The absence of molybdenum cofactor sulfuration is the primary cause of the flacca phenotype in tomato plants

The molybdenum cofactor (MoCo)-containing enzymes aldehyde oxidase (AO; EC 1.2.3.1) and xanthine dehydrogenase (XDH; EC 1.2.1.37) require for activity a sulfuration step that inserts a terminal sulfur ligand into the MoCo. The tomato flacca mutation was originally isolated as a wilty phenotype due to a lack of abscisic acid (ABA) that is related to simultaneous loss of AO and XDH activities. An expressed sequence tag candidate from tomato was selected on the basis of homology to sulfurases from animals, fungi and the recently isolated Arabidopsis genes LOS5/ABA3. The tomato homologue maps as a single gene to the bottom of chromosome 7, consistent with the genetic location of the flacca mutation. The structure of FLACCA shows a multidomain protein with an N-terminal NifS-like sulfurase domain; a mammal-specific intermediate section; and a C-terminus containing conserved motifs. Prominent among these are molybdopterin oxidoreductases and thioredoxin redox-active centre/iron-sulfur-binding region signatures which may be relevant to the specific sulfuration of MoCo. Indeed, the molecular analysis of flacca identifies the mutation in a highly conserved motif located in the C-terminus. Activity gel assays show that FLACCA is expressed throughout the plant. Transient and stable complementation of flacca and the Arabidopsis aba3 mutants with Aspergillus nidulans hxB and FLACCA yielded full, partial and tissue-specific types of Mo-hydroxylase activities. Restoration of activity in the root alone is sufficient to augment plant ABA content and rectify the wild-type phenotype. Thus the pleiotropic flacca phenotype is due to the loss of activity of enzymes requiring a sulfurated MoCo.     

Different patterns of interleukin-1alpha and interleukin-1beta expression in organs of normal young and old mice

The different physiological roles of interleukin-1alpha (IL-1alpha) and interleukin-1beta (IL-1beta) are not well understood, especially when considering the apparent overlap and redundancy of the two IL-1 molecules. Characterization of IL-1alpha and IL-1beta expression was performed in this study in organs from young and old mice, using immunohistochemistry and ELISA (enzyme-linked immunosorbent assay). The results indicate that organ IL-1alpha and IL-1beta display different patterns of expression: IL-1alpha is manifested more prominently in lymphoreticular organs (lungs, small intestine, spleen, liver), while IL-1beta is more evident in highly specialized and more vulnerable organs, which do not play a leading role in defense against infections and intoxication (heart, brain, skeletal muscle, kidney). This differential expression is more accentuated in old mice, possibly pointing to the special relevance of these cytokines to organ homeostasis in old age. These findings may shed new light on the physiological functions of IL-1alpha and IL-1beta, and may also lead to the development of improved therapeutic approaches, based on the specific manipulation of these cytokines.     

Development of an improved selective agar medium for isolation of Yersinia pestis

Existing media designed for selective isolation of clinically important members of the genus Yersinia were found to be unsatisfactory for the growth and isolation of Yersinia pestis. We report the development of a new selective agar medium (termed BIN) that supports the growth of Y. pestis. The development of the formulation of this medium was based on a fluorescence screening system designed for monitoring bacterial growth on semisolid media, using a green fluorescent protein-expressing strain. High-throughput combinatorial experiments can be conducted for the quantitative evaluation of the effect of different medium components on growth. Generation of fluorescence plots in this system, using microplates, allowed the quantitative evaluation of the growth rate of Y. pestis EV76 cultures in different agar compositions. The final BIN formulation is based on brain heart infusion agar, to which the selective agents irgasan, cholate salts, crystal violet, and nystatin were introduced. It was found that BIN agar is more efficient in supporting colony formation and recovery of Y. pestis than are the conventional semisolid media MacConkey agar and Yersinia-selective agar (cefsulodin-irgasan-novobiocin agar). The advantage of BIN over other media has been also demonstrated in recovering virulent Y. pestis from the mixed bacterial populations found in decaying carcasses of infected mice. The BIN medium is suggested as a selective medium for isolation and recovery of Y. pestis from various backgrounds.     

Effects of the marine unicellular alga Nannochloropsis sp. to reduce the plasma and liver cholesterol levels in male rats fed on diets with cholesterol

The effects of Nannochloropsis were studied on rats consuming hypercholesterolemic diets. The whole biomass and the hexane/ethanol extract increased the plasma and hepatic eicosapentaenoic and docosahexaenoic acids levels, and reduced the cholesterol levels. We also observed a higher level of propionate, and a lower ratio between acetate and propionate. These data suggest the efficacy of Nannochloropsis in reducing cholesterol levels.     

Characteristics of the aerobic respiratory chains of the microaerophiles Campylobacter jejuni and Helicobacter pylori

The respiratory chain enzymes of microaerophilic bacteria should play a major role in their adaptation to growth at low oxygen tensions. The genes encoding the putative NADH:quinone reductases (NDH-1), the ubiquinol:cytochrome c oxidoreductases (bc1 complex) and the terminal oxidases of the microaerophiles Campylobacter jejuni and Helicobacter pylori were analysed to identify structural elements that may be required for their unique energy metabolism. The gene clusters encoding NDH-1 in both C. jejuni and H. pylori lacked nuoE and nuoF, and in their place were genes encoding two unknown proteins. The NuoG subunit in these microaerophilic bacteria appeared to have an additional Fe-S cluster that is not present in NDH-1 from other organisms; but C. jejuni and H. pylori differed from each other in a cysteine-rich segment in this subunit, which is present in some but not all NDH-1. Both organisms lacked genes orthologous to those encoding NDH-2. The subunits of the bc1 complex of both bacteria were similar, and the Rieske Fe-S and cytochrome b subunits had significant similarity to those of Paracoccus denitrificans and Rhodobacter capsulatus, well-studied bacterial bc1 complexes. The composition of the terminal oxidases of C. jejuni and H. pylori was different; both bacteria had cytochrome cbb3 oxidases, but C. jejuni also contained a bd-type quinol oxidase. The primary structures of the major subunits of the cbb3-type (terminal) oxidase of C. jejuni and H. pylori indicated that they form a separate group within the cbb3 protein family. The implications of the results for the function of the enzymes and their adaptation to microaerophilic growth are discussed.     

Reproducibility and complications in gene searches: linkage on chromosome 6, heterogeneity, association, and maternal inheritance in juvenile myoclonic epilepsy

Evidence for genetic influences in epilepsy is strong, but reports identifying specific chromosomal origins of those influences conflict. One early study reported that human leukocyte antigen (HLA) markers were genetically linked to juvenile myoclonic epilepsy (JME); this was confirmed in a later study. Other reports did not find linkage to HLA markers. One found evidence of linkage to markers on chromosome 15, another to markers on chromosome 6, centromeric to HLA. We identified families through a patient with JME and genotyped markers throughout chromosome 6. Linkage analysis assuming equal male-female recombination probabilities showed evidence for linkage (LOD score 2.5), but at a high recombination fraction (theta), suggesting heterogeneity. When linkage analysis was redone to allow independent male-female thetas, the LOD score was significantly higher (4.2) at a male-female theta of.5,.01. Although the overall pattern of LOD scores with respect to male-female theta could not be explained solely by heterogeneity, the presence of heterogeneity and predominantly maternal inheritance of JME might explain it. By analyzing loci between HLA-DP and HLA-DR and stratifying the families on the basis of evidence for or against linkage, we were able to show evidence of heterogeneity within JME and to propose a marker associated with the linked form. These data also suggest that JME may be predominantly maternally inherited and that the HLA-linked form is more likely to occur in families of European origin.