Proposal for a standard representation of two-dimensional gel electrophoresis data

The global analysis of proteins is now feasible due to improvements in techniques such as two-dimensional gel electrophoresis (2-DE), mass spectrometry, yeast two-hybrid systems and the development of bioinformatics applications. The experiments form the basis of proteomics, and present significant challenges in data analysis, storage and querying. We argue that a standard format for proteome data is required to enable the storage, exchange and subsequent re-analysis of large datasets. We describe the criteria that must be met for the development of a standard for proteomics. We have developed a model to represent data from 2-DE experiments, including difference gel electrophoresis along with image analysis and statistical analysis across multiple gels. This part of proteomics analysis is not represented in current proposals for proteomics standards. We are working with the Proteomics Standards Initiative to develop a model encompassing biological sample origin, experimental protocols, a number of separation techniques and mass spectrometry. The standard format will facilitate the development of central repositories of data, enabling results to be verified or re-analysed, and the correlation of results produced by different research groups using a variety of laboratory techniques.     

Do we want our data raw? Including binary mass spectrometry data in public proteomics data repositories

With the human Plasma Proteome Project (PPP) pilot phase completed, the largest and most ambitious proteomics experiment to date has reached its first milestone. The correspondingly impressive amount of data that came from this pilot project emphasized the need for a centralized dissemination mechanism and led to the development of a detailed, PPP specific data gathering infrastructure at the University of Michigan, Ann Arbor as well as the protein identifications database project at the European Bioinformatics Institute as a general proteomics data repository. One issue that crept up while discussing which data to store for the PPP concerns whether the raw, binary data coming from the mass spectrometers should be stored, or rather the more compact and already significantly processed peak lists. As this debate is not restricted to the PPP but relates to the proteomics community in general, we will attempt to detail the relative merits and caveats associated with centralized storage and dissemination of raw data and/or peak lists, building on the extensive experience gained during the PPP pilot phase. Finally, some suggestions are made for both immediate and future storage of MS data in public repositories.     

Proposed standard for image cytometry data files

A number of different types of computers running a variety of operating systems are presently used for the collection and analysis of image cytometry data. In order to facilitate the development of sharable data analysis programs, to allow for the transport of image cytometry data from one installation to another, and to provide a uniform and controlled means for including textual information in data files, this document describes a data storage format that is proposed as a standard for use in image cytometry. In this standard, data from an image measurement are stored in a minimum of two files. One file is written in ASCII to include information about the way the image data are written and optionally, information about the sample, experiment, equipment, etc. The image data are written separately into a binary file. This standard is proposed with the intention that it will be used internationally for the storage and handling of biomedical image cytometry data. The method of data storage described in this paper is similar to those methods published in American Association of Physicists in Medicine (AAPM) Report Number 10 and in ACR-NEMA Standards Publication Number 300-1985.     

The effect of inosine, inorganic phosphates and pyruvate on erythrocyte glycolysis metabolites under conditions of preservation]

Human erythrocytes were stored as resuspensions in solutions containing citrate (Z), inosine + citrate (I), inosine + phosphate (IP), and inosine + phosphate + pyruvate (IPP). The storage was made at + 4 degrees C for 6 weeks; the initial pH-value amounted to 7.4 at + 4 degrees C. The cellular concentrations of 2.3 DPG, ATP, G6P, FDP and DOAP + GAP were determined. The following results were obtained: 1. During the storage in stored Z-blood the 2.3 DPG concentration will fall below 10% of its initial value; it will remain nearly unchanged in stored I-blood and will increase to 170% in stored IP-blood, to 270% of its initial value in stored IPP-blood. 2. The ATP concentration of cells will fall to about 50% of its initial value at the beginning of the storage of all stored blood. After that it will only increase to about 80% of its initial value in stored IP- and IPP-blood. 3. During the storage the G6P concentration will increase to the highest degree in stored IPP-blood and if high pyruvate concentrations are not present, it will have a reciprocal behaviour towards the FDP and triosephosphate level. The results were discussed in view of the regulation of glycolysis under storage conditions.     

Formation and Maintenance of Robust Long-Term Information Storage in the Presence of Synaptic Turnover

A long-standing problem is how memories can be stored for very long times despite the volatility of the underlying neural substrate, most notably the high turnover of dendritic spines and synapses. To address this problem, here we are using a generic and simple probabilistic model for the creation and removal of synapses. We show that information can be stored for several months when utilizing the intrinsic dynamics of multi-synapse connections. In such systems, single synapses can still show high turnover, which enables fast learning of new information, but this will not perturb prior stored information (slow forgetting), which is represented by the compound state of the connections. The model matches the time course of recent experimental spine data during learning and memory in mice supporting the assumption of multi-synapse connections as the basis for long-term storage.     

