A novel screening method for microorganisms that efficiently remove heavy metals from aqueous solution

A novel bioassay system for estimating concentrations of several heavy metal ions was carried out with yeast mutants which are highly sensitive to heavy metal ions. The method does not need an atomic adsorption spectrometer or other special equipment. It is suitable for screening of microorganisms that efficiently remove heavy metal ions from aqueous solution.     

Expression of a soybean nodule‐enhanced phosphoenolpyruvate carboxylase gene that shows striking similarity to another gene for a house‐keeping isoform

Three different cDNAs for phosphoenolpyruvate carboxylase (PEPC) were isolated from soybean root nodules. The full-length cDNA of the most abundant isoform (GmPEPC7) was very similar to another one (GmPEPC15), the nucleotide sequence of which is identical to that of a reported clone (gmppc1) (Vazquez-Tello, A., Whittier, R.F., Kawasaki, T., Sugimoto, T., Kawamura, Y., Shibata, D. (1993) Plant Physiol. 103, 1025–1026). In the coding region, the newly isolated GmPEPC7 and the previously reported were gmppc1 99% and 98% identical at the amino acid and nucleotide levels, respectively. In contrast, they exhibited only 39% identity in the 3′ non-coding region, indicating that they are encoded by distinct genes. Northern blot analysis with 3′ non-coding regions as isoform-specific probes showed that GmPEPC7 is nodule-enhanced whereas GmPEPC15 (gmppc1) is expressed in most soybean tissues. The third clone (GmPEPC4) was much less homologous to the above two clones and thus was not further characterized. It was also shown by in situ hybridization that the nodule-enhanced isoform is expressed in all cell types in nodules, including in Bradyrhizobium-infected and uninfected cells and cortical cells. A relatively strong hybridization signal was detected in the vascular bundle pericycle. Southern blot analysis indicated that there are only two PEPC genes exhibiting a high degree of similarity in the soybean genome, one for the nodule-enhanced GmPEPC7 and the other for the constitutively expressed gmppc1. A phylogenetic tree based on the amino acid sequences of soybean PEPCs and nodule-enhanced PEPCs of alfalfa and pea suggested that the soybean nodule-enhanced isoform evolved from the housekeeping PEPC gene after the ureid-translocating and amide-translocating legumes diverged from each other.     

Effect of [His7]-corazonin on the number of antennal sensilla in Locusta migratoria

Abstract.  Certain types of antennal sensilla are known to be more abundant in solitarious individuals than in gregarious ones in the migratory locust, Locusta migratoria . We tested the hypothesis that injection of a neurohormone, [His⁷]-corazonin, into isolated-reared nymphs of this species mimics the effect of crowding on the frequencies of various types of antennal sensilla. One nmol of the hormone was injected into nymphs on two occasions, on the third days of the second and third stadia, respectively. Upon adult emergence, the numbers of different types of sensilla on the eighth antennal segment were compared with those of oil-injected controls. [His⁷]-corazonin did not influence the numbers of basiconic sensilla type A, basiconic sensilla type B and trichoid sensilla significantly compared to oil-injected controls. However, the number of coeloconic sensilla was reduced significantly by the hormone injections. Because the length of the antennal segment was not affected by the hormone injection, it appears that the hormone influenced the development of coeloconic sensilla. The results support the hypothesis tested and are consistent with the idea that [His⁷]-corazonin plays an important role in the control of phase polymorphism in L. migratoria .     

Increased Serum LIGHT Levels Correlate with Disease Progression and Severity of Interstitial Pneumonia in Patients with Dermatomyositis: A Case Control Study

BackgroundActivated CD8+ T cells play an important role in the pathogenesis of dermatomyositis (DM) with interstitial pneumonia (IP). Serum CD8+ T-cell activator, LIGHT, and Th1/Th2/Th17 cytokines were measured in DM-IP patients and compared with clinical parameters to investigate their usefulness.MethodsThe correlations between the clinical findings and serum LIGHT and Th1/Th2/Th17 cytokine levels were investigated in 21 patients with DM-IP (14 with rapidly progressive IP [RPIP] and 7 with chronic IP [CIP], including 4 fatal cases of IP).ResultsThe median serum LIGHT level was 119 (16–335.4) pg/ml, which was higher than that in healthy control subjects and DM patients without IP. The median serum IL–6 level was 14.7 (2.4–154.5) pg/ml (n = 13). The other cytokines were detected in only a few patients. The median serum LIGHT level in DM-RPIP patients (156 [49.6–335.4] pg/ml) was significantly higher than that in DM-CIP patients (94.3 [16–164.2] pg/ml) (P = 0.02). The serum IL–6 level did not correlate with either progression or outcome of DM-IP. ROC curve analysis determined a serum LIGHT level of ≥120 pg/ml to be the cut-off value for the rapid progression of DM-IP. Serum LIGHT levels correlated significantly with %DLco (R = 0.55, P = 0.04) and total ground-glass opacity scores (R = 0.72, P = 0.0002). The serum LIGHT level significantly decreased to 100.5 (12.4–259.3) pg/ml 4 weeks after treatment initiation (P = 0.04).ConclusionsThe serum LIGHT level may be a promising marker of disease progression and severity in patients with DM-IP.     

