Differential in vitro inhibition of polyphenoloxidase from a wild edible mushroom Lactarius salmonicolor

The polyphenol oxidase (LsPPO) from a wild edible mushroom Lactarius salmonicolor was purified using a Sepharose 4B-L-tyrosine-p-amino benzoic acid affinity column. At the optimum pH and temperature, the K(M) and V(Max) values of LsPPO towards catechol, 4-methylcatechol and pyrogallol were determined as 0.025 M & 0.748 EU/mL, 1.809 x 10(- 3) M & 0.723 EU/mL and 9.465 x 10(- 3) M & 0.722 EU/mL, respectively. Optimum pH and temperature values of LsPPO for the three substrates above ranged between the pH 4.5-11.0 and 5-50 degrees C. Enzyme activity decreased due to heat denaturation with increasing temperature. Effects of a variety of classical PPO inhibitors were investigated opon the activity of LsPPO using catechol as the substrate. IC(50) values for glutathione, p-aminobenzenesulfonamide, L-cysteine, L-tyrosine, oxalic acid, beta-mercaptoethanol and syringic acid were determined as 9.1 x 10(- 4), 2.3 x 10(- 4) M, 1.5 x 10(- 4) M, 3.8 x 10(- 7) M, 1.2 x 10(- 4) M, 4.9 x 10(- 4) M, and 4 x 10(- 4) M respectively. Thus L-tyrosine was by far the most effective inhibitor. Interestingly, sulfosalicylic acid behaved as an activator of LsPPO in this study.     

<Emphasis Type="Italic">FHIT</Emphasis> Gene Sequence Variants and Reduced Fhit Protein Expression in Glioblastoma Multiforme

Molecular studies have an important role in the elucidation of the mechanisms involved in Glioblastoma multiforme (GBM) development. The occurrence of FHIT gene alterations, which has an important role in different cancers, has not yet been studied well in GBM. We aimed to investigate the occurrence of alterations of FHIT gene sequence and protein expression in the GBMs. Sequence alterations in exons 5–9 of the FHIT gene were screened in 63 GBMs using the single-strand conformational polymorphism method, followed by DNA sequencing. Additionally, the level of Fhit protein expression in tissues of 48 tumors was assessed by immunohistochemistry (IHC). In our investigation, FHIT gene alterations in the coding region were detected in 11 of the 63 GBM cases (17.5%). Two different sequence variants were determined: one novel missense variant (G→C transition at codon 49) and one previously described silent alteration (C→T transition at codon 88). Using web-based programs, such as SIFT and ESEfinder, it was determined that both alterations might have caused significant modification on protein function. In addition, we identified a previously reported an intronic polymorphism (T→A transition at IVS8-17) in 47.5% of cases as a similar rate (45%) in the control group. Moreover, it was observed that Fhit protein expression was reduced in 87.5% of tumors. In conclusion, the reduction or loss of Fhit protein expression by genetic alterations or epigenetic mechanisms in GBM might be associated with brain tumorigenesis.     

Modeling above‐ground litterfall in eastern Mediterranean conifer forests using fractional tree cover,and remotely sensed and ground data

Question: How can we model above‐ground litterfall in Mediterranean conifer forests using remotely sensed and ground data, and geographic information systems (GIS)? Location: Eastern Mediterranean conifer forest of Turkey. Methods: Above‐ground litterfall from Mediterranean forest stands of Pinus nigra, Cedrus libani, Pinus brutia and Juniperus excelsa and mixed Abies cilicica, C. libani and P. nigra was modeled as a function of fractional tree cover using a regression tree algorithm, based on IKONOS and Landsat TM/ETM+data. Landsat TM/ETM+images for the study area were used to map actual stand patterns, based on a land‐cover map of species stands using a supervised classification. Results: Total amount of annual above‐ground litterfall for the entire study area (12 260 km²) was estimated at 417.2 Mg ha^?1 for P. brutia, 291.1 Mg ha^?1 for the mixed stand, 115.5 Mg ha^?1 for P. nigra, 54.6 Mg ha^?1 for J. excelsa and 45.9 Mg ha^?1 for C. libani. The maps generated indicate the distribution of the seasonal amount of total above‐ground litterfall for different species and the distribution of species stands in the study area. There was an increase in the amount of above‐ground litterfall for P. brutia stand in summer, for J. excelsa in autumn and for C. libani, P. nigra and the mixed stand of A. cilicica, P. nigra and C. libani in winter. Conclusion: Application of this model helps to improve the accuracy of estimated litterfall input to soil organic carbon pools in the Mediterranean conifer forests.     

Presenilin-based genetic screens in Drosophila melanogaster identify novel notch pathway modifiers

Presenilin is the enzymatic component of gamma-secretase, a multisubunit intramembrane protease that processes several transmembrane receptors, such as the amyloid precursor protein (APP). Mutations in human Presenilins lead to altered APP cleavage and early-onset Alzheimer's disease. Presenilins also play an essential role in Notch receptor cleavage and signaling. The Notch pathway is a highly conserved signaling pathway that functions during the development of multicellular organisms, including vertebrates, Drosophila, and C. elegans. Recent studies have shown that Notch signaling is sensitive to perturbations in subcellular trafficking, although the specific mechanisms are largely unknown. To identify genes that regulate Notch pathway function, we have performed two genetic screens in Drosophila for modifiers of Presenilin-dependent Notch phenotypes. We describe here the cloning and identification of 19 modifiers, including nicastrin and several genes with previously undescribed involvement in Notch biology. The predicted functions of these newly identified genes are consistent with extracellular matrix and vesicular trafficking mechanisms in Presenilin and Notch pathway regulation and suggest a novel role for gamma-tubulin in the pathway.     

