Uptake kinetics of different arsenic species by Brassica carinata

The time- and concentration-dependent uptake kinetics for arsenate and arsenite were determined in 15-day-old excised roots. In both cases, arsenite showed a mono-phasic influx with the isotherm data fitting a linear model better than a non-linear one. The time- and the concentration-dependent uptake of arsenate displayed a hyperbolic kinetic. Greater uptake of arsenate, compared with arsenite, was found especially at lower external substrate concentrations. Competitive inhibition of uptake with phosphate showed that arsenite and arsenate were taken up by different uptake systems because arsenate uptake was strongly inhibited in the presence of phosphate, whereas arsenite uptake was not affected.     

Ago1 and Ago2 differentially affect cell proliferation, motility and apoptosis when overexpressed in SH-SY5Y neuroblastoma cells

Argonaute are a conserved class of proteins central to the microRNA pathway. We have highlighted a novel and non-redundant function of Ago1 versus Ago2; the two core factors of the miRNA-associated RISC complex. Stable overexpression of Ago1 in neuroblastoma cells causes the cell cycle to slow down, a decrease in cellular motility and a stronger apoptotic response upon UV irradiation. These effects, together with a significant increase in p53 levels, suggest that Ago1 may act as a tumor-suppressor factor, a function also supported by GEO Profiles microarrays that inversely correlate Ago1 expression levels with cell proliferation rates.     

High sensitivity C-reactive protein is associated with lower tibial cartilage volume but not lower patella cartilage volume in healthy women at mid-life

Introduction  

Elevated serum high sensitivity C-reactive protein (hsCRP) has been reported in established osteoarthritis (OA). The aim of this study was to determine whether serum levels of hsCRP are associated with the variation in tibial and patella cartilage volumes in women without evidence of OA.     

The role of conserved residues of chagasin in the inhibition of cysteine peptidases

We have evaluated the roles of key amino acids to the action of the natural inhibitor chagasin of papain-family cysteine peptidases. A W93A substitution decreased inhibitor affinity for human cathepsin L 100-fold, while substitutions of T31 resulted in 10-100-fold increases in the K(i) for cruzipain of Trypanosoma cruzi. A T31A/T32A double mutant had increased affinity for cathepsin L but not for cruzipain, while the T31-T32 deletion drastically affected inhibition of both human and parasite peptidases. These differential effects reflect the occurrence of direct interactions between chagasin and helix 8 of cathepsin L, interactions that do not occur with cruzipain.     

The primate-specific protein TBC1D3 is required for optimal macropinocytosis in a novel ARF6-dependent pathway

The generation of novel genes and proteins throughout evolution has been proposed to occur as a result of whole genome and gene duplications, exon shuffling, and retrotransposition events. The analysis of such genes might thus shed light into the functional complexity associated with highly evolved species. One such case is represented by TBC1D3, a primate-specific gene, harboring a TBC domain. Because TBC domains encode Rab-specific GAP activities, TBC-containing proteins are predicted to play a major role in endocytosis and intracellular traffic. Here, we show that the TBC1D3 gene originated late in evolution, likely through a duplication of the RNTRE locus, and underwent gene amplification during primate speciation. Despite possessing a TBC domain, TBC1D3 is apparently devoid of Rab-GAP activity. However, TBC1D3 regulates the optimal rate of epidermal growth factor–mediated macropinocytosis by participating in a novel pathway involving ARF6 and RAB5. In addition, TBC1D3 binds and colocalize to GGA3, an ARF6-effector, in an ARF6-dependent manner, and synergize with it in promoting macropinocytosis, suggesting that the two proteins act together in this process. Accordingly, GGA3 siRNA-mediated ablation impaired TBC1D3-induced macropinocytosis. We thus uncover a novel signaling pathway that appeared after primate speciation. Within this pathway, a TBC1D3:GGA3 complex contributes to optimal propagation of signals, ultimately facilitating the macropinocytic process.     

In Vitro Evaluation of Putative Virulence Attributes of Oral Isolates of <Emphasis Type="Italic">Candida</Emphasis> spp. Obtained from Elderly Healthy Individuals

Identification of Candida isolates obtained from oral cavity of elderly healthy individuals revealed the predominance of non-albicans Candida species (88.9%) compared to Candida albicans (11%). CHROMagar Candida differential medium and PCR revealed the presence of Candida tropicalis (33.3%), Candida glabrata (27.8%), and Candida krusei (16.7%). We investigated the presence of virulence attributes in a total of 18 isolates, including acid protease and phospholipase production, hemolytic activity, and biofilm production. Extracellular protease was found in five isolates (27.8%) whereas extracellular phospholipase was found in three isolates (17%). All isolates showed hemolytic activity. About 56% of the isolates were weakly positive for biofilm formation (score +) whereas a minority (5.6%) of them showed strong biofilm formation (score 4+). Susceptibility in vitro of the isolates to fluconazole was carried out by microdilution method. Fluconazole showed a strong inhibition against most buccal isolates. The resistant isolates were 2 C. tropicalis, 2 C. glabrata, and 1 C. krusei.     

Molecular interactions of ASPP1 and ASPP2 with the p53 protein family and the apoptotic promoters PUMA and Bax

The apoptosis stimulating p53 proteins, ASPP1 and ASPP2, are the first two common activators of the p53 protein family that selectively enable the latter to regulate specific apoptotic target genes, which facilitates yes yet unknown mechanisms for discrimination between cell cycle arrest and apoptosis. To better understand the interplay between ASPP- and p53-family of proteins we investigated the molecular interactions between them using biochemical methods and structure-based homology modelling. The data demonstrate that: (i) the binding of ASPP1 and ASPP2 to p53, p63 and p73 is direct; (ii) the C-termini of ASPP1 and ASPP2 interact with the DNA-binding domains of p53 protein family with dissociation constants, K_d, in the lower micro-molar range; (iii) the stoichiometry of binding is 1:1; (iv) the DNA-binding domains of p53 family members are sufficient for these protein–protein interactions; (v) EMSA titrations revealed that while tri-complex formation between ASPPs, p53 family of proteins and PUMA/Bax is mutually exclusive, ASPP2 (but not ASPP1) formed a complex with PUMA (but not Bax) and displaced p53 and p73. The structure-based homology modelling revealed subtle differences between ASPP2 and ASPP1 and together with the experimental data provide novel mechanistic insights.