Sigrid Schmitt

Justus-Liebig-Universität Gießen, Giessen, Hesse, Germany

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Publications (23)80.01 Total impact

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    ABSTRACT: Chronic alveolar hypoxia induces vascular remodeling processes in the lung resulting in pulmonary hypertension (PH). However, the mechanisms underlying pulmonary remodeling processes are not fully resolved yet. To investigate functional changes occurring during hypoxia exposure we applied 2-dimensional gel electrophoresis (2-DE) to compare protein expression in lungs from mice subjected to 3 hours of alveolar hypoxia and those kept under normoxic conditions. Already after this short-time period several proteins were significantly regulated. Subsequent analysis by MALDI-MS identified cofilin as one of the most prominently upregulated proteins. The regulation was confirmed by western blotting and its cellular localization was determined by immunohisto- and immunocytochemistry. Interestingly, enhanced cofilin serine 3 phosphorylation was observed after short-term and after chronic hypoxia-induced PH in mice, in pulmonary arterial smooth muscle cells (PASMC) from monocrotaline-induced PH in rats, in lungs of idiopathic pulmonary arterial hypertension patients and in hypoxic or platelet-derived growth factor BB-treated human PASMC. Furthermore, elevated cofilin phosphorylation was attenuated by curative treatment of monocrotaline-induced PH in rats and hypoxia-induced PH in mice with the PDGF-BB receptor antagonist imatinib. In conclusion, short-term hypoxic exposure induced prominent changes in lung protein regulation. These very early changes allowed us to identify potential triggers of PH. Thus, respective 2-DE analysis can lead to the identification of new target proteins for the possible treatment of PH.
    No preview · Article · Jan 2013 · Proteomics
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    ABSTRACT: Idiopathic pulmonary fibrosis (IPF) is a progressive and fatal disease for which no effective therapy exists to date. To identify the molecular mechanisms underlying IPF, we performed comparative proteome analysis of lung tissue from patients with sporadic IPF (n = 14) and human donor lungs (controls, n = 10) using two-dimensional gel electrophoresis and MALDI-TOF-MS. Eighty-nine differentially expressed proteins were identified, from which 51 were up-regulated and 38 down-regulated in IPF. Increased expression of markers for the unfolded protein response (UPR), heat-shock proteins, and DNA damage stress markers indicated a chronic cell stress-response in IPF lungs. By means of immunohistochemistry, induction of UPR markers was encountered in type-II alveolar epithelial cells of IPF but not of control lungs. In contrast, up-regulation of heat-shock protein 27 (Hsp27) was exclusively observed in proliferating bronchiolar basal cells and associated with aberrant re-epithelialization at the bronchiolo-alveolar junctions. Among the down-regulated proteins in IPF were antioxidants, members of the annexin family, and structural epithelial proteins. In summary, our results indicate that IPF is characterized by epithelial cell injury, apoptosis, and aberrant epithelial proliferation.
    No preview · Article · Feb 2011 · Journal of Proteome Research
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    ABSTRACT: The proteome of Eimeria bovis meront I-carrying host cells was analyzed by two-dimensional gel electrophoresis (2DE) at 14 days p.i. and compared to non-infected control cells. A total of 221 protein spots were modulated in their abundance in E. bovis-infected host cells and were subsequently analyzed by matrix-assisted laser desorption ionization time-of-flight mass spectometry (MALDI-TOF-MS). These analyses identified 104 proteins in total with 25 host cell proteins being up-regulated and 79 proteins being down-regulated in E. bovis-infected host cells. Moreover, 20 newly expressed proteins were identified exclusively in E. bovis-infected host cells and were most likely of parasite origin. Parasite-induced differences in protein abundance concerned distinct functional categories, with most proteins being involved in host cell metabolism, cell structure, protein fate and gene transcription. Some of the modulated molecules also indicated regulatory processes on the level of host cell stress response (HSP70, HSP90), host cell apoptosis (caspase 8) and actin elongation/depolymerization (α-actinin-1, gelsonin, tropomodulin-3, transgelin). Since merozoites I were already released shortly after cell sampling, the current data reflect the situation at the end of first merogony. This is the first proteomic approach on E. bovis-infected host cells that was undertaken to gain a rather broad insight into Eimeria-induced host cell modulation. The data processed in this investigation should provide a useful basis for more detailed analyses concerning Eimeria-host cell interactions.
    Full-text · Article · Jan 2011 · Molecular and Biochemical Parasitology
  • Christian Zörb · Sigrid Schmitt · Karl H Mühling
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    ABSTRACT: It is of fundamental importance to understand adaptation processes leading to salt resistance. The initial effects on maize roots in the first hour after the adjustment to saline conditions were monitored to elucidate initial responses. The subsequent proteome change was monitored using a 2-D proteomic approach. We found several new salt-inducible proteins, whose expression has not been previously reported to be modulated by salt. A set of phosphoproteins in maize was detected but only ten proteins were phosphorylated and six proteins were dephosphorylated after the application of 25 mM NaCl for 1 h. Some of the phosphorylated maize proteins such as fructokinase, UDP-glucosyl transferase BX9, and 2-Cys-peroxyredoxine were enhanced, whereas an isocitrate-dehydrogenase, calmodulin, maturase, and a 40-S-ribosomal protein were dephosphorylated after adjustment to saline conditions. The initial reaction of the proteome and phosphoproteome of maize after adjustment to saline conditions reveals members of sugar signalling and cell signalling pathways such as calmodulin, and gave hint to a transduction chain which is involved in NaCl-induced signalling. An alteration of 14-3-3 proteins as detected may change plasma membrane ATPase activity and cell wall growth regulators such as xyloglucane endotransglycosylase were also found to be changed immediately after the adjustment to salt stress.
    No preview · Article · Dec 2010 · Proteomics

