Sequence Determinants for Amyloid Fibrillogenesis of Human alpha-Synuclein

Parkinson's disease (PD) and dementia with Lewy bodies (DLB) are characterized by the presence of filamentous inclusions in nerve cells. These filaments are amyloid fibrils that are made of the protein α-synuclein, which is genetically linked to rare cases of PD and DLB. β-Synuclein, which shares 60% identity with α-synuclein, is not found in the inclusions. Furthermore, while recombinant α-synuclein readily assembles into amyloid fibrils, β-synuclein fails to do so. It has been suggested that this may be due to the lack in β-synuclein of a hydrophobic region that spans residues 73-83 of α-synuclein. Here, fibril assembly of recombinant human α-synuclein, α-synuclein deletion mutants, β-synuclein and β/α-synuclein chimeras was assayed quantitatively by thioflavin T fluorescence and semi-quantitatively by transmission electron microscopy. Deletion of residues 73-83 from α-synuclein did not abolish filament formation. Furthermore, a chimera of β-synuclein with α-synuclein(73-83) inserted was significantly less fibrillogenic than wild-type α-synuclein. These findings, together with results obtained using a number of recombinant synucleins, showed a correlation between fibrillogenesis and mean β-strand propensity, hydrophilicity and charge of the amino acid sequences. The combination of these simple physicochemical properties with a previously described calculation of β-strand contiguity allowed us to design mutations that changed the fibrillogenic propensity of α-synuclein in predictable ways.     

A pathway separate from the central channel through the nuclear pore complex for inorganic ions and small macromolecules

Nuclear pore complexes (NPCs) are supramolecular nanomachines that mediate the exchange of macromolecules and inorganic ions between the nucleus and the cytosol. Although there is no doubt that large cargo is transported through the centrally located channel, the route of ions and small molecules remains debatable. We thus tested the hypothesis that there are two separate pathways by imaging NPCs using atomic force microscopy, NPC electrical conductivity measurements, and macromolecule permeability assays. Our data indicate a spatial separation between the active transport of macromolecules through the central channel and the passive transport of ions and small macromolecules through the pore periphery.     

Seed-coat microsculpturing and its systematic application in Isatis (Brassicaceae) and allied genera in Iran

In order to examine the systematic application of seed-coat microsculpturing in Isatis, seed surfaces of 23 species (41 populations) in four genera of tribe Isatideae were examined using scanning electron microscopy (SEM). Eight types of basic ornamentation patterns were recognized among the studied specimens. Of these, the reticulate–areolate type was the most common and was found in the genera Isatis, Pachypterygium, Samerari and Tauscheria and 15 species (e.g., I. cappodocica, I. kotschyana and I. tinctoria). The reticulate type, the second most frequent, occurred in 7 species while other types each were represented by only one or 2 species. Although different populations of a given species show similar seed-surface sculpturing in most cases, in some polymorphic species like I. cappadocica and I. kotschyana these patterns were variable among populations. To some extent the variation corresponds to infraspecific taxa for some species, but the differences are not significant enough to be useful in the delimitation of the subspecies recognized by previous workers. Moreover, seed-coat characters do not support the separation of genera Isatis, Pachypterygium, Sameraria and Tauscheria.     

Targeting MAPKs,NF-κB,and PI3K/AKT pathways by methyl palmitate ameliorates ethanol-induced gastric mucosal injury in rats

