Human Brain Proteome Project Research Articles

Deep Phosphoproteome Landscape of Interhemispheric Functionality of Neuroanatomical Regions of the Human Brain.

Post-translational modifications (PTMs) are one of the compulsive and predominant biological processes that regulate the diverse molecular mechanism, modulate the onset of disease, and are the reason behind the functional diversity of proteins. Despite the widespread research findings in neuroproteomics, one of the key drawbacks has been the lack of proteome-level knowledge of hemispheric lateralization. We have investigated the proteome level expression in different neuroanatomical regions under the Human Brain Proteome Project (HBPP) and developed the global interhemispheric brain proteome map (Brainprot) earlier. Furthermore, this study has extended to decipher the phosphoproteome map of human brain interhemispheric regions through high-resolution mass spectrometry. The phosphoproteomics examination of 12 unique interhemispheric neurological brain regions using Orbitrap fusion liquid chromatography with tandem mass spectrometry provided comprehensive coverage of 996 phosphoproteins, 2010 phosphopeptides, and 3567 phosphosites. Moreover, interhemispheric phosphoproteome profiling has been categorized according to synaptic ontologies and interhemispheric expression to understand the functionality. Finally, we have integrated the phosphosites data under the PhosphoMap section in the Inter-Hemispheric Brain Proteome Map Portal (https://www.brainprot.org/) for the advancement and support of the ongoing neuroproteomics research worldwide. Data is available via ProteomeXchange with the identifier PXD031188.

Smelling the Dark Proteome: Functional Characterization of PITH Domain-Containing Protein 1 (C1orf128) in Olfactory Metabolism.

The Human Proteome Project (HPP) consortium aims to functionally characterize the dark proteome. On the basis of the relevance of olfaction in early neurodegeneration, we have analyzed the dark proteome using data mining in public resources and omics data sets derived from the human olfactory system. Multiple dark proteins localize at synaptic terminals and may be involved in amyloidopathies such as Alzheimer's disease (AD). We have characterized the dark PITH domain-containing protein 1 (PITHD1) in olfactory metabolism using bioinformatics, proteomics, in vitro and in vivo studies, and neuropathology. PITHD1-/- mice exhibit olfactory bulb (OB) proteome changes related to synaptic transmission, cognition, and memory. OB PITHD1 expression increases with age in wild-type (WT) mice and decreases in Tg2576 AD mice at late stages. The analysis across 6 neurological disorders reveals that olfactory tract (OT) PITHD1 is specifically upregulated in human AD. Stimulation of olfactory neuroepithelial (ON) cells with PITHD1 alters the ON phosphoproteome, modifies the proliferation rate, and induces a pro-inflammatory phenotype. This workflow applied by the Spanish C-HPP and Human Brain Proteome Project (HBPP) teams across the ON-OB-OT axis can be adapted as a guidance to decipher functional features of dark proteins. Data are available via ProteomeXchange with identifiers PXD018784 and PXD021634.

The Human Brain Proteome Project: Biological and Technological Challenges.

Brain proteomics has become a method of choice that allows zooming-in where neuropathophysiological alterations are taking place, detecting protein mediators that might eventually be measured in cerebrospinal fluid (CSF) as potential neuropathologically derived biomarkers. Following this hypothesis, mass spectrometry-based neuroproteomics has emerged as a powerful approach to profile neural proteomes derived from brain structures and CSF in order to map the extensive protein catalog of the human brain. This chapter provides a historical perspective on the Human Brain Proteome Project (HBPP), some recommendation to the experimental design in neuroproteomic projects, and a brief description of relevant technological and computational innovations that are emerging in the neurobiology field thanks to the proteomics community. Importantly, this chapter highlights recent discoveries from the biology- and disease-oriented branch of the HBPP (B/D-HBPP) focused on spatiotemporal proteomic characterizations of mouse models of neurodegenerative diseases, elucidation of proteostatic networks in different types of dementia, the characterization of unresolved clinical phenotypes, and the discovery of novel biomarker candidates in CSF.

