Heme biosynthesis in Methanosarcina barkeri via a pathway involving two methylation reactions

The methanogenic archaeon Methanosarcina barkeri synthesizes protoheme via precorrin-2, which is formed from uroporphyrinogen III in two consecutive methylation reactions utilizing S-adenosyl-L-methionine. The existence of this pathway, previously exclusively found in the sulfate-reducing delta-proteobacterium Desulfovibrio vulgaris, was demonstrated for M. barkeri via the incorporation of two methyl groups from methionine into protoheme.     

Visualizing cytoskeleton dynamics in mammalian cells using a humanized variant of monomeric red fluorescent protein

Fluorescent proteins are versatile tools for live cell imaging studies. In particular, recent progress was achieved in the development of monomeric red fluorescent proteins (mRFPs) that show improved properties in respect to maturation and intracellular fluorescence. mRFPmars, a red fluorescent protein designed especially for the use in Dictyostelium, proved to be a brilliant label for different cytoskeletal elements. Here we report on the synthesis of a humanized version of a monomeric RFP, mRFPruby, which differs in sequence from mRFPmars in four amino acids and has a codon usage that is optimized for the application in mammalian cells. In order to demonstrate the usefulness of this new mRFP variant, mRFPruby fused to beta-actin was expressed in different mouse cell lines and used to visualize actin cytoskeleton dynamics by live cell microscopy.     

Biochemical,physicochemical and molecular characterization of a genuine 2-Cys-peroxiredoxin purified from cowpea [Vigna unguiculata (L.) Walpers] leaves

Background

Peroxiredoxins have diverse functions in cellular defense-signaling pathways. 2-Cys-peroxiredoxins (2-Cys-Prx) reduce H₂O₂ and alkyl-hydroperoxide. This study describes the purification and characterization of a genuine 2-Cys-Prx from Vigna unguiculata (Vu-2-Cys-Prx).

Methods

Vu-2-Cys-Prx was purified from leaves by ammonium sulfate fractionation, chitin affinity and ion exchange chromatography.

Results

Vu-2-Cys-Prx reduces H₂O₂ using NADPH and DTT. Vu-2-Cys-Prx is a 44 kDa (SDS-PAGE)/46 kDa (exclusion chromatography) protein that appears as a 22 kDa molecule under reducing conditions, indicating that it is a homodimer linked intermolecularly by disulfide bonds and has a pI range of 4.56–4.72; its NH₂-terminal sequence was similar to 2-Cys-Prx from Phaseolus vulgaris (96%) and Populus tricocarpa (96%). Analysis by ESI-Q-TOF MS/MS showed a molecular mass/pI of 28.622 kDa/5.18. Vu-2-Cys-Prx has 8% α-helix, 39% β-sheet, 22% of turns and 31% of unordered forms. Vu-2-Cys-Prx was heat stable, has optimal activity at pH 7.0, and prevented plasmid DNA degradation. Atomic force microscopy shows that Vu-2-Cys-Prx oligomerized in decamers which might be associated with its molecular chaperone activity that prevented denaturation of insulin and citrate synthase. Its cDNA analysis showed that the redox-active Cys⁵² residue and the amino acids Pro⁴⁵, Thr⁴⁹ and Arg¹²⁸ are conserved as in other 2-Cys-Prx.

General significance

The biochemical and molecular features of Vu-2-Cys-Prx are similar to other members of 2-Cys-Prx family. To date, only one publication reported on the purification of native 2-Cys-Prx from leaves and the subsequent analysis by N-terminal Edman sequencing, which is crucial for construction of stromal recombinant 2-Cys-Prx proteins.     

Antiviral and regulatory T cell immunity in a patient with stromal interaction molecule 1 deficiency

Stromal interaction molecule 1 (STIM1) deficiency is a rare genetic disorder of store-operated calcium entry, associated with a complex syndrome including immunodeficiency and immune dysregulation. The link from the molecular defect to these clinical manifestations is incompletely understood. We report two patients with a homozygous R429C point mutation in STIM1 completely abolishing store-operated calcium entry in T cells. Immunological analysis of one patient revealed that despite the expected defect of T cell proliferation and cytokine production in vitro, significant antiviral T cell populations were generated in vivo. These T cells proliferated in response to viral Ags and showed normal antiviral cytotoxicity. However, antiviral immunity was insufficient to prevent chronic CMV and EBV infections with a possible contribution of impaired NK cell function and a lack of NKT cells. Furthermore, autoimmune cytopenia, eczema, and intermittent diarrhea suggested impaired immune regulation. FOXP3-positive regulatory T (Treg) cells were present but showed an abnormal phenotype. The suppressive function of STIM1-deficient Treg cells in vitro, however, was normal. Given these partial defects in cytotoxic and Treg cell function, impairment of other immune cell populations probably contributes more to the pathogenesis of immunodeficiency and autoimmunity in STIM1 deficiency than previously appreciated.     

Intergroup conflicts among chimpanzees in Taï National Park: lethal violence and the female perspective

Intercommunity aggression in chimpanzees and primitive warfare in humans possess striking similarities, such as the common occurrence of large male coalitions, systematic control of territory boundaries, and lethal attacks on isolated individuals from neighboring groups. However, an important apparent contrast is the absence of recurrent peaceful interactions between neighboring groups of chimpanzees. We observed a remarkable range of behavior in intergroup encounters among three habituated communities of chimpanzees in Taï National Park, Côte d'Ivoire. Lethal attacks are documented in these study groups for the first time, as well as year‐long exchanges of parous adult females and peaceful intergroup visits of mothers with infants. Demographic factors, including group size and number of adult males, are shown to affect the nature of intergroup interactions in ways not considered previously. A reconsideration of the difference in intergroup interactions between eastern and western chimpanzees is proposed including a more important consideration of the female's perspective. The inclusion of the new complexities in intergroup interactions in chimpanzees allows new parallels to be drawn with the evolution of primitive warfare in humans. Am. J. Primatol. 70:519–532, 2008. © 2008 Wiley‐Liss, Inc.     

MHC motif viewer

In vertebrates, the major histocompatibility complex (MHC) presents peptides to the immune system. In humans, MHCs are called human leukocyte antigens (HLAs), and some of the loci encoding them are the most polymorphic in the human genome. Different MHC molecules present different subsets of peptides, and knowledge of their binding specificities is important for understanding the differences in the immune response between individuals. Knowledge of motifs may be used to identify epitopes, to understand the MHC restriction of epitopes, and to compare the specificities of different MHC molecules. Algorithms that predict which peptides MHC molecules bind have recently been developed and cover many different alleles, but the utility of these algorithms is hampered by the lack of tools for browsing and comparing the specificity of these molecules. We have, therefore, developed a web server, MHC motif viewer, that allows the display of the likely binding motif for all human class I proteins of the loci HLA A, B, C, and E and for MHC class I molecules from chimpanzee (Pan troglodytes), rhesus monkey (Macaca mulatta), and mouse (Mus musculus). Furthermore, it covers all HLA-DR protein sequences. A special viewing feature, MHC fight, allows for display of the specificity of two different MHC molecules side by side. We show how the web server can be used to discover and display surprising similarities as well as differences between MHC molecules within and between different species. The MHC motif viewer is available at .