Mechanisms of a ring shaped helicase

Bacteriophage T7 helicase (T7 gene 4 helicase-primase) is a prototypical member of the ring-shaped family of helicases, whose structure and biochemical mechanisms have been studied in detail. T7 helicase assembles into a homohexameric ring that binds single-stranded DNA in its central channel. Using RecA-type nucleotide binding and sensing motifs, T7 helicase binds and hydrolyzes several NTPs, among which dTTP supports optimal protein assembly, DNA binding and unwinding activities. During translocation along single stranded DNA, the subunits of the ring go through dTTP hydrolysis cycles one at a time, and this probably occurs also during DNA unwinding. Interestingly, the unwinding speed of T7 helicase is an order of magnitude slower than its translocation rate along single stranded DNA. The slow unwinding rate is greatly stimulated when DNA synthesis by T7 DNA polymerase is coupled to DNA unwinding. Using the T7 helicase as an example, we highlight critical findings and discuss possible mechanisms of helicase action.     

Nitric oxide levels in patients with psoriasis treated with methotrexate

Psoriasis is a chronic, recurrent, inflammatory, and hyperproliferative disease. Recently there have been studies regarding increases in the levels of NO in inflammatory dermatoses including psoriasis. In this study, 22 patients with psoriasis were scored with PASI (psoriasis area and severity index) and the levels of serum nitrite-nitrate were evaluated before and after therapy with methotrexate (Mtx). The results were compared with age- and sex-matched healthy volunteers. The relation of the results with the clinical severity and the cumulative Mtx dose were also evaluated. The serum levels of nitrite-nitrate of the psoriatic patients with active lesions were found to be significantly higher than the levels of the healthy volunteers and the patients after therapy. The elevated nitrite-nitrate serum levels in the inflammatory period may suggest the possible role of this mediator in the etiopathogenesis of psoriasis and the potential future use of NO inhibitors in the treatment of psoriasis.     

Effects of ether vs. ester linkage on lipid bilayer structure and water permeability

The structure and water permeability of bilayers composed of the ether-linked lipid, dihexadecylphosphatidylcholine (DHPC), were studied and compared with the ester-linked lipid, dipalmitoylphosphaditdylcholine (DPPC). Wide angle X-ray scattering on oriented bilayers in the fluid phase indicate that the area per lipid A is slightly larger for DHPC than for DPPC. Low angle X-ray scattering yields A = 65.1 Å² for DHPC at 48 °C. LAXS data provide the bending modulus, K_C = 4.2 × 10⁻¹³ erg, and the Hamaker parameter H = 7.2 × 10⁻¹⁴ erg for the van der Waals attractive interaction between neighboring bilayers. For the low temperature phases with ordered hydrocarbon chains, we confirm the transition from a tilted L_β′ gel phase to an untilted, interdigitated L_βI phase as the sample hydrates at 20 °C. Our measurement of water permeability, P_f = 0.022 cm/s at 48 °C for fluid phase DHPC is slightly smaller than that of DPPC (P_f = 0.027 cm/s) at 50 °C, consistent with our triple slab theory of permeability.     

The C-type lectin receptor CLECSF8 (CLEC4D) is expressed by myeloid cells and triggers cellular activation through Syk kinase

CLECSF8 is a poorly characterized member of the "Dectin-2 cluster" of C-type lectin receptors and was originally thought to be expressed exclusively by macrophages. We show here that CLECSF8 is primarily expressed by peripheral blood neutrophils and monocytes and weakly by several subsets of peripheral blood dendritic cells. However, expression of this receptor is lost upon in vitro differentiation of monocytes into dendritic cells or macrophages. Like the other members of the Dectin-2 family, which require association of their transmembrane domains with signaling adaptors for surface expression, CLECSF8 is retained intracellularly when expressed in non-myeloid cells. However, we demonstrate that CLECSF8 does not associate with any known signaling adaptor molecule, including DAP10, DAP12, or the FcRγ chain, and we found that the C-type lectin domain of CLECSF8 was responsible for its intracellular retention. Although CLECSF8 does not contain a signaling motif in its cytoplasmic domain, we show that this receptor is capable of inducing signaling via Syk kinase in myeloid cells and that it can induce phagocytosis, proinflammatory cytokine production, and the respiratory burst. These data therefore indicate that CLECSF8 functions as an activation receptor on myeloid cells and associates with a novel adaptor molecule. Characterization of the CLECSF8-deficient mice and screening for ligands using oligosaccharide microarrays did not provide further insights into the physiological function of this receptor.     

Hallermann-Streiff syndrome with hemihypertrophy

We report on a five year old girl with Hallermann-Streiff syndrome and hemihypertrophy. Hemihypertrophy does not appear to have ever been associated with Hallermann-Streiff syndrome.