Synaptosomal Proteome of the Orbitofrontal Cortex from Schizophrenia Patients Using Quantitative Label-Free and iTRAQ-Based Shotgun Proteomics.

Schizophrenia is a chronic and incurable neuropsychiatric disorder that affects about one percent of the world population. The proteomic characterization of the synaptosome fraction of the orbitofrontal cortex is useful for providing valuable information about the molecular mechanisms of synaptic functions in these patients. Quantitative analyses of synaptic proteins were made with eight paranoid schizophrenia patients and a pool of eight healthy controls free of mental diseases. Label-free and iTRAQ labeling identified a total of 2018 protein groups. Statistical analyses revealed 12 and 55 significantly dysregulated proteins by iTRAQ and label-free, respectively. Quantitative proteome analyses showed an imbalance in the calcium signaling pathway and proteins such as reticulon-1 and cytochrome c, related to endoplasmic reticulum stress and programmed cell death. Also, it was found that there is a significant increase in limbic-system-associated membrane protein and α-calcium/calmodulin-dependent protein kinase II, associated with the regulation of human behavior. Our data contribute to a better understanding about apoptosis as a possible pathophysiological mechanism of this disease as well as neural systems supporting social behavior in schizophrenia. This study also is a joint effort of the Chr 15 C-HPP team and the Human Brain Proteome Project of B/D-HPP. All MS proteomics data are deposited in the ProteomeXchange Repository under PXD006798.

Proteomic atlas of the human olfactory bulb

The olfactory bulb (OB) is the first site for the processing of olfactory information in the brain and its deregulation is associated with neurodegenerative disorders. Although different efforts have been made to characterize the human brain proteome in depth, the protein composition of the human OB remains largely unexplored. We have performed a comprehensive analysis of the human OB proteome employing protein and peptide fractionation methods followed by LC-MS/MS, identifying 1529 protein species, corresponding to 1466 unique proteins, which represents a 7-fold increase in proteome coverage with respect to previous OB proteome descriptions from translational models. Bioinformatic analyses revealed that protein components of the OB participated in a plethora of biological process highlighting hydrolase and phosphatase activities and nucleotide and RNA binding activities. Interestingly, 631 OB proteins identified were not previously described in protein datasets derived from large-scale Human Brain Proteome Project (HBPP) studies. In particular, a subset of these differential proteins was mainly involved in axon guidance, opioid signaling, neurotransmitter receptor binding, and synaptic plasticity. Taken together, these results increase our knowledge about the molecular composition of the human OB and may be useful to understand the molecular basis of the olfactory system and the etiology of its disorders.

Human Brain Proteome Project – 12th HUPO BPP Workshop 26 September 2009, Toronto, Canada

The HUPO Brain Proteome Project (HUPO BPP) held its 12th workshop in Toronto on 26 September 2009 prior to the HUPO VIII World Congress. The principal aim of this project is to obtain a better understanding of neurodiseases and ageing, with the ultimate objective of discovering prognostic and diagnostic biomarkers, in addition to the development of novel diagnostic techniques and new medications. The attendees came together to discuss progress in the human clinical neuroproteomics and to define the needs and guidelines required for more advanced proteomic approaches.

Human Microglial Cells Synthesize Albumin in Brain

Albumin, an abundant plasma protein with multifunctional properties, is mainly synthesized in the liver. Albumin has been implicated in Alzheimer's disease (AD) since it can bind to and transport amyloid beta (Aβ), the causative agent of AD; albumin is also a potent inhibitor of Aβ polymerization. Despite evidence of non-hepatic transcription of albumin in many tissues including kidney and pancreas, non-hepatic synthesis of albumin at the protein level has been rarely confirmed. In a pilot phase study of Human Brain Proteome Project, we found evidence that microglial cells in brain may synthesize albumin. Here we report, for the first time, the de novo synthesis of albumin in human microglial cells in brain. Furtherore, we demonstrate that the synthesis and secretion of albumin from microglial cells is enhanced upon microgial activation by Aβ1–42- or lipopolysaccharide (LPS)-treatment. These data indicate that microglial cells may play a beneficial role in AD by secreting albumin that not only inhibits Aβ polymerization but also increases its clearance.

