Expression of L3T4 molecules on Lyt-2+, class II-restricted antigen-specific T suppressor lymphocytes

Three bovine serum albumin-specific Lyt-2⁺ T suppressor (Ts) cell clones from CBA/J mice have been analyzed with regard to expression of L3T4 molecules. All three Ts-cell clones can be stained with monoclonal antibodies (mAb) to L3T4. Tested for the two clones restricted to recognition of E^k determinants, antigen-specific proliferation on antigen-presenting cells, but not the proliferation induced by conditioned medium can be inhibited by L314-specific mAb. In a similar way, Ts-cell cytolytic effector functions can be blocked by L3T4-specific mAb. Thus L3T4 structures seem to play a role in Ts-cell functions. Furthermore, the data support the view that L3T4 expression can be a property of class II-restricted T cells irrespective of their Lyt phenotype.     

Increased Abundance of IncP-1β Plasmids and Mercury Resistance Genes in Mercury-Polluted River Sediments: First Discovery of IncP-1β Plasmids with a Complex mer Transposon as the Sole Accessory Element

Although it is generally assumed that mobile genetic elements facilitate the adaptation of microbial communities to environmental stresses, environmental data supporting this assumption are rare. In this study, river sediment samples taken from two mercury-polluted (A and B) and two nonpolluted or less-polluted (C and D) areas of the river Nura (Kazakhstan) were analyzed by PCR for the presence and abundance of mercury resistance genes and of broad-host-range plasmids. PCR-based detection revealed that mercury pollution corresponded to an increased abundance of mercury resistance genes and of IncP-1β replicon-specific sequences detected in total community DNA. The isolation of IncP-1β plasmids from contaminated sediments was attempted in order to determine whether they carry mercury resistance genes and thus contribute to an adaptation of bacterial populations to Hg pollution. We failed to detect IncP-1β plasmids in the genomic DNA of the cultured Hg-resistant bacterial isolates. However, without selection for mercury resistance, three different IncP-1β plasmids (pTP6, pTP7, and pTP8) were captured directly from contaminated sediment slurry in Cupriavidus necator JMP228 based on their ability to mobilize the IncQ plasmid pIE723. These plasmids hybridized with the merRTΔP probe and conferred Hg resistance to their host. A broad host range and high stability under conditions of nonselective growth were observed for pTP6 and pTP7. The full sequence of plasmid pTP6 was determined and revealed a backbone almost identical to that of the IncP-1β plasmids R751 and pB8. However, this is the first example of an IncP-1β plasmid which had acquired only a mercury resistance transposon but no antibiotic resistance or biodegradation genes. This transposon carries a rather complex set of mer genes and is inserted between Tra1 and Tra2.     

Mismatch-induced DNA unbending upon duplex opening

A DNA duplex can be torn open at a specific position by introducing a branch or bulge to create an asymmetric three-way junction (TWJ). The opened duplex manifests a bent conformation (bending angle approximately 60 degrees , relative to the unopened form), which leads to a dramatic decrease in gel electrophoretic mobility. In the presence of a basepair mismatch at the opening position, the DNA backbone becomes less bent and assumes a distorted T-shaped structure, resulting in an increase in polyacrylamide gel electrophoretic mobility. Both conformational changes are confirmed using fluorescence resonance energy transfer experiments and found to be similar to the signature conformational changes of DNA duplex upon MutS protein binding. Our results imply that some structural rearrangements essential for mismatch recognition are achievable without protein interference. The gel electrophoretic mobility data for DNA TWJs with and without base mismatches correlates well with rotational diffusivity, computed by taking into account the conformational change of TWJ induced by base mismatch.     

Opposite fates of the purine metabolite allantoin under water and nitrogen limitations in bread wheat

Plant Molecular Biology - Degradation of nitrogen-rich purines is tightly and oppositely regulated under drought and low nitrogen supply in bread wheat. Allantoin is a key target metabolite for...     

