Proinflammatory cytokines promote glial heme oxygenase-1 expression and mitochondrial iron deposition: implications for multiple sclerosis

Proinflammatory cytokines, pathological iron deposition, and oxidative stress have been implicated in the pathogenesis of multiple sclerosis (MS) and experimental autoimmune encephalomyelitis (EAE). HO-1 mRNA levels and mitochondrial uptake of [(55)Fe]Cl(3)-derived iron were measured in rat astroglial cultures exposed to interleukin-1beta (IL-1beta) or tumor necrosis factor-alpha (TNF-alpha) alone or in combination with the heme oxygenase-1 (HO-1) inhibitors, tin mesoporphyrin (SnMP) or dexamthasone (DEX), or interferon beta1b (INF-beta). HO-1 expression in astrocytes was evaluated by immunohistochemical staining of spinal cord tissue derived from MS and control subjects. IL-1beta or TNF-alpha promoted sequestration of non-transferrin-derived (55)Fe by astroglial mitochondria. HO-1 inhibitors, mitochondrial permeability transition pore (MTP) blockers and antioxidants significantly attenuated cytokine-related mitochondrial iron sequestration in these cells. IFN-beta decreased HO-1 expression and mitochondrial iron sequestration in IL-1beta- and TNF-alpha-challenged astroglia. The percentage of astrocytes coexpressing HO-1 in affected spinal cord from MS patients (57.3% +/- 12.8%) was significantly greater (p < 0.05) than in normal spinal cord derived from controls subjects (15.4% +/- 8.4%). HO-1 is over-expressed in MS spinal cord astroglia and may promote mitochondrial iron deposition in MS plaques. In MS, IFN-beta may attenuate glial HO-1 gene induction and aberrant mitochondrial iron deposition accruing from exposure to proinflammatory cytokines.     

Recognition of nucleic acid bases and base-pairs by hydrogen bonding to amino acid side-chains

Sequence-specific protein-nucleic acid recognition is determined, in part, by hydrogen bonding interactions between amino acid side-chains and nucleotide bases. To examine the repertoire of possible interactions, we have calculated geometrically plausible arrangements in which amino acids hydrogen bond to unpaired bases, such as those found in RNA bulges and loops, or to the 53 possible RNA base-pairs. We find 32 possible interactions that involve two or more hydrogen bonds to the six unpaired bases (including protonated A and C), 17 of which have been observed. We find 186 "spanning" interactions to base-pairs in which the amino acid hydrogen bonds to both bases, in principle allowing particular base-pairs to be selectively targeted, and nine of these have been observed. Four calculated interactions span the Watson-Crick pairs and 15 span the G:U wobble pair, including two interesting arrangements with three hydrogen bonds to the Arg guanidinum group that have not yet been observed. The inherent donor-acceptor arrangements of the bases support many possible interactions to Asn (or Gln) and Ser (or Thr or Tyr), few interactions to Asp (or Glu) even though several already have been observed, and interactions to U (or T) only if the base is in an unpaired context, as also observed in several cases. This study highlights how complementary arrangements of donors and acceptors can contribute to base-specific recognition of RNA, predicts interactions not yet observed, and provides tools to analyze proposed contacts or design novel interactions.     

A monoclonal antibody to viral glycoprotein blocks virus-immune effector T cells operating at H-2Dd but not at H-2Kd1.

A particular monoclonal antibody that binds to the influenza virus HA molecule inhibits HA-specific thymus-derived lymphocytes mediating cytotoxicity in the context of H-2Dd but not of H-2Kd. Another monoclonal antibody blocks both sets of HA-specific effector T cells. This observation, together with related findings from other laboratories, is considered to support the idea that T cell recognition is directed against some association of viral and H-2 glycoproteins, as proposed in the original formulation of the "altered self" concept.     