The HUPO Brain Proteome Project Wish List – Summary of the 9th HUPO BPP Workshop 9–10 January 2008, Barbados

The Human Brain Proteome Project (HUPO BPP) aims at advancing knowledge and the understanding of neurodiseases and aging with the purpose of identifying prognostic and diagnostic biomarkers, as well as to push new diagnostic approaches and medications. The participating groups meet in semi-annual workshops to discuss the progress, as well as the needs, within the field of proteomics. The 9(th) HUPO BPP workshop took place in Barbados from 9-10 January, 2008. Discussing the future HUPO BPP Roadmap, the attendees drafted the so called HUPO BPP wish list containing timelines, suggestions and missions. This wish list will be updated regularly and will serve as a guideline for the next phase.

Multidimensional advancement of neuroproteomics

Tim Veenstra is the Director of the Laboratory of Proteomics and Analytical Technologies at the National Cancer Institute at Frederick, MD, USA. Veenstra acquired his PhD in biochemistry from the University of Windsor, Canada, in 1994 under the guidance of Lana Lee. He then moved to the laboratory of Rajiv Kumar at the Mayo Clinic in Rochester, MN, USA, where he completed a postdoctoral fellowship in molecular biology. He has been at his current position for 7 years. The focus of his research deals primarily with the discovery of novel biomarkers for diseases such as cancer. To accomplish this goal, his laboratory has developed methods to analyze the proteomes and metabolomes of thin sections obtained from both fresh-frozen and formalin-fixed paraffin-embedded tissues. His laboratory is also interested in developing and applying methods to more effectively characterize the proteomes and metabolomes of various biofluids for the discovery of both diagnostic and therapeutic biomarkers.Katrin Marcus studied Biochemistry at the Ruhr-University Bochum, Germany. After finishing her diploma thesis, she elaborated her PhD at the Proteinstrukturlabor of Professor HE Meyer in Bochum, Germany, analyzing the phosphoproteome of human thrombin-stimulated platelets. Since August 2002, she has been group leader at the Medizinisches Proteom-Center, supervising several projects such as the ‘Human Brain Proteome Project’ and ‘Clinical Neuroproteomics of Neurodegenerative Diseases’. In August 2003, she was appointed as an Assistant Professor for proteomics at the Medical Faculty of the Ruhr-University. In December 2007, she became full Professor and now leads the Department of Functional Proteomics. Her scientific work is focused on the discovery of biomarkers for Alzheimer’s and Parkinson’s disease, and the analysis of integral membrane proteins of human hepatocytes that play a pivotal role in all phases of xenobiotics metabolism.

Human microglial cells synthesize albumin in brain

AbstractAlbumin has been implicated in Alzheimer's disease since it can bind to and transport amyloid beta, the causative agent; albumin is also a potent inhibitor of amyloid beta polymerization. In a pilot phase study of Human Brain Proteome Project, we found evidence that albumin may be synthesized in immortalized human microglial cells, human primary microglial cells, and human fetal and adult brain tissues. We also found the synthesis and secretion is enhanced upon microglial activation by Amyloid [beta]~1-42~, lipopolysaccharide treatment or human Alzheimer's brain.

BrainProfileDB – a platform for integration of functional genomics data

BrainProfileDB is a database system for integrating large sets of high throughput functional genomics data of the Human Brain Proteome Project (HBPP). Within HBPP (http://www.smp-proteomics.de/) the molecular pathology of neurodegenerative diseases is investigated, using complementary methods from transcriptomics, proteomics, toponomics and interaction measurements. Aim of the database system is to provide a broad spectrum of scientific users joined in the consortium with a practical integrated view on their data. Employing appropriate mapping techniques and levels of data representation the user is relieved from technical details of gene identification or experimental measurement technique.

