Identifying the minimal copper- and zinc-binding site sequence in amyloid-beta peptides

With a combination of complementary experimental techniques, namely sedimentation assay, Fourier transform infrared spectroscopy, and x-ray absorption spectroscopy, we are able to determine the atomic structure around the metal-binding site in samples where amyloid-beta (Abeta) peptides are complexed with either Cu(II) or Zn(II). Exploiting information obtained on a selected set of fragments of the Abeta peptide, we identify along the sequence the histidine residues coordinated to the metal in the various peptides we have studied (Abeta(1-40), Abeta(1-16), Abeta(1-28), Abeta(5-23), and Abeta(17-40)). Our data can be consistently interpreted assuming that all of the peptides encompassing the minimal 1-16 amino acidic sequence display a copper coordination mode that involves three histidines (His(6), His(13), and His(14)). In zinc-Abeta complexes, despite the fact that the metal coordination appears to be more sensitive to solution condition and shows a less rigid geometry around the binding site, a four-histidine coordination mode is seen to be preferred. Lacking a fourth histidine along the Abeta peptide sequence, this geometrical arrangement hints at a Zn(II)-promoted interpeptide aggregation mode.     

Influence of external factors on secondary embryogenesis and germination in somatic embryos from leaves of Quercus suber

Somatic embryogenesis was obtained in cultures of leaves from young seedlings of Quercus suber L. A two-stage process, in which benzyladenine and naphthaleneacetic acid were added first at high and then at low concentrations, was required to initiate the process. Somatic embryos arose when the explants were subsequently placed on medium lacking plant growth regulators. The embryogenic lines remained productive, by means of secondary embryogenesis, on medium without growth regulators. However, this repetitive induction was influenced by the macronutrient composition of the culture medium. Both low total nitrogen content and high reduced nitrogen concentration decreased the percentage of somatic embryos that showed secondary embryogenesis. Our results suggest that alternate culture on medium that increases embryo proliferation and a low salt medium prohibiting embryo formation will partially synchronize embryo development. Chilling slightly reduced secondary embryogenesis but gave a modest increase in germination. Maturation under light followed by storage at 4 °C for at least 30 days gave the best results in switching embryos from an embryogenic pathway to a germinative one. Under these conditions 15% of embryos showed coordinated root and shoot growth and 35% formed either shoots or mostly roots. These percentages were higher than those of embryos matured in darkness. This result indicates that a specific treatment is required after maturation and before chilling to activate the switch from secondary embryo formation to germination.Abbreviations BA benzyladenine - NAA naphthaleneacetic acid - 2,4-D 2,4-dichlorophenoxyacetic acid - BA indolebutyric acid - MS Murashige & Skoog (1962) medium - SH Schenk & Hildebrandt (1972) medium - G Gamborg (1966, PRL-4-C) medium (macronutrients in mg l^–1: NaH₂PO₄·H₂O, 90; Na₂HPO₄, 30; KCl, 300; (NH₄)₂SO₄, 200; MgSO₄·7H₂O, 250; KNO₃, 1000, CaCl₂·2H₂O, 150) - PGR plant growth regulator     

Joint distribution of first exit times of a two dimensional Wiener process with jumps with application to a pair of coupled neurons

Motivated by a neuronal modeling problem, a bivariate Wiener process with two independent components is considered. Each component evolves independently until one of them reaches a threshold value. If the first component crosses the threshold value, it is reset while the dynamics of the other component remains unchanged. But, if this happens to the second component, the first one has a jump of constant amplitude; the second component is then reset to its starting value and its evolution restarts. Both processes evolve once again until one of them reaches again its boundary. In this work, the coupling of the first exit times of the two connected processes is studied.     

The Role of Protein Denaturation Energetics and Molecular Chaperones in the Aggregation and Mistargeting of Mutants Causing Primary Hyperoxaluria Type I

Primary hyperoxaluria type I (PH1) is a conformational disease which result in the loss of alanine:glyoxylate aminotransferase (AGT) function. The study of AGT has important implications for protein folding and trafficking because PH1 mutants may cause protein aggregation and mitochondrial mistargeting. We herein describe a multidisciplinary study aimed to understand the molecular basis of protein aggregation and mistargeting in PH1 by studying twelve AGT variants. Expression studies in cell cultures reveal strong protein folding defects in PH1 causing mutants leading to enhanced aggregation, and in two cases, mitochondrial mistargeting. Immunoprecipitation studies in a cell-free system reveal that most mutants enhance the interactions with Hsc70 chaperones along their folding process, while in vitro binding experiments show no changes in the interaction of folded AGT dimers with the peroxisomal receptor Pex5p. Thermal denaturation studies by calorimetry support that PH1 causing mutants often kinetically destabilize the folded apo-protein through significant changes in the denaturation free energy barrier, whereas coenzyme binding overcomes this destabilization. Modeling of the mutations on a 1.9 Å crystal structure suggests that PH1 causing mutants perturb locally the native structure. Our work support that a misbalance between denaturation energetics and interactions with chaperones underlie aggregation and mistargeting in PH1, suggesting that native state stabilizers and protein homeostasis modulators are potential drugs to restore the complex and delicate balance of AGT protein homeostasis in PH1.     

