The peroxisome represents a ubiquitous single membrane-bound key organelle that executes various metabolic pathways such as fatty acid degradation by ␣-and ␤-oxidation, ether-phospholipid biosynthesis, metabolism of reactive oxygen species, and detoxification of glyoxylate in mammals. To fulfil this vast array of metabolic functions, peroxisomes accommodate ϳ50 different enzymes at least as identified until now. Interest in peroxisomes has been fueled by the discovery of a group of genetic diseases in humans, which are caused by either a defect in peroxisome biogenesis or the deficient activity of a distinct peroxisomal enzyme or transporter. Although this research has greatly improved our understanding of peroxisomes and their role in mammalian metabolism, deeper insight into biochemistry and functions of peroxisomes is required to expand our knowledge of this low abundance but vital organelle. In this work, we used classical subcellular fractionation in combination with MS-based proteomics methodologies to characterize the proteome of mouse kidney peroxisomes. We could identify virtually all known components involved in peroxisomal metabolism and biogenesis. Moreover through protein localization studies by using a quantitative MS screen combined with statistical analyses, we identified 15 new peroxisomal candidates. Of these, we further investigated five candidates by immunocytochemistry, which confirmed their localization in peroxisomes. As a result of this joint effort, we believe to have compiled the so far most comprehensive protein catalogue of mammalian peroxisomes.

Dysferlin is a muscle protein involved in cell membrane repair and its deficiency is associated with muscular dystrophy. We describe that dysferlin is also expressed in leaky endothelial cells. In the normal central nervous system (CNS), dysferlin is only present in endothelial cells of circumventricular organs. In the inflamed CNS of patients with multiple sclerosis (MS) or in animals with experimental autoimmune encephalomyelitis, dysferlin reactivity is induced in endothelial cells and the expression is associated with vascular leakage of serum proteins. In MS, dysferlin expression in endothelial cells is not restricted to vessels with inflammatory cuffs but is also present in noninflamed vessels. In addition, many blood vessels with perivascular inflammatory infiltrates lack dysferlin expression in inactive lesions or in the normal-appearing white matter. In vitro, dysferlin can be induced in endothelial cells by stimulation with tumor necrosis factor-alpha. Hence, dysferlin is not only a marker for leaky brain vessels, but also reveals dissociation of perivascular inflammatory infiltrates and blood-brain barrier disturbance in multiple sclerosis.

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Nuclear-matrix proteins were prepared from different rat and human cells and separated by twodimensional gel electrophoresis. By computer-assisted analysis of the images, two of the proteins were identified as ubiquitously occurring (common) nuclear-matrix proteins, which appeared in tissue-dependent concentrations. The two proteins that originated from human blood mononuclear cells were analyzed further. Tryptic digests of the blotted proteins were analyzed by partial peptide sequencing and matrixassisted laser-desorption ionization-time-of-flight mass spectrometry. The two human common nuclearmatrix proteins were identified as heterogeneous nuclear ribonucleoproteins (hnRNP) H and H' or their variants. Furthermore, mass analysis revealed details on the N terminus of hnRNP H.Keywords: nuclear matrix ; common nuclear-matrix protein ; heterogeneous nuclear ribonucleoprotein ; tissue specificity ; peptide sequencing.The nuclear matrix is operationally defined as the residual nuclear structure that is yielded by sequential treatment of isolated nuclei with detergents, nucleases and buffers of high ionic strength 11 1. It is considered to represent the three-dimensional fibrillar protein structure constituting the framework of the interphase nucleus. In addition to its role in maintaining the nuclear architecture and the higher order structure of chromatin, the nuclear matrix has been reported as being involved in various nuclear activities, such as DNA replication, DNA transcription, RNA processing and steroid-hormone action. Studies on selected cell types and tissues have indicated that some nuclearmatrix proteins are cell and differentiation specific, while others are shared among various cell types [2]. Stuurman et al. [3] have presented evidence for the presence of a common set of polypeptides termed minimal matrix proteins in the nuclear matrices of mouse cells. Nakayasu and Berezney [4] have reported that eight proteins named nuclear matrins are major constituents of the internal nuclear matrix of rat liver cells. Belgrader et al. were the first to determine the sequence of a defined nuclear-matrix protein, matrin 3, by cloning and sequencing [5]. The detection of other common nuclear-matrix proteins, such as nuclear lamin B [3, 41, nucleolar protein B23 [3, 41, or the nuclear-mitotic apparatus protein [6, 71, was mainly based on immunological methods. However, the number and identity of common nuclearmatrix proteins have not been established unequivocally (reviewed in [8, 91).In systematic studies, we are investigating the nuclear-matrix proteins of various human and rat cells and tissues. ComputerCorrespondence to G. Sauermann,

Nuclear matrix proteins have been defined as insoluble residual proteins resulting from treatment of isolated nuclei with nucleases, detergents and high ionic strength buffers. They are considered as in part representing the proteins constituting the three-dimensional framework of the interphase nucleus. Though cell-specific nuclear matrix proteins have been differentiated from ubiquitously occurring (common) nuclear matrix proteins, the number and types of common nuclear matrix proteins have not yet been unequivocally established. In the present study nuclear matrix proteins were prepared from isolated nuclei of rat kidney, liver, lung, spleen and testes. The matrix proteins were separated by two-dimensional (2-D) electrophoresis and silver stained. Then the spot patterns were compared by computer-assisted image analysis. Composite images were derived for nuclear matrix proteins of individual tissues. Finding between 396-483 spots per tissue, a total of 964 individual spots were registered. Of these, 102 were common nuclear matrix proteins, as appearing in each of the tissue-characteristic images. The apparent molecular mass and pI data may serve for further identification of these nuclear proteins.