Seven successful years of Omics research: The Human Brain Proteome Project within the National German Research Network (NGFN)

Seven successful years of Omics research: The Human Brain Proteome Project within the National German Research Network (NGFN)

Systematic gene expression profiling of mouse model series reveals coexpressed genes

A major aim of the Human Brain Proteome Project (HBPP) is a better understanding of the molecular etiology and progression of neurodegenerative diseases. Transgenic and loss-of-function mouse mutant lines (MMLs) serve as experimental models. Transcriptome and proteome regulate each other in a complex and controlled way, and their comparative analysis is an essential aspect. As a fundamental study, we have assessed transcript profiles using a microarray containing 21 000 cDNA probes in a series of disease models within the German Mouse Clinic (GMC). Seventeen distinct organs of one adult stage were systematically collected for each submitted MML. Samples for gene expression profiling are individually selected based on conspicuous phenotypes in at least one of 14 GMC phenotype screens or on previous knowledge of the mutant phenotype. By microarray experiments expression patterns of 90 organs from 46 MMLs were analysed, identifying up to 232 differentially expressed genes in 45 organs. Here we present an overview of the results of all MMLs analysed and demonstrate the efficiency of systematic genome-wide expression profiling for the detection of molecular phenotypes in organs of a mammalian model organism. We identify the recurring regulation of particular genes and groups of coexpressed genes in apparently unrelated MMLs.

Inside SMP proteomics: Six years German Human Brain Proteome Project (HBPP) – A summary

In 2001, the German Federal Ministry of Education and Research (BMBF) initiated the National Genome Research Network (NGFN; www.ngfn.de) as a nation-wide multidisciplinary networking platform aiming at the analysis of common human diseases and aging. Within the NGFN the Human Brain Proteome Project (HBPP; www.smp-proteomics.de) focuses on the analysis of the human brain in health and disease. The concept is based on two consecutive steps: (i) Elaborating and establishing the necessary technology platforms. (ii) Application of the established technologies for research in Alzheimer's disease and Parkinson's disease. In the first funding period, HBPP1, running from 2001 to 2004, necessary technologies were established and optimized. In HBPP2, which started 2004 and will end in May 2008, the developed technologies are used for large-scale experiments, offering new links for disease related research and therapies. The following overview describes structure, aims and outcome of this unique German Brain Proteome Project.

HUPO Brain Proteome Project: Toward a Code of Conduct

The prevalence of neurodegenerative diseases in all societies increases steadily and so do the costs for comprehensive scientific proteomics studies. This is due to immense expenses for equipment and consumables as well as to a high number of repetitions to fulfill the statistical imperative. At the same time, techniques become even more complex, whereas suitable samples are scarce, especially when dealing with the valuable human brain. What is the consequence? What should be done to overcome this contradiction within the triangle validated protein analyses, time, and money? The Human Brain Proteome Project (BPP) is one of the initial initiatives of the Human Proteome Organisation (HUPO). Operating since 2003, it gathers a critical mass of researchers from academia and industry aiming at advancing knowledge of neurodiseases and aging to elaborate new diagnostic approaches and medications. HUPO BPP wants to perform suitable quantitative proteomics on disease-related brain areas and bodily fluids to analyze the brain proteome of human as well as mouse models with healthy, neurodiseased, and aged statuses with a focus on Alzheimer and Parkinson disease. Recent work has shown that standards in proteomics and especially in bioinformatics are mandatory to allow comparable analyses. To address this challenge, the HUPO BPP is closely working together with the HUPO Proteome Standards Initiative (HUPO PSI). Two standardization pilot studies are already finished and documented in a special edition (Proteomics, Volume 6, Issue 18). Due to the combined effort of 18 laboratories worldwide a huge amount of data sets with current proteomics technologies were gained and analyzed. Only a very few of the identified differentially expressed proteins could be found by many or all of the participating groups, although each of the individual laboratories could show to have detected several dozens of reproducible differences. The yield of an individual protein is very dependent on the conditions for sample preparation, i.e. salt concentration, pH, detergents, temperature, etc., and every little change will result in a different sample composition and thus another result. Therefore it is important that experiments are repeated independently several times to show their reproducibility as it was already suggested by Wilkins et al. (1). During the studies, participants agreed that the standards and standard operating procedures are essential for successful work. Annotations concerning sample handling, preparation, separation, and identification have to be considered and documented. Different approaches and search engines are complementary. The separation features of the different techniques do overlap and can be applied successively. Nevertheless every study has to show the reproducibility of its results. As the overlap of the identified proteins in regard to the different laboratories is not optimal, it is extremely important that their own data are handled very critically. The validation of the proteins identified is the next essential step. These and other considerations are also discussed in some of the latest publications of the HUPO BPP consortium (2–4). To fix these maxims, the next workshops will be essential. The 9th HUPO BPP workshop will take place in Barbados from January 9 to 10, 2008. In this workshop, involved colleagues will discuss and lay down the HUPO BPP roadmap for the next years, stressing standard operating procedures, suitable models, and study designs. Funding and further integration into HUPO’s world also will be an important issue. The 10th HUPO BPP congress is scheduled for the HUPO World Congress in Amsterdam, August 2008. Interested colleagues are always welcome and may visit the HUPO Brain Proteome Project website for the latest news.