Excessive drinking of alcohol has been frequently associated with gastric injury; however, its underlying molecular mechanisms have been inadequately investigated. Methyl palmitate (MP) has demonstrated marked hepato-, cardio- and pulmonary protective features; however, its effects on ethanol-induced gastric injury have not been studied. The aim of the present study was to evaluate the potential gastroprotective activity of MP against ethanol-evoked gastric mucosal damage in rats and associated molecular mechanisms, for example, mitogen-activated protein kinases (MAPKs), nuclear factor κB (NF-κB), and phosphoinositide 3 kinase/protein kinase B (PI3K/AKT) pathways. The rat stomachs were examined in terms of the inflammatory, oxidative, and apoptotic perturbations. Current data demonstrated that pretreatment with MP attenuated the gross gastric damage, scores of ulcer index, area of mucosal lesions and histopathology outcomes; actions which were similar to the reference antiulcer omeprazole. MP inhibited NF-κB expression, its nuclear translocation, and the expression of its downstream signals, for example, tumor necrosis factor-α and myeloperoxidase besides restoration of interleukin-10 levels. Western blot analysis revealed that MP counteracted the disruption of MAPKs signaling via lowering p-c-Jun N-terminal kinase 1/2 (p-JNK1/2) expression and restoring the phospho-extracellular signal-regulated kinase 1/2 (p-ERK1/2) levels without affecting p-p38MAPK levels. Additionally, MP improved the antioxidant milieu via diminishing lipid peroxides and enhancing glutathione, glutathione peroxidase, total antioxidant capacity and mucosal nitric oxide. In the context of apoptosis, MP inhibited the cleavage of caspase-3 and poly(ADP-ribose)polymerase (PARP) and Bax protein expression with upregulating B cell lymphoma-2 expression (Bcl-2), thus, promoting gastric cellular survival. This was confirmed by MP activation of the PI3K/AKT pathway manifested by enhanced expression of PI3K p110α and p-AKT. Together, the present findings report the gastroprotective actions of MP mediated via its anti-inflammatory, antioxidant, and antiapoptotic actions. The underlying molecular mechanisms involve, at least partly, the modulation of MAPKs, NF-κB and PI3K/AKT transduction.     

Effect of resveratrol on the radiosensitivity of 5-FU in human breast cancer MCF-7 cells

The aim of this study was to assess the efficacy of resveratrol (Res) on radiosensitivity of 5-fluorouracil (5-FU) in the spheroid culture of MCF-7 breast cancer cell line using colony formation examination. Spheroids on day 9 with 300 µm diameters were treated with 20 µM resveratrol and/or 1 µM 5-FU for one volume doubling time (VDT) (42 hours) and then irradiated with 2 Gy gamma radiation (⁶⁰Co) in various groups. Then the viability of the cells and clonogenic ability were acquired by blue dye exclusion and colony formation assay, respectively. The population doubling time in the monolayer culture and the VDT of spheroid culture was 22.48 0.23 hours and 42 0.63 hours respectively. None of the drugs and combination of them had any effect on the viability of cells. The combination treatment of 5-FU+Res+ radiation significantly reduced the colony formation ability of spheroid cells in comparison with each treatment alone. Our results indicated that resveratrol can significantly decrease colony number of breast cancer spheroid cells treated with 5-FU in combination with gamma-rays. Thus, resveratrol as a hypoxia-inducible factor-1-alpha inhibitor increased the radiosensitization of breast cancer spheroid cells.     

Influence of the construction of porous spargers on lovastatin production by Aspergillus terreus ATCC 20,542 in a laboratory bubble column

Bioprocess and Biosystems Engineering - In bubble column bioreactors, the hydrodynamic behavior like mixing time, bubble size and morphology of filamentous fungi are influenced by the construction...     

Molecular phylogenetic analysis of Arenaria (Caryophyllaceae: tribe Arenarieae) and its allies inferred from nuclear DNA internal transcribed spacer and plastid DNA rps16 sequences