  • No preview · Conference Paper · May 2010

  • No preview · Conference Paper · May 2010

  • No preview · Article · Mar 2010 · Pneumologie
  • C. Zörb · S. Schmitt · K. H. Mühling
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    No preview · Conference Paper · Sep 2009
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    ABSTRACT: Staphylococcus aureus Clp ATPases (molecular chaperones) alter normal physiological functions including an aconitase-mediated effect on post-stationary growth, acetate catabolism, and entry into death phase (Chatterjee et al., J. Bacteriol. 2005, 187, 4488-4496). In the present study, the global function of ClpC in physiology, metabolism, and late-stationary phase survival was examined using DNA microarrays and 2-D PAGE followed by MALDI-TOF MS. The results suggest that ClpC is involved in regulating the expression of genes and/or proteins of gluconeogenesis, the pentose-phosphate pathway, pyruvate metabolism, the electron transport chain, nucleotide metabolism, oxidative stress, metal ion homeostasis, stringent response, and programmed cell death. Thus, one major function of ClpC is balancing late growth phase carbon metabolism. Furthermore, these changes in carbon metabolism result in alterations of the intracellular concentration of free NADH, the amount of cell-associated iron, and fatty acid metabolism. This study provides strong evidence for ClpC as a critical factor in staphylococcal energy metabolism, stress regulation, and late-stationary phase survival; therefore, these data provide important insight into the adaptation of S. aureus toward a persister state in chronic infections.
    No preview · Article · Mar 2009 · Proteomics