International Symposium on High Performance Proteomics 2007: A contribution to Proteomics May 14–16, 2007 Dortmund, Germany

The symposium High Performance Proteomics was held in Dortmund on May 14-16, 2007, to celebrate the opening of the Zentrum für Angewandte Proteomik as well as the 6(th) anniversary of the German Human Brain Proteome Project. It offered an outstanding opportunity to obtain a broad overview about all fields of proteomics and related fields, combining the expertise of biochemists, physicians, bioinformatics, mathematicians and other researchers in Life Sciences. The main topics were the presentation of state-of-the-art proteomics technologies as well as possible transfer models for industrial applications. An accompanying industrial exhibition, as well as a discussion panel, offered the possibility to get in contact with colleagues and potential industrial partners. A visit to the former colliery Zeche Zollern and the social event at the Harenberg City-Center with an excellent view around Dortmund also left time for further communication between the more than 200 attendees.

Neuroproteomics comes of age

Neuroproteomics comes of age

Evaluation of 2‐DE protein patterns from pre‐ and postnatal stages of the mouse brain

Brains of the mouse from three developmental stages, embryo day 16 (Ed16), postnatal stage one week (1W) and eight weeks (8W), were distributed to different laboratories for a collaborative proteome analysis (The Human Brain Proteome Project). As one of the laboratories involved in this project, we separated total protein extracts of the brains by large gel 2-DE. From the 2-DE protein patterns a section was evaluated for each of the three stages according to resolution, reproducibility and quantitative changes using an image analysis software. The evaluated pattern section was selected to allow comparisons of 2-DE patterns between different laboratories on the basis of optimum separation. Changes in protein expression were analysed within two phases of development: Stage Ed16 versus stage 1W and stage 1W versus stage 8W. Out of the 200 protein spots evaluated 5-6% showed quantitative changes in the range of > or = 30% between two stages. The relationship in the frequency of up- and down-regulated protein spots differed between the two investigated phases. Most of the protein spots which showed altered expression between two stages were identified by MS. High quality in protein separation and evaluation is demonstrated.

Great mood in proteomics: Beijing and the HUPO Human Brain Proteome Project

More than 1200 attendees came together at the 3(rd) HUPO World Congess in Beijing, October 25-27, 2004. In numerous different sessions the wide range of proteomic areas became visible. The HUPO Brain Proteome Project (HUPO BPP) organized an evening session on October 23, presenting the first results of two pilot studies as well as the newest, very positive international development in this field. The rising importance became even more apparent in the plenary presentation of all HUPO initiatives and the following congress activities.

Towards data management of the HUPO Human Brain Proteome Project pilot phase.

The pilot phase of the Human Brain Proteome Project as a part of the Human Proteome Organisation has just been started. In two pilot studies, 18 different laboratories are analyzing mouse brains of three age stages and human brain autopsy versus biopsy material, respectively. The overall aim is to elucidate the portfolio of available techniques as well as to elaborate common standards. As a first step, it was decided to use the common bioinformatics platform ProteinScape that was introduced to the participating groups in a two day course in Bochum, Germany.