). This open access knowledge-base can be used to explore targets for next generation antibody therapeutics, as well as a discovery tool to find potential biomarkers and drug targets for disease (4,5). A focus has been to use a new drug development platform based on the affibody molecule developed in our group and use this concept for applications in cancer, autoimmune diseases and neurodegenerative diseases. Recently, we have set-up an animal cell factory using CHO cells with the aim to produce full-length proteins representing all the 2,000 secreted proteins encoded in human genome. The Human Protein Atlas program has already contributed to several thousands of publications in the field of human biology and disease and it was recently selected by the organization ELIXIR as a European core resource, due to its fundamental importance for a wider life science community. All the data in the knowledge resource is open access to allow scientists both in academia and industry to freely access the data for exploration of the human proteome. Selected recent references: 1. Uhlen et al (2015) Science 347: 394 2. Thul et al (2017), Science 356:6340 3. Uhlen et al (2017) Science (August 18) 4. Uhlen et al (2016) Mol Systems Biol. 12: 862 5. Lee et al (2016) Cell Metabolism 12;24(1):172-8

Human protein secretory pathway genes are expressed in a tissue-specific pattern to match processing demands of the secretome

Protein secretory pathway in eukaryal cells is responsible for delivering functional secretory proteins. The dysfunction of this pathway causes a range of important human diseases from congenital disorders to cancer. Despite the piled-up knowledge on the molecular biology and biochemistry level, the tissue-specific expression of the secretory pathway genes has not been analyzed on the transcriptome level. Based on the recent RNA-sequencing studies, the largest fraction of tissue-specific transcriptome encodes for the secretome (secretory proteins). Here, the question arises that if the expression levels of the secretory pathway genes have a tissue-specific tuning. In this study, we tackled this question by performing a meta-analysis of the recently published transcriptome data on human tissues. As a result, we detected 68 as called “extreme genes” which show an unusual expression pattern in specific gene families of the secretory pathway. We also inspected the potential functional link between detected extreme genes and the corresponding tissues enriched secretome. As a result, the detected extreme genes showed correlation with the enrichment of the nature and number of specific post-translational modifications in each tissue’s secretome. Our findings conciliate both the housekeeping and tissue-specific nature of the protein secretory pathway, which we attribute to a fine-tuned regulation of defined gene families to support the diversity of secreted proteins and their modifications

Scheffersomyces stipitis: a comparative systems biology study with the Crabtree positive yeast Saccharomyces cerevisiae

<p>Abstract</p> <p>Background</p> <p><it>Scheffersomyces stipitis</it> is a Crabtree negative yeast, commonly known for its capacity to ferment pentose sugars. Differently from Crabtree positive yeasts such as <it>Saccharomyces cerevisiae</it>, the onset of fermentation in <it>S. stipitis</it> is not dependent on the sugar concentration, but is regulated by a decrease in oxygen levels. Even though <it>S. stipitis</it> has been extensively studied due to its potential application in pentoses fermentation, a limited amount of information is available about its metabolism during aerobic growth on glucose. Here, we provide a systems biology based comparison between the two yeasts, uncovering the metabolism of <it>S. stipitis</it> during aerobic growth on glucose under batch and chemostat cultivations.</p> <p>Results</p> <p>Starting from the analysis of physiological data, we confirmed through ¹³C-based flux analysis the fully respiratory metabolism of <it>S. stipitis</it> when growing both under glucose limited or glucose excess conditions. The patterns observed showed similarity to the fully respiratory metabolism observed for <it>S. cerevisiae</it> under chemostat cultivations however, intracellular metabolome analysis uncovered the presence of several differences in metabolite patterns. To describe gene expression levels under the two conditions, we performed RNA sequencing and the results were used to quantify transcript abundances of genes from the central carbon metabolism and compared with those obtained with <it>S. cerevisiae</it>. Interestingly, genes involved in central pathways showed different patterns of expression, suggesting different regulatory networks between the two yeasts. Efforts were focused on identifying shared and unique families of transcription factors between the two yeasts through <it>in silico</it> transcription factors analysis, suggesting a different regulation of glycolytic and glucoenogenic pathways.</p> <p>Conclusions</p> <p>The work presented addresses the impact of high-throughput methods in describing and comparing the physiology of Crabtree positive and Crabtree negative yeasts. Based on physiological data and flux analysis we identified the presence of one metabolic condition for <it>S. stipitis</it> under aerobic batch and chemostat cultivations, which shows similarities to the oxidative metabolism observed for <it>S. cerevisiae</it> under chemostat cultivations. Through metabolome analysis and genome-wide transcriptomic analysis several differences were identified. Interestingly, <it>in silico</it> analysis of transciption factors was useful to address a different regulation of mRNAs of genes involved in the central carbon metabolism. To our knowledge, this is the first time that the metabolism of <it>S. stiptis</it> is investigated in details and is compared to <it>S. cerevisiae</it>. Our study provides useful results and allows for the possibility to incorporate these data into recently developed genome-scaled metabolic, thus contributing to improve future industrial applications of <it>S. stipitis</it> as cell factory.</p

