Solution Structure and DNA-binding Properties of the Winged Helix Domain of the Meiotic Recombination HOP2 Protein

The HOP2 protein is required for efficient double-strand break repair which ensures the proper synapsis of homologous chromosomes and normal meiotic progression. We previously showed that in vitro HOP2 shows two distinctive activities: when it is incorporated into a HOP2-MND1 heterodimer, it stimulates DMC1 and RAD51 recombination activities, and the purified HOP2 alone is proficient in promoting strand invasion. The structural and biochemical basis of HOP2 action in recombination are poorly understood; therefore, they are the focus of this work. Herein, we present the solution structure of the amino-terminal portion of mouse HOP2, which contains a typical winged helix DNA-binding domain. Together with NMR spectral changes in the presence of double-stranded DNA, protein docking on DNA, and mutation analysis to identify the amino acids involved in DNA coordination, our results on the three-dimensional structure of HOP2 provide key information on the fundamental structural and biochemical requirements directing the interaction of HOP2 with DNA. These results, in combination with mutational experiments showing the role of a coiled-coil structural feature involved in HOP2 self-association, allow us to explain important aspects of the function of HOP2 in recombination.     

Biodiversity hotspots are not congruent with conservation areas in the Gulf of California

As marine systems are threatened by increasing human impacts, mechanisms to maintain biodiversity and ecosystem functions and services are needed. Protecting areas of conservation importance may serve as a proxy for maintaining these functions, while also facilitating efficient use and management of