SPLASH: systematic proteomics laboratory analysis and storage hub

In the field of proteomics, the increasing difficulty to unify the data format, due to the different platforms/instrumentation and laboratory documentation systems, greatly hinders experimental data verification, exchange, and comparison. Therefore, it is essential to establish standard formats for every necessary aspect of proteomics data. One of the recently published data models is the proteomics experiment data repository [Taylor, C. F., Paton, N. W., Garwood, K. L., Kirby, P. D. et al., Nat. Biotechnol. 2003, 21, 247-254]. Compliant with this format, we developed the systematic proteomics laboratory analysis and storage hub (SPLASH) database system as an informatics infrastructure to support proteomics studies. It consists of three modules and provides proteomics researchers a common platform to store, manage, search, analyze, and exchange their data. (i) Data maintenance includes experimental data entry and update, uploading of experimental results in batch mode, and data exchange in the original PEDRo format. (ii) The data search module provides several means to search the database, to view either the protein information or the differential expression display by clicking on a gel image. (iii) The data mining module contains tools that perform biochemical pathway, statistics-associated gene ontology, and other comparative analyses for all the sample sets to interpret its biological meaning. These features make SPLASH a practical and powerful tool for the proteomics community.     

Effects of Moisture, Temperature, and Time on Seed Germination of Five Wetland Carices: Implications for Restoration

Successful restoration of sedge meadow wetlands is limited by lack of information regarding reintroduction of sedge (Carex) propagules. While restoration from seed is common for prairie restorations, little is known about the germination characteristics of many wetland plants, including sedges. We present the results of a 2.5-year study on seed germination and viability for five species of Carex common to sedge meadow and prairie pothole wetlands in temperate North America. Seed storage and germination conditions were investigated to determine the optimum combination for maintaining seed viability and stimulating germination rates over time. Seeds were germinated under seven different temperature and three moisture regimes after storage for 4, 10, and 14 months under one of four different storage regimes (dry-warm, dry-cold, moist-cold, and wet-cold). The efficacy of short-term wet-cold stratification to stimulate germination of 2.5-year-old seed after long-term dry storage was also investigated. Carex stricta, Carex comosa, and Carex lacustris showed the greatest germination response after wet-cold or moist-cold storage, while Carex lasiocarpa and Carex rostrata showed similar rates of germination after either wet-cold or dry-warm storage. Wet-cold long-term storage was associated with a high level of viability in all five species after 2.5 years. Viability and germination rates were reduced in Carex stricta, Carex comosa, and Carex lasiocarpa after long-term dry-cold storage. Germination rates of seeds stored dry for 2.5 years are not improved by short-term wet-cold treatment in any species tested. Carex seeds should be stored under wet-cold conditions to maintain seed viability over time, thus increasing the likelihood of seeding success for sedge meadow restoration.     

Mass spectrometry in the study of lysosomal storage disorders.

Lysosomal storage disorders represent a group of over 45 distinct genetic diseases, each one resulting from a deficiency of a particular lysosomal protein or, in a few cases, from non-lysosomal proteins that are involved in lysosomal biogenesis. A common biochemical feature of this group of disorders is the accumulation within lysosomes of undegraded or partially degraded substrates that are normally degraded within, and transported out of the lysosome. The particular substrates stored and the site(s) of storage vary with disease type and enzyme/protein deficiency. The nature of the substrate can be used to group the disorders into broad categories including the mucopolysaccharidoses, lipidoses, glycogenoses and oligosaccharidoses. These categories show many clinical similarities within groups as well as significant similarities between groups. For most lysosomal storage disorders the relationship between the stored substrates (type, amount and location) and the disease pathology is not well understood. The use of mass spectrometry and in particular tandem mass spectrometry provides a powerful tool for the investigation of stored substrates in this group of disorders. In this review we will describe the use of mass spectrometry for the analysis of stored substrates. We will discuss progress in the field, limitations of current methods, and summarise issues relating to the diagnosis and treatment of some of the more prevalent lysosomal storage disorders.     

The proteomics standards initiative

The Proteomics Standards Initiative (PSI) aims to define community standards for data representation in proteomics and to facilitate data comparison, exchange and verification. Progress has been made in the development of common standards for data exchange in the fields of both mass spectrometry and protein-protein interaction. A proteomics-specific extension is being created for the emerging American Society for Tests and Measurements mass spectrometry standard, which will be supported by manufacturers of both hardware and software. A data model for proteomics experimentation is under development and discussions on a public repository for published proteomics data are underway. The Protein-Protein Interactions group expects to publish the Level 1 PSI data exchange format for protein-protein interactions soon and discussions as to the content of Level 2 have been initiated.     

The 4th Human Kidney and Urine Proteome Project (HKUPP) Workshop 26 September 2009, Toronto,Canada

The Human Kidney and Urine Proteome Project (HKUPP) was initiated to promote proteomics research in the nephrology field, to better understand kidney functions as well as pathogenic mechanisms of kidney diseases, and to define novel biomarkers and therapeutic targets. The 4th workshop held in September 2009 discussed problems of proteomics analysis for kidney tissues and urine samples and a standard protocol for collection, storage and protein concentration of urine samples was decided upon.     