Discovery of dachshund 2 protein as a novel biomarker of poor prognosis in epithelial ovarian cancer

<p>Abstract</p> <p>Background</p> <p>The Dachshund homolog 2 (<it>DACH2</it>) gene has been implicated in development of the female genital tract in mouse models and premature ovarian failure syndrome, but to date, its expression in human normal and cancerous tissue remains unexplored. Using the Human Protein Atlas as a tool for cancer biomarker discovery, DACH2 protein was found to be differentially expressed in epithelial ovarian cancer (EOC). Here, the expression and prognostic significance of DACH2 was further evaluated in ovarian cancer cell lines and human EOC samples.</p> <p>Methods</p> <p>Immunohistochemical expression of DACH2 was examined in tissue microarrays with 143 incident EOC cases from two prospective, population-based cohorts, including a subset of benign-appearing fallopian tubes (n = 32). A nuclear score (NS), i.e. multiplier of staining fraction and intensity, was calculated. For survival analyses, cases were dichotomized into low (NS < = 3) and high (NS > 3) using classification and regression tree analysis. Kaplan Meier analysis and Cox proportional hazards modelling were used to assess the impact of DACH2 expression on survival. DACH2 expression was analysed in the cisplatin sensitive ovarian cancer cell line A2780 and its cisplatin resistant derivative A2780-Cp70. The specificity of the DACH2 antibody was tested using siRNA-mediated silencing of DACH2 in A2780-Cp70 cells.</p> <p>Results</p> <p>DACH2 expression was considerably higher in the cisplatin resistant A2780-Cp70 cells compared to the cisplatin-sensitive A2780 cells. While present in all sampled fallopian tubes, DACH2 expression ranged from negative to strong in EOC. In EOC, DACH2 expression correlated with several proteins involved in DNA integrity and repair, and proliferation. DACH2 expression was significantly higher in carcinoma of the serous subtype compared to non-serous carcinoma. In the full cohort, high DACH2 expression was significantly associated with poor prognosis in univariable analysis, and in carcinoma of the serous subtype, DACH2 remained an independent factor of poor prognosis.</p> <p>Conclusions</p> <p>This study provides a first demonstration of DACH2 protein being expressed in human fallopian tubes and EOC, with the highest expression in serous carcinoma where DACH2 was found to be an independent biomarker of poor prognosis. Future research should expand on the role of DACH2 in ovarian carcinogenesis and chemotherapy resistance.</p

The mucosa-associated bacteria from the sigmoid colon of nine healthy 60 years old individuals, identified by bacterial 16S rDNA

The bacterial flora of the gastro intestinal (GI) tract may be involved in chronic inflammation and colon cancer and affected by antibiotics, cytotoxic drugs and radiotherapy, trauma and intensive care therapy. It is important to map the mucosa-associated flora in healthy individuals to clarify the pathogenic risk under stressed conditions. The aim was to achieve an overview of the mucosa-associated bacterial flora in the sigmoid colon by direct 16S rDNA identification by sampling nine 60-years old volunteers, without clinical symptoms or medication. The bacterial flora was estimated by sequence analysis of cloned 16S rDNA as enriched by PCR from biopsies. 26% of the clones had ≥99% similarity to known species (36% had ≥98% similarity). The largest number of identified clones was related to Escherichia coli, Bacteroides vulgatus and Ruminicoccus torques. Most frequently distributed between the volunteers were Bacteroides uniformis and Bacteroides vulgatus (7 individuals). Bacteroides caccae, Bacteroides distasonis, Bacteroides putredinis, Bacteroides thetaiotaomicron and Ruminicoccus torques were found in 5 persons. Opportunistic pathogens found in more than one individual were Bacteroides fragilis, Escherichia coli and Bilophila wadsworthia. Acinetobacter baumannii, Brachyspira aalborgi, Cardiobacterium hominis, Clostridium perfringens, Klebsiella pneumoniae and Veillonella parvula were found in single individuals. A majority of the individuals had a heterogeneous flora but in one person, 91% of the clones were related to E. coli. The GI-flora differs between healthy individuals in respect to both composition and diversity, and it can include several opportunistic pathogens