  • No preview · Article · Feb 2009 · Pneumologie
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    ABSTRACT: Pulmonary hypertension (PH) is a severe disease with a poor prognosis. Different forms of PH are characterized by pronounced vascular remodeling, resulting in increased vascular resistance and subsequent right heart failure. The molecular pathways triggering the remodeling process are poorly understood. We hypothesized that underlying key factors can be identified at the onset of the disease. Thus, we screened for alterations to protein expression in lung tissue at the onset of PH in a mouse model of hypoxia-induced PH. Using 2-dimensional polyacrylamide gel electrophoresis in combination with matrix-assisted laser desorption/ionization time-of-flight analysis, we identified 36 proteins that exhibited significantly altered expression after short-term hypoxic exposure. Among these, Fhl-1, which is known to be involved in muscle development, was one of the most prominently upregulated proteins. Further analysis by immunohistochemistry, Western blot, and laser-assisted microdissection followed by quantitative polymerase chain reaction confirmed the upregulation of Fhl-1, particularly in the pulmonary vasculature. Comparable upregulation was confirmed (1) after full establishment of hypoxia-induced PH, (2) in 2 rat models of PH (monocrotaline-treated and hypoxic rats treated with the vascular endothelial growth factor receptor antagonist SU5416), and (3) in lungs from patients with idiopathic pulmonary arterial hypertension. Furthermore, we demonstrated that regulation of Fhl-1 was hypoxia-inducible transcription factor dependent. Abrogation of Fhl-1 expression in primary human pulmonary artery smooth muscle cells by small-interfering RNA suppressed, whereas Fhl-1 overexpression increased, migration and proliferation. Coimmunoprecipitation experiments identified Talin1 as a new interacting partner of Fhl-1. Protein screening identified Fhl-1 as a novel protein regulated in various forms of PH, including idiopathic pulmonary arterial hypertension.
    Preview · Article · Oct 2008 · Circulation
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    ABSTRACT: The posttranslational modification of extra- and intracellular proteins by non-enzymatic glycation results in the formation of advanced glycation end products (AGEs) in physiological systems and is associated with the loss of protein structure and function. Modification by N (epsilon)-carboxymethyl lysine (CML) correlates with the risk for retinopathy in diabetes mellitus and has been discussed as a marker for the prediction of mortality in hemodialysis patients. AGEing of proteins is particularly increased under hyperglycemia associated with different late complications of diabetes mellitus. Modification of proteins to form AGE residues is significantly more enhanced in patients suffering from chronic renal disease than in hyperglycemia and is associated with increased risk for cardiovascular complications and inflammation in patients with chronic renal insuffiency. In order to identify and define the protein "substrates" for non-enzymatic glycation we used a proteome approach combining two-dimensional gel electrophoresis and immunoblotting with Edman protein sequencing to identify specific CML-modified proteins in human hemofiltrate, which essentially resembles plasma with respect to protein composition. Albumin, Ig kappa chain, prostaglandin D2 synthase, lysozyme C, plasma retinol binding protein and beta-2-microglobulin were identified as the major CML-modified proteins. CML-modified fragments of these proteins were also found in hemofiltrate. All identified proteins have in common that they appeared in hemofiltrate predominantly in their CML-modified form(s). Further studies of the functional roles of proteins identified by this new experimental approach could lead to the development of diagnostic tools to follow the progression of diabetes and contribute to the understanding of the pathogenesis of AGE-related diseases.
    No preview · Article · Feb 2008 · Experimental and Clinical Endocrinology & Diabetes

  • No preview · Article · Jan 2008 · Pneumologie
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    ABSTRACT: Opsoclonus-myoclonus syndrome (OMS) is a rare neurologic disorder comprising the main symptoms of eye-movement disturbances, muscle jerks, and severe ataxia. In children and adults, some cases are associated with a tumor as a paraneoplastic syndrome, whereas in children the paraneoplastic form is almost exclusively associated with neuroblastoma. The detection of autoantibodies in some OMS sera led to the hypothesis that the syndrome is of autoimmune origin. Beside autoantibodies against intracellular proteins, such as anti-Hu, alpha-enolase, and KHSRP, specific binding of autoantibodies to the surface of neuroblastoma cells and cerebellar granular neurons have been found. Antiproliferative and proapoptotic effects of these autoantibodies on neuroblastoma cell lines were noted as well. These results support the concept of a humoral autoimmune process in the pathogenesis of OMS.
    No preview · Article · Oct 2007 · Annals of the New York Academy of Sciences