The role of bioregenerative life-support systems in a manned future in space

Thus far in the manned space program, human life support has depended on storage of air, water, food, and energy. There are no refrigerators on Shuttle, and fresh foods are limited to what can be stowed in lockers for the first 3 days of a mission, when spoilage becomes a factor. Oxygen is stored, CO2 is scrubbed, and water is stored and treated. As we approach the Space Station era, life support will be a combination of storage and resupply. Duty cycles will be 90 days, and physico-chemical (P/C) systems will be important for recycling oxygen and water. Nutritionists seek a capability for refrigerated storage of fresh food on Station. However, most food still will be thermostabilized, rehydratables that can be stored at room temperature. Present Shuttle food is not much more sophisticated than repackaged camp food, and tends to be high in salt content. Hopefully, menus will be healthier on Station, where dietary countermeasures against biomedical responses to chronic microgravity might be implemented, and certainly need to be studied.     

Further steps in standardisation. Report of the second annual Proteomics Standards Initiative Spring Workshop (Siena, Italy 17-20th April 2005)

The spring workshop of the HUPO-PSI convened in Siena to further progress the data standards which are already making an impact on data exchange and deposition in the field of proteomics. Separate work groups pushed forward existing XML standards for the exchange of Molecular Interaction data (PSI-MI, MIF) and Mass Spectrometry data (PSI-MS, mzData) whilst significant progress was made on PSI-MS' mzIdent, which will allow the capture of data from analytical tools such as peak list search engines. A new focus for PSI (GPS, gel electrophoresis) was explored; as was the need for a common representation of protein modifications by all workers in the field of proteomics and beyond. All these efforts are contextualised by the work of the General Proteomics Standards workgroup; which in addition to the MIAPE reporting guidelines, is continually evolving an object model (PSI-OM) from which will be derived the general standard XML format for exchanging data between researchers, and for submission to repositories or journals.     

Transfusion medicine in the era of proteomics

Blood components (BCs) are highly complex mixtures of plasma proteins and cells. At present, BC and blood derivatives (BDs) quality control is mainly focused on standardized quantitative assessment, providing relatively limited information about products. Unfortunately, during the production, inactivation, and storage processes there is the risk of changes in their integrity, especially at the protein level, which could cause negative effects on transfusion. It is therefore a major challenge to identify significant alterations of these products, and, in this context, proteomics can play a potentially relevant role in transfusion medicine (TM) to assess the protein composition of blood-derived therapeutics, particularly for identifying modified proteins. It can provide comprehensive information about changes occurring during processing and storage of BCs and BDs and can be applied to assess or improve them, therefore potentially enabling a global assessment of processing, inactivation and storage methods, as well as of possible contaminants and neoantigens that may influence the immunogenic capacity of blood-derived therapeutics. Thus, proteomics could become a relevant part of quality-control process to verify the identity, purity, safety, and potency of various blood therapeutics. A more detailed understanding of the proteins found in blood and blood products, and the identification of their interactions, may also yield important information for the design of new small molecule therapeutics and also for future improvements in TM.

Proteomics, together with genomics in the near future, will presumably have an impact on disease diagnosis and prognosis as well as on further advances in the production, pathogen inactivation and storage processes of blood-based therapeutics.     

Contribution of proteomics of Leishmania spp. to the understanding of differentiation, drug resistance mechanisms, vaccine and drug development

Leishmania spp., protozoan parasites with a digenetic life cycle, cause a spectrum of diseases in humans. Recently several Leishmania spp. have been sequenced which significantly boosted the number and quality of proteomic studies conducted. Here a historic review will summarize work of the pre-genomic era and then focus on studies after genome information became available. Firstly works comparing the different life cycle stages, in order to identify stage specific proteins, will be discussed. Identifying post-translational modifications by proteomics especially phosphorylation events will be discussed. Further the contribution of proteomics to the understanding of the molecular mechanism of drug resistance and the investigation of immunogenic proteins for the identification of vaccine candidates will be summarized. Approaches of how potentially secreted proteins were identified are discussed. So far 30-35% of the total predicted proteome of Leishmania spp. have been identified. This comprises mainly the abundant proteins, therefore the last section will look into technological approaches on how this coverage may be increased and what the gel-free and gel-based proteomics have to offer will be compared.     

Role and challenges of proteomics in pharma and biotech: technical, scientific and commercial perspective

Contemporary proteomics, currently in its exponential growth phase, is a bewildering array of tools. Proteomic methods are the result of a convergence of rapidly improving mass spectrometry technologies, protein chemistry and separation sciences, genomics and bioinformatics. Strides in improving proteomics technologies to map and measure proteomes and subproteomes are being made. However, no single proteomic platform appears ideally suited to address all research needs or accomplish ambitious goals satisfactorily. However, proteomics is in a unique position to contribute to protein discovery and to public health in terms of better biomarkers, diagnostics and treatment of disease. While the potential is great, many challenges and issues remain to be solved. Fundamental issues, such as biological variability, pre-analytic factors and analytical reproducibility, remain to be resolved. Neither an all-genetic approach nor an all-proteomic approach will solve biological complexity. Proteomics will be the foundation for constructing and extracting useful knowledge to pharma and biotech depicted in the following path: data --> structured data --> information --> information architecture --> knowledge --> useful knowledge.