Cytokine Profiling in Immigrants with Clinical Malaria after Extended Periods of Interrupted Exposure to Plasmodium falciparum

Immunity to malaria is believed to wane with time in the absence of exposure to Plasmodium falciparum infection, but immunoepidemiological data on longevity of immunity remain controversial. We quantified serum cytokines and chemokines by suspension array technology as potential biomarkers for durability of immunity in immigrants with clinical malaria after years without parasite exposure. These were compared to serum/plasma profiles in naïve adults (travelers) and semi-immune adults under continuous exposure, with malaria, along with immigrant and traveler patients without malaria. Immigrants had higher levels of IL-2, IL-5 and IL-8 compared to semi-immune adults with malaria (P≤0.0200). Time since immigration correlated with increased IL-2 (rho=0.2738P=0.0495) and IFN-γ (rho=0.3044P=0.0282). However, immigrants did not show as high IFN-γ concentrations as travelers during a first malaria episode (P<0.0001). Immigrants and travelers with malaria had higher levels of IFN-γ, IL-6, and IL-10 (P<0.0100) than patients with other diseases, and IL-8 and IL-1β were elevated in immigrants with malaria (P<0.0500). Therefore, malaria patients had a characteristic strong pro-inflammatory/Th1 signature. Upon loss of exposure, control of pro-inflammatory responses and tolerance to P. falciparum appeared to be reduced. Understanding the mechanisms to maintain non-pathogenic effector responses is important to develop new malaria control strategies.     

Episodic ataxia type 1 mutations affect fast inactivation of K+ channels by a reduction in either subunit surface expression or affinity for inactivation domain

Episodic ataxia type 1 (EA1) is an autosomal dominant disorder characterized by continuous myokymia and episodic attacks of ataxia. Mutations in the gene KCNA1 that encodes the voltage-gated potassium channel Kv1.1 are responsible for EA1. In several brain areas, Kv1.1 coassembles with Kv1.4, which confers N-type inactivating properties to heteromeric channels. It is therefore likely that the rate of inactivation will be determined by the number of Kv1.4 inactivation particles, as set by the precise subunit stoichiometry. We propose that EA1 mutations affect the rate of N-type inactivation either by reduced subunit surface expression, giving rise to a reduced number of Kv1.1 subunits in heterotetramer Kv1.1-Kv1.4 channels, or by reduced affinity for the Kv1.4 inactivation domain. To test this hypothesis, quantified amounts of mRNA for Kv1.4 or Kv1.1 containing selected EA1 mutations either in the inner vestibule of Kv1.1 on S6 or in the transmembrane regions were injected into Xenopus laevis oocytes and the relative rates of inactivation and stoichiometry were determined. The S6 mutations, V404I and V408A, which had normal surface expression, reduced the rate of inactivation by a decreased affinity for the inactivation domain while the mutations I177N in S1 and E325D in S5, which had reduced subunit surface expression, increased the rate of N-type inactivation due to a stoichiometric increase in the number of Kv1.4 subunits.     

A chromosome-level genome assembly enables the identification of the follicule stimulating hormone receptor as the master sex-determining gene in the flatfish Solea senegalensis

Sex determination (SD) shows huge variation among fish and a high evolutionary rate, as illustrated by the Pleuronectiformes (flatfishes). This order is characterized by its adaptation to demersal life, compact genomes and diversity of SD mechanisms. Here, we assembled the Solea senegalensis genome, a flatfish of great commercial value, into 82 contigs (614 Mb) combining long- and short-read sequencing, which were next scaffolded using a highly dense genetic map (28,838 markers, 21 linkage groups), representing 98.9% of the assembly. Further, we established the correspondence between the assembly and the 21 chromosomes by using BAC-FISH. Whole genome resequencing of six males and six females enabled the identification of 41 single nucleotide polymorphism variants in the follicle stimulating hormone receptor (fshr) consistent with an XX/XY SD system. The observed sex association was validated in a broader independent sample, providing a novel molecular sexing tool. The fshr gene displayed differential expression between male and female gonads from 86 days post-fertilization, when the gonad is still an undifferentiated primordium, concomitant with the activation of amh and cyp19a1a, testis and ovary marker genes, respectively, in males and females. The Y-linked fshr allele, which included 24 nonsynonymous variants and showed a highly divergent 3D protein structure, was overexpressed in males compared to the X-linked allele at all stages of gonadal differentiation. We hypothesize a mechanism hampering the action of the follicle stimulating hormone driving the undifferentiated gonad toward testis.     

Molecular Identification of Candida Species in the Oral Microbiota of Individuals with Down Syndrome: A Case–Control Study

Mycopathologia - Candida species are common in the human oral microbiota and may cause oral candidiasis (OC) when the microbiota equilibrium is disturbed. Immunosuppressed individuals are...     

Plant teratologies as a result of phytoplasma infections

The direct correlation between teratological cases and phytoplasma infections was ascertained in spontaneous and cultivated plant species. Plants, belonging to 31 species and 12 families, showing symptoms of growth abnormalities were collected and analysed. Attempted detection of Rhodococcus fascians by isolation, PCR indexing and 16S rRNA sequencing from fasciated tissues allowed to exclude its presence. Nested PCR by universal primers and 16S rRNA sequence analyses indicated the presence of phytoplasmas, belonging to six groups, in the 44% of symptomatic samples. Among the infected species, Austrocylindropuntia exaltata, Opuntia subulata, Euphorbia characias, Euphorbia dendroides, Euphorbia linifolia, Euphorbia myrsinites, Rumex buchephalophorus, Linaria multicaulis and Fedia cornucopiae represent new phytoplasma hosts world-wide. Moreover this is the first report of phytoplasma belonging to subgroup 16SrRNA II-I in Italy. These findings together with the known erratic distribution in plant tissues of these phloem-restricted prokaryotes indicate a close correlation between fasciation and similar growth disorders and phytoplasma infections.