Transcriptomics resources of human tissues and organs

Quantifying the differential expression of genes in various human organs, tissues, and cell types is vital to understand human physiology and disease. Recently, several large-scale transcriptomics studies have analyzed the expression of protein-coding genes across tissues. These datasets provide a framework for defining the molecular constituents of the human body as well as for generating comprehensive lists of proteins expressed across tissues or in a tissue-restricted manner. Here, we review publicly available human transcriptome resources and discuss body-wide data from independent genome-wide transcriptome analyses of different tissues. Gene expression measurements from these independent datasets, generated using samples from fresh frozen surgical specimens and postmortem tissues, are consistent. Overall, the different genome-wide analyses support a distribution in which many proteins are found in all tissues and relatively few in a tissue-restricted manner. Moreover, we discuss the applications of publicly available omics data for building genome-scale metabolic models, used for analyzing cell and tissue functions both in physiological and in disease contexts

Correlations between RNA and protein expression profiles in 23 human cell lines

<p>Abstract</p> <p>Background</p> <p>The Central Dogma of biology holds, in famously simplified terms, that DNA makes RNA makes proteins, but there is considerable uncertainty regarding the general, genome-wide correlation between levels of RNA and corresponding proteins. Therefore, to assess degrees of this correlation we compared the RNA profiles (determined using both cDNA- and oligo-based microarrays) and protein profiles (determined immunohistochemically in tissue microarrays) of 1066 gene products in 23 human cell lines.</p> <p>Results</p> <p>A high mean correlation coefficient (0.52) was obtained from the pairwise comparison of RNA levels determined by the two platforms. Significant correlations, with correlation coefficients exceeding 0.445, between protein and RNA levels were also obtained for a third of the specific gene products. However, the correlation coefficients between levels of RNA and protein products of specific genes varied widely, and the mean correlations between the protein and corresponding RNA levels determined using the cDNA- and oligo-based microarrays were 0.25 and 0.20, respectively.</p> <p>Conclusion</p> <p>Significant correlations were found in one third of the examined RNA species and corresponding proteins. These results suggest that RNA profiling might provide indirect support to antibodies' specificity, since whenever a evident correlation between the RNA and protein profiles exists, this can sustain that the antibodies used in the immunoassay recognized their cognate antigens.</p

Low RBM3 protein expression correlates with tumour progression and poor prognosis in malignant melanoma: An analysis of 215 cases from the Malmö Diet and Cancer Study

<p>Abstract</p> <p>Background</p> <p>We have previously reported that expression of the RNA- and DNA-binding protein RBM3 is associated with a good prognosis in breast cancer and ovarian cancer. In this study, the prognostic value of immunohistochemical RBM3 expression was assessed in incident cases of malignant melanoma from a prospective population-based cohort study.</p> <p>Methods</p> <p>Until Dec 31^st2008, 264 incident cases of primary invasive melanoma had been registered in the Malmö Diet and Cancer Study. Histopathological and clinical information was obtained for available cases and tissue microarrays (TMAs) constructed from 226 (85.6%) suitable paraffin-embedded tumours and 31 metastases. RBM3 expression was analysed by immunohistochemistry on the TMAs and a subset of full-face sections. Chi-square and Mann-Whitney U tests were used for comparison of RBM3 expression and relevant clinicopathological characteristics. Kaplan Meier analysis and Cox proportional hazards modelling were used to assess the relationship between RBM3 and recurrence free survival (RFS) and overall survival (OS).</p> <p>Results</p> <p>RBM3 could be assessed in 215/226 (95.1%) of primary tumours and all metastases. Longitudinal analysis revealed that 16/31 (51.6%) of metastases lacked RBM3 expression, in contrast to the primary tumours in which RBM3 was absent in 3/215 (1.4%) cases and strongly expressed in 120/215 (55.8%) cases. Strong nuclear RBM3 expression in the primary tumour was significantly associated with favourable clinicopathological parameters; i.e. non-ulcerated tumours, lower depth of invasion, lower Clark level, less advanced clinical stage, low mitotic activity and non-nodular histological type, and a prolonged RFS (RR = 0.50; 95% CI = 0.27-0.91) and OS (RR = 0.36, 95%CI = 0.20-0.64). Multivariate analysis demonstrated that the beneficial prognostic value of RBM3 remained significant for OS (RR = 0.33; 95%CI = 0.18-0.61).</p> <p>Conclusions</p> <p>In line with previous in vitro data, we here show that RBM3 is down-regulated in metastatic melanoma and high nuclear RBM3 expression in the primary tumour is an independent marker of a prolonged OS. The potential utility of RBM3 in treatment stratification of patients with melanoma should be pursued in future studies.</p