  • No preview · Article · Jan 2007 · Pneumologie
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    ABSTRACT: The middle-sized (M) surface proteins of hepatitis B virus (HBV) and other orthohepadnaviruses contain a conserved N-glycan in their pre-S2 domain, which is essential for the secretion of viral particles. Recently, we also found O-glycans in the pre-S2 domain of M protein from woodchuck hepatitis virus (WHV) and HBV genotype D. Since the O-glycosylation motif is not conserved in all genotypes of HBV, the glycosylation patterns of HBV genotypes A and C were analysed. Pre-S2 (glyco)peptides were released from HBV-carrier-derived HBV subviral particles by tryptic digestion, purified by reversed-phase HPLC and identified by amino acid and amino-terminal sequence analysis as well as matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS). Pre-S2 N-glycans were characterized by anion-exchange chromatography, methylation analysis and on-target sequential exoglycosidase digestions in combination with MALDI-TOF-MS, demonstrating the presence of partially sialylated diantennary complex-type oligosaccharides in all genotypes examined. Pre-S2 O-glycans were characterized by on-target sequential exoglycosidase digestions in combination with MALDI-TOF-MS. The pre-S2 domain of M protein and, to a minor extent, of L (large) protein from HBV genotype C and D was partially O-glycosylated by Neu5Ac(alpha2-3)Gal(beta1-3)GalNAcalpha- or Gal(beta1-3)GalNAcalpha-units at Thr-37 within a conserved sequence context. Genotype A, containing no Thr at position 37 or 38, was not O-glycosylated. Analytical data further revealed that M protein is mostly amino-terminally acetylated in all examined genotypes and that the terminal methionine is partially oxidized. The findings may be relevant for the secretion and the immunogenicity of HBV.
    Full-text · Article · Aug 2004 · Journal of General Virology
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    ABSTRACT: The biochemical reaction of maize (Zea mays L.) to salt stress at the level of proteins in roots and shoots is shown here for the first time. Maize is considered as a salt-sensitive plant. We used an Na+-excluding maize inbred line and low NaCl concentrations to minimize ion effects. Protein patterns were analyzed using 2D polyacrylamide gel electrophoresis. High, as well as low, NaCl treatment of maize led to an unexpected high number of differentially regulated proteins in roots and shoots. Moderate salt stress (25 mM NaCl) already led to a differential regulation of 31% of shoot proteins and 45% of root proteins, without an effect on the morphology and the Na+ and Cl− concentrations of the plants. High stress (100 mM NaCl) led to an uncontrolled change of more than 80% of the separated proteins. Fourteen proteins which were increased by salt stress were identified by in-gel digestion and peptide mass fingerprinting using a matrix-assisted laser desorption/ionization time-of-flight mass spectrometer (MALDI-TOF). We detected three groups of differentially regulated proteins under low salts stress. (A) proteins which are involved in protein biosynthesis and protein modifications by kinases, (B) enzymes of carbon metabolism, and (C) enzymes of the nitrogen metabolism. Eight of these 14 proteins have been reported in the literature to be differentially regulated under high NaCl stress at the level of transcription, translation or metabolism. According to our data there appears to be no specific adaptation to salt stress in maize at the level of proteins.
    No preview · Article · Jul 2004 · Plant Science
  • Andrea Hermann · Sigrid Schmitt · Albert Jeltsch
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    ABSTRACT: The human Dnmt2 protein is one member of a protein family conserved from Schizosaccharomyces pombe and Drosophila melanogaster to Mus musculus and Homo sapiens. It contains all of the amino acid motifs characteristic for DNA-(Cytosine-C5) methyltransferases, and its structure is very similar to prokaryotic DNA methyltransferases. Nevertheless, so far all attempts to detect catalytic activity of this protein have failed. We show here by two independent assay systems that the purified Dnmt2 protein has weak DNA methyltransferase activity. Methylation was observed at CG sites in a loose ttnCGga(g/a) consensus sequence, suggesting that Dnmt2 has a more specialized role than other mammalian DNA methyltransferases.