Non-synonymous FGD3 Variant as Positional Candidate for Disproportional Tall Stature Accounting for a Carcass Weight QTL (CW-3) and Skeletal Dysplasia in Japanese Black Cattle

Recessive skeletal dysplasia, characterized by joint- and/or hip bone-enlargement, was mapped within the critical region for a major quantitative trait locus (QTL) influencing carcass weight; previously named CW-3 in Japanese Black cattle. The risk allele was on the same chromosome as the Q allele that increases carcass weight. Phenotypic characterization revealed that the risk allele causes disproportional tall stature and bone size that increases carcass weight in heterozygous individuals but causes disproportionately narrow chest width in homozygotes. A non-synonymous variant of FGD3 was identified as a positional candidate quantitative trait nucleotide (QTN) and the corresponding mutant protein showed reduced activity as a guanine nucleotide exchange factor for Cdc42. FGD3 is expressed in the growth plate cartilage of femurs from bovine and mouse. Thus, loss of FDG3 activity may lead to subsequent loss of Cdc42 function. This would be consistent with the columnar disorganization of proliferating chondrocytes in chondrocyte-specific inactivated Cdc42 mutant mice. This is the first report showing association of FGD3 with skeletal dysplasia.     

The Centrosomal Linker and Microtubules Provide Dual Levels of Spatial Coordination of Centrosomes

The centrosome is the principal microtubule organizing center in most animal cells. It consists of a pair of centrioles surrounded by pericentriolar material. The centrosome, like DNA, duplicates exactly once per cell cycle. During interphase duplicated centrosomes remain closely linked by a proteinaceous linker. This centrosomal linker is composed of rootletin filaments that are anchored to the centrioles via the protein C-Nap1. At the onset of mitosis the linker is dissolved by Nek2A kinase to support the formation of the bipolar mitotic spindle. The importance of the centrosomal linker for cell function during interphase awaits characterization. Here we assessed the phenotype of human RPE1 C-Nap1 knockout (KO) cells. The absence of the linker led to a modest increase in the average centrosome separation from 1 to 2.5 μm. This small impact on the degree of separation is indicative of a second level of spatial organization of centrosomes. Microtubule depolymerisation or stabilization in C-Nap1 KO cells dramatically increased the inter-centrosomal separation (> 8 μm). Thus, microtubules position centrosomes relatively close to one another in the absence of linker function. C-Nap1 KO cells had a Golgi organization defect with a two-fold expansion of the area occupied by the Golgi. When the centrosomes of C-Nap1 KO cells showed considerable separation, two spatially distinct Golgi stacks could be observed. Furthermore, migration of C-Nap1 KO cells was slower than their wild type RPE1 counterparts. These data show that the spatial organization of centrosomes is modulated by a combination of centrosomal cohesion and microtubule forces. Furthermore a modest increase in centrosome separation has major impact on Golgi organization and cell migration.     

Display of both N- and C-terminal target fusion proteins on the Aspergillus oryzae cell surface using a chitin-binding module

A novel cell surface display system in Aspergillus oryzae was established by using a chitin-binding module (CBM) from Saccharomyces cerevisiae as an anchor protein. CBM was fused to the N or C terminus of green fluorescent protein (GFP) and the fusion proteins (GFP-CBM and CBM-GFP) were expressed using A. oryzae as a host. Western blotting and fluorescence microscopy analysis showed that both GFP-CBM and CBM-GFP were successfully expressed on the cell surface. In addition, cell surface display of triacylglycerol lipase from A. oryzae (tglA), while retaining its activity, was also successfully demonstrated using CBM as an anchor protein. The activity of tglA was significantly higher when tglA was fused to the C terminus than N terminus of CBM. Together, these results show that CBM used as a first anchor protein enables the fusion of both the N and/or C terminus of a target protein.     

Computational characterization of structural role of the non-active site mutation M36I of human immunodeficiency virus type 1 protease

A prominent characteristic of human immunodeficiency virus type 1 (HIV-1) is its high genetic variability, which generates diversity of the virus and often causes a serious problem of the emergence of drug-resistant mutants. Subtype B HIV-1 is dominant in advanced countries, and the mortality rate due to subtype B HIV-1 has been decreased during the past decade. In contrast, the number of patients with non-subtype B viruses is still increasing in developing countries. One of the reasons for the prevalence of non-subtype B viruses is lack of information about the biological and therapeutic differences between subtype B and non-subtype B viruses. M36I is the most frequently observed polymorphism in non-subtype B HIV-1 proteases. However, since the 36th residue is located at a non-active site of the protease and has no direct interaction with any ligands, the structural role of M36I remains unclear. Here, we performed molecular dynamics (MD) simulations of M36I protease in complex with nelfinavir and revealed the influence of the M36I mutation. The results show that M36I regulates the size of the binding cavity of the protease. The reason for the rare emergence of D30N variants in non-subtype B HIV-1 proteases was also clarified from our computational analysis.