Effect of high temperature and water stress on pollen germination and spikelet fertility in rice

In future climates, rice could more frequently be subjected to simultaneous high temperature and water stress during sensitive developmental stages such as flowering. In this study, five rice genotypes were exposed to high temperature, water stress and combined high temperature and water stress during flowering to quantify their response through spikelet fertility. Microscopic analyses revealed significant differences in anther dehiscence between treatments and genotypes, with a moderately high association with the number of germinated pollen grains on the stigma. There was a strong relationship between spikelet fertility and the number of germinated pollen on stigmas. Although, all three stress treatments resulted in spikelet sterility, high-temperature stress caused the highest sterility in all five genotypes. A cumulative linear decline in spikelet fertility with increasing duration of independent high-temperature stress and in combination with water stress was quantified. Better anther dehiscence, higher in vivo pollen germination, and higher spikelet fertility were observed in both the N22 accessions compared with IR64, Apo and Moroberekan under high temperature, water stress and combined stress, indicating its ability to tolerate multiple abiotic stresses.     

PCR Detection of Oxytetracycline Resistance Genes otr(A) and otr(B) in Tetracycline-Resistant Streptomycete Isolates from Diverse Habitats

A range of European habitats was screened by PCR for detection of the oxytetracycline resistance genes otr(A) and otr(B), found in the oxytetracycline-producing strain Streptomyces rimosus. Primers were developed to detect these otr genes in tetracycline-resistant (Tc^R) streptomycete isolates from environmental samples. Samples were obtained from bulk and rhizosphere soil, manure, activated sludge and seawater. The majority of Tc^R streptomycetes originated from bulk and rhizosphere soil. Fewer Tc^R streptomycetes were isolated from manure and seawater and none from sewage. By PCR, three out of 217 isolates were shown to contain the otr(A) gene and 13 out of 217 the otr(B) gene. Surprisingly, these genes were detected in taxonomic groups not known as tetracycline-producing strains. The majority of the otr gene–carrying strains was assigned to S. exfoliatus or S. rochei and originated from all habitats from which Tc^R streptomycetes were obtained. Our results indicated that the occurrence of otr(A) and otr(B) genes in natural environments was limited and that otr(B), in comparison to otr(A), seemed to be more common.     

Multiplex proteomic approaches to sepsis research: case studies employing new technologies

Sepsis is a multifactorial disease that provides unique challenges to the critical care physician. Diagnosis is hampered by the lack of a quantitative in vitro diagnostic test, instead, it relies on a series of clinical measures. The complex nature of the disease, with involvement of several physiologic systems, suggests a need to simultaneously monitor many clinical parameters. Novel proteomic technologies now exist that enable the multiplex measurement of multiple protein analytes from the same sample. Integration of these analytical measures with patient clinical data may provide the foundation for a better understanding of disease diagnosis, disease progression and the selection of optimal therapeutic regimen. The future challenge is the translation of these multiplex approaches from investigative research to clinical diagnostics for the greatest impact on patient treatment decisions.     

Analysis of the thyrotropin-releasing hormone-degrading ectoenzyme by site-directed mutagenesis of cysteine residues. Cys68 is involved in disulfide-linked dimerization.

Thyrotropin-releasing hormone-degrading ectoenzyme is a member of the M1 family of Zn-dependent aminopeptidases and catalyzes the degradation of thyrotropin-releasing hormone (TRH; Glp-His-Pro-NH2). Cloning of the cDNA of this enzyme and biochemical studies revealed that the large extracellular domain of the enzyme with the catalytically active site contains nine cysteine residues that are highly conserved among species. To investigate the functional role of these cysteines in TRH-DE we used a site-directed mutagenesis approach and replaced individually each cysteine by a serine residue. The results revealed that the proteolytically truncated and enzymatically fully active enzyme consists of two identical subunits that are associated noncovalently by protein-protein interactions but not via interchain S-S bridges. The eight cysteines contained within this region are all important for the structure of the individual subunit and the enzymatic activity, which is dramatically reduced in all mutant enzymes. This is even true for the four cysteines that are clustered within the C-terminal domain remote from the Zn-binding consensus sequence HEICH. In contrast, Cys68, which resides within the stalk region seven residues from the end of the hydrophobic membrane-spanning domain, can be replaced by serine without a significant change in the enzymatic activity. Interestingly, this residue is involved in the formation of an interchain disulfide bridge. Covalent dimerization of the subunits, however, does not seem to be essential for efficient biosynthesis, enzymatic activity and trafficking to the cell surface.