    No preview · Article · Sep 2003 · Journal of Biological Chemistry
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    ABSTRACT: Twelve MAbs were generated by immunization of BALB/c mice with plasma-derived hepatitis B virus surface spherical antigen particles subtype ayw2 (HBsAg/ayw2 genotype D). Their epitopes were mapped by analysis of reactivity with plasma-derived HBsAg/ayw2 and HBsAg/adw2 (genotype A) in enzyme immunoassays and blots. Mapping was supported by nested sets of truncated preS2 proteins and preS2 peptides. Five antibodies were S domain-specific, seven were preS2-specific and 11 had a preference for genotype D. According to our data, group I of the three known epitope groups of preS2 has to be divided into IA and IB. Three preS2-specific MAbs forming the new group IA reacted with genotype D residues 3-15 which have not yet been described as an epitope region. IA antibodies strongly inhibited the binding of polymerized human serum albumin. Two antibodies (group II) reacted with the glycosylated N-terminal region of preS2 in plasma-derived HBsAg, but not with a preparation from transfected murine cells. One group III antibody was subtype-specific and reacted with the highly variable preS2 sequence 38-48. Only one antibody (group IB) mapped to the region (old group I) which was believed to be immunodominant and genotype-independent. Geno(sub)type-specific epitopes of preS2 are obviously the immunodominant components of natural HBsAg in BALB/c mice, but these epitopes may be masked by serum albumins in humans. The data may explain why it is difficult to detect anti-preS2 antibodies in human recipients of preS2-containing vaccines, in spite of the preS2 immunodominance in mice.
    Full-text · Article · Mar 2000 · Journal of General Virology
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    ABSTRACT: The surface antigen of hepatitis B virus comprises a nested set of small (S), middle (M), and large (L) proteins, all of which are partially glycosylated in their S domains. The pre-S2 domain, present only in M and L proteins, is further N-glycosylated at Asn-4 exclusively in the M protein. Since the pre-S2 N-glycan appears to play a crucial role in the secretion of viral particles, the M protein may be considered as a potential target for antiviral therapy. For characterization of the pre-S2 glycosylation, pre-S2 (glyco)peptides were released from native, patient-derived hepatitis B virus subviral particles by tryptic digestion, separated from remaining particles, purified by reversed-phase high performance liquid chromatography, and identified by amino acid and N-terminal sequence analysis as well as matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS). Pre-S2 N-glycans were characterized by anion exchange chromatography, methylation analysis, and on target sequential exoglycosidase digestions in combination with MALDI-TOF-MS, demonstrating the presence of partially sialylated diantennary complex-type oligosaccharides. In addition, the pre-S2 domain of M protein, but not that of L protein, was found to be partially O-glycosylated by a Gal(beta1-3)GalNAcalpha-, Neu5Ac(alpha2-3)Gal(beta1-3)GalNAcalpha-, or GalNAcalpha-residue. The respective O-glycosylation site was assigned to Thr-37 by digestion with carboxypeptidases in combination with MALDI-TOF-MS and by quadrupole time-of-flight electrospray mass spectrometry. Analytical data further revealed that about 90% of M protein is N-terminally acetylated.
    No preview · Article · May 1999 · Journal of Biological Chemistry

Publication Stats

672 Citations
80.01 Total Impact Points


  • 2004-2013
    • Justus-Liebig-Universität Gießen
      • • Department of Internal Medicine
      • • Institute of Parasitology
      • • Faculty of Medicine
      Giessen, Hesse, Germany