The estrogen-receptor complex is bound at unusual chromatin regions

The estrogen receptor of MCF-7 cells labeled with high specific activity estradiol was used to mark the chromatin binding sites for this regulatory molecule. Many of these sites are especially sensitive to nuclease, and produce on digestion a series of uniquely sedimenting products. Several of these have been examined in some detail in this paper. These include a form of receptor that sediments in trace digests at 9S but in more extensive digests at 7S, fast mononucleosomes of about 12.5S, and a species at 15S. Two components of digests, fast mononucleosomes and dinucleosomes were isolated and subjected to further digestion. Much of the hormone on these isolated particles was found to be sensitive to additional hydrolysis, although some was nuclease resistant. It appears that a major fraction of the hormone receptor complexes bound to MCF-7 cell chromatin occurs at nucleosome-free regions which can be detected as transient hydrolysis intermediates.     

Structure of the bundle-forming pilus from enteropathogenic Escherichia coli

Bundle-forming pili (BFP) are essential for the full virulence of enteropathogenic Escherichia coli (EPEC) because they are required for localized adherence to epithelial cells and auto-aggregation. We report the high resolution structure of bundlin, the monomer of BFP, solved by NMR. The structure reveals a new variation in the topology of type IVb pilins with significant differences in the composition and relative orientation of elements of secondary structure. In addition, the structural parameters of native BFP filaments were determined by electron microscopy after negative staining. The solution structure of bundlin was assembled according to these helical parameters to provide a plausible atomic resolution model for the BFP filament. We show that EPEC and Vibriocholerae type IVb pili display distinct differences in their monomer subunits consistent with data showing that bundlin and TcpA cannot complement each other, but assemble into filaments with similar helical organization.     

Screening in vivo for RNA-binding peptides from combinatorial libraries.

We have modified a previously developed genetic assay system for RNA-polypeptide interactions in a attempt to more readily identify RNA-binding peptides. The first modification involved the design of a "complex" library that would contain a variety of RNA-binding polypeptides. The second modification involved the use of neomycin phosphotransferase (NPT II) as the reporter gene, therefore allowing "selection" of RNA-binding peptides by kanamycin resistance. The improved screening system should allow the identification of peptides that bind to a variety of RNA structures.     

A protein arginine N-methyltransferase 1 (PRMT1) and 2 heteromeric interaction increases PRMT1 enzymatic activity

Protein arginine N-methyltransferases (PRMTs) act in signaling pathways and gene expression by methylating arginine residues within target proteins. PRMT1 is responsible for most cellular arginine methylation activity and can work independently or in collaboration with other PRMTs. In this study, we demonstrate a direct interaction between PRMT1 and PRMT2 using co-immunoprecipitation, bimolecular fluorescence complementation, and enzymatic assays. As a result of this interaction, PRMT2 stimulated PRMT1 activity, affecting its apparent V(max) and K(M) values in vitro and increasing the production of methylarginines in cells. Active site mutations and regional deletions from PRMT1 and -2 were also investigated, which demonstrated that complex formation required full-length, active PRMT1. Although the inhibition of methylation by adenosine dialdehyde prevented the interaction between PRMT1 and -2, it did not prevent the interaction between PRMT1 and a truncation mutant of PRMT2 lacking its Src homology 3 (SH3) domain. This result suggests that the SH3 domain may mediate an interaction between PRMT1 and -2 in a methylation-dependent fashion. On the basis of our findings, we propose that PRMT1 serves as the major methyltransferase in cells by forming higher-order oligomers with itself, PRMT2, and possibly other PRMTs.     

Copper association with iron sulfide magnetosomes in a magnetotactic bacterium

Greigite (Fe₃S₄) and pyrite (FeS₂) particles in the magnetosomes of a many-celled, magnetotactic prokaryote (MMP), common in brackish-to-marine, sulfidic, aquatic habitats, contained relatively high concentrations of copper which ranged from about 0.1 to 10 atomic per cent relative to iron. In contrast, the greigite particles in the magnetosomes of a curved magnetotactic bacterium collected from the same sampling site did not contain significant levels of copper. The ability of the MMP to biomineralize copper within its magnetosomes appeared to be limited to that organism and dependent upon the site from which it was collected. Although the chemical mechanism and physiological function of copper accumulation in the magnetosomes of the MMP is unclear, the presence of copper is the first evidence that another transition metal ion could be incorporated in the mineral phase of the magnetosomes of a magnetotactic bacterium.Abbreviation MMP many-celled magnetotactic prokaryote