The systematics and phylogeny of the genus Arenaria and allied genera are unresolved. The use of morphological data has resulted in contradictory taxonomic concepts in the past due to their homoplastic nature. We present a phylogenetic analysis based on internal transcribed spacer (ITS) and rps16 sequence data of 140 (132 taxa) and 131 (120 taxa) accessions, respectively. Maximum parsimony and Bayesian analyses of each marker produced nearly congruent trees. Monophyly of Arenaria s.s. and Eremogone is confirmed here. Our results corroborate earlier results indicating that Arenaria subgenus Odontostemma is monophyletic, but outside the core group of Arenaria. Arenaria subgenus Solitaria is sister to Odontostemma and also not closely related to the latter; both of these subgenera are excluded from Arenaria and treated as distinct genera. The molecular data indicate that the ‘Arenaria s.s. clade’ consists of a few well‐supported subgroups and that the current subgeneric classification of the genus does not reflect evolutionary history. Arenaria subgenus Leiosperma is clearly monophyletic, but we reduce it to sectional level. Our molecular data show that the monotypic Arenaria subgenera Porphyrantha and Arenariastrum are nested in A. subgenus Arenaria, whereas subgenus Eremogoneastrum is included in Eremogone. None of the species‐rich sections in subgenus Arenaria is monophyletic. © 2015 The Linnean Society of London, Botanical Journal of the Linnean Society, 2015, 178 , 648–669.     

Nuclear and chloroplast DNA variability and phylogeny of Iranian apples (Malus domestica)

This study was conducted to reveal genetic diversity among 23 local apple genotypes using nuclear (RAPD) and chloroplast DNA (PCR-cpRFLP) markers. Eleven RAPD primers and four cpDNA primer combinations were used in this study. RAPD primers produced a total of 77 polymorphic fragments with an average of seven bands per primer. The percentage of polymorphic bands (68.14 %) showed the efficiency of used RAPD primers in distinguishing all the genotypes considered. Genetic similarity between studied genotypes varied from 0.38 to 0.72 and cluster analysis showed the abundant diversity, indicating high intraspecific genetic variation between Iranian apple genotypes. From the four universal chloroplast primer pairs, three primer pairs amplified the fragments and their combinations showed polymorphic patterns and revealed intraspecific chloroplast variation. The information will facilitate germplasm identification, conservation and new cultivar development.     

Exploring the generality of associations between plant functional traits: evidence within ecological groups along an altitudinal gradient in Hyrcanian forest

We hypothesized that associations among plant functional traits may differ within different ecological assemblages and plant communities. Association among plant traits including plant maximum height, seed weight, fruit type, pollination mode, mean leaf area, and leaf type were explored within life forms, plant strategy groups along with lowland and montane forest vegetation. In total, 83 sampling plots of 400 m² were placed along a 2400 m altitudinal gradient in Hyrcanian forest. Importance‐values of species within vegetation types were used for weighting data and trait associations were explored using categorical principal component analysis. A G‐test and Fisher's exact test of independence were used to retest significance of the correlations. Different paired trait associations (association lines) including height–leaf, height–seed, height–pollination, leaf–seed, seed–fruit and fruit–pollination were observed and their ecological or physiological basis was discussed. Life forms, strategy types and vegetation types differed based on association lines. Some of the well‐known trade‐offs appear by increasing scale from ecological groups to vegetation types in Hyrcanian forest. The observed patterns of trait associations in Hyrcanian forest and several other ecosystems of the world call the generality of previously accepted trait correlations into question.     

Hydrophobic Substitution of Surface Residues Affects Lipase Stability in Organic Solvents

A novel lipase has been recently isolated from a local Pseudomonas sp. (GQ243724). In the present study, we have tried to increase the organic solvent stability of this lipase using site-directed mutagenesis. Eight variants N219L, N219I, N219P, N219A, N219R, N219D, S251L, and S251K were designed to change the surface hydrophobicity of this enzyme with respect to the wild-type. Among these variants, the stability of N219L and N219I significantly increased in the presence of all tested organic solvents, whereas two mutants (N219R and N219D) significantly exhibited decreased stabilities in all the organic solvent studied, suggesting that improvement of hydrophobic patches on the enzyme surface enhances the stability in organic media. Furthermore, replacing Ser²⁵¹ with hydrophobic residues on the enzyme surface dramatically diminished its stability in the tested condition. In spite of the distance of the mutated sites from the active site, the values of k _cat and K _m were affected. Finally, structural analysis of the wild-type and mutated variants was carried out in the presence and absence of some organic solvents using circular dichroism and fluorescence spectroscopy.