A disappearing oasis in the semi-arid Chaco: Deficient palm regeneration and establishment

Palm forests of Copernicia alba are a rare habitat in the semi-arid Chaco of Northwestern Argentina, are centres of high species diversity, and provide key resources for many species. Our goal was to assess the conservation status of five C. alba patches in Northwestern Argentina: Reserve; Embarcación; Palma Sola; Vinalito; and, Talar. We compared patches to identify the sites with greatest conservation needs based on four criteria: population size structure; palm density (of individuals with height >1.30 m); probability of seedlings being browsed; and, presence of potential seed dispersers. We found that three (Embarcación, Reserve, and Talar) out of five sites had palm densities greater than 200 individuals/ha and only one site (Embarcación) showed a reverse J-shaped size structure for height. Reserve and Embarcación had the greatest probability of seedlings being browsed (0.99 ± 0.01 and 0.88 ± 0.12, respectively). A total of 14 potential disperser species of mammals and birds were recorded across the five sites. Only Reserve harboured all of the potential dispersers, but at least two potential disperser species were recorded at the other sites. None of the palm patches studied had an adequate conservation status. However, Embarcación met three out of four criteria, and therefore it can be considered to have the best conservation status in the semi-arid Chaco. Palma Sola and Vinalito have the greatest conservation needs. To conserve C. alba in Northwestern Argentina, strategies are needed that ensure seedling establishment for future populations to reach an adequate density and structure.     

Testing Models of the APC Tumor Suppressor/β-Catenin Interaction Reshapes Our View of the Destruction Complex in Wnt Signaling

The Wnt pathway is a conserved signal transduction pathway that contributes to normal development and adult homeostasis, but is also misregulated in human diseases such as cancer. The tumor suppressor adenomatous polyposis coli (APC) is an essential negative regulator of Wnt signaling inactivated in >80% of colorectal cancers. APC participates in a multiprotein “destruction complex” that targets the proto-oncogene β-catenin for ubiquitin-mediated proteolysis; however, the mechanistic role of APC in the destruction complex remains unknown. Several models of APC function have recently been proposed, many of which have emphasized the importance of phosphorylation of high-affinity β-catenin-binding sites [20-amino-acid repeats (20Rs)] on APC. Here we test these models by generating a Drosophila APC2 mutant lacking all β-catenin-binding 20Rs and performing functional studies in human colon cancer cell lines and Drosophila embryos. Our results are inconsistent with current models, as we find that β-catenin binding to the 20Rs of APC is not required for destruction complex activity. In addition, we generate an APC2 mutant lacking all β-catenin-binding sites (including the 15Rs) and find that a direct β-catenin/APC interaction is also not essential for β-catenin destruction, although it increases destruction complex efficiency in certain developmental contexts. Overall, our findings support a model whereby β-catenin-binding sites on APC do not provide a critical mechanistic function per se, but rather dock β-catenin in the destruction complex to increase the efficiency of β-catenin destruction. Furthermore, in Drosophila embryos expressing some APC2 mutant transgenes we observe a separation of β-catenin destruction and Wg/Wnt signaling outputs and suggest that cytoplasmic retention of β-catenin likely accounts for this difference.     

Current analytical methods to study plant water extracts: the example of two mushrooms species, Inonotus hispidus and Sparassis crispa

The analytical study of two hot-water extracts from the mushrooms Inonotus hispidus (Bull.) P. Karst and Sparassis crispa Wulf.:Fr was performed by NMR, HPLC-PAD-MS and GC-MS. The simultaneous use of different analytical techniques highlighted the diverse classes of natural products contained in these extracts. This study describes an attempt to adapt a useful phytochemical method to the direct investigation of plant water extracts, which represent the typical traditional manner for the administration of natural remedies. The heritage concerning plant processing procedures, known as traditional pharmaceutical knowledge, could play an important role in future research on medicinal species. This kind of study could be used as an update for current and future perspectives in this research field.     

Crowding alone cannot account for cosolute effect on amyloid aggregation

Amyloid fiber formation is a specific form of protein aggregation, often resulting from the misfolding of native proteins. Aimed at modeling the crowded environment of the cell, recent experiments showed a reduction in fibrillation halftimes for amyloid-forming peptides in the presence of cosolutes that are preferentially excluded from proteins and peptides. The effect of excluded cosolutes has previously been attributed to the large volume excluded by such inert cellular solutes, sometimes termed "macromolecular crowding". Here, we studied a model peptide that can fold to a stable monomeric β-hairpin conformation, but under certain solution conditions aggregates in the form of amyloid fibrils. Using Circular Dichroism spectroscopy (CD), we found that, in the presence of polyols and polyethylene glycols acting as excluded cosolutes, the monomeric β-hairpin conformation was stabilized with respect to the unfolded state. Stabilization free energy was linear with cosolute concentration, and grew with molecular volume, as would also be predicted by crowding models. After initiating the aggregation process with a pH jump, fibrillation in the presence and absence of cosolutes was followed by ThT fluorescence, transmission electron microscopy, and CD spectroscopy. Polyols (glycerol and sorbitol) increased the lag time for fibril formation and elevated the amount of aggregated peptide at equilibrium, in a cosolute size and concentration dependent manner. However, fibrillation rates remained almost unaffected by a wide range of molecular weights of soluble polyethylene glycols. Our results highlight the importance of other forces beyond the excluded volume interactions responsible for crowding that may contribute to the cosolute effects acting on amyloid formation.     

Mathematical modeling of nucleotide excision repair reveals efficiency of sequential assembly strategies

Nucleotide excision repair (NER) requires the concerted action of many different proteins that assemble at sites of damaged DNA in a sequential fashion. We have constructed a mathematical model delineating hallmarks and general characteristics for NER. We measured the assembly kinetics of the putative damage-recognition factor XPC-HR23B at sites of DNA damage in the nuclei of living cells. These and other in vivo kinetic data allowed us to scrutinize the dynamic behavior of the nucleotide excision repair process in detail. A sequential assembly mechanism appears remarkably advantageous in terms of repair efficiency. Alternative mechanisms for repairosome formation, including random assembly and preassembly, can readily become kinetically unfavorable. Based on the model, new experiments can be defined to gain further insight into this complex process and to critically test model predictions. Our work provides a kinetic framework for NER and rationalizes why many multiprotein processes within the cell nucleus show sequential assembly strategy.     

Useful applications of DOSY experiments for the study of mushroom polysaccharides

DOSY analysis has been performed on different mushroom extracts and fractions with the aim to describe a general method for the study of high molecular weight polysaccharides. These NMR experiments can be exploited to monitor the fractionation pathways performed on crude extracts in order to isolate polysaccharides. DOSY can also rapidly verify the purity of the isolated compounds, as well as evaluate their molecular size. In spite of the complexity of DOSY spectra of mixtures, this NMR technique seems to be a valid analytical tool that could be adopted as a routine method for the study of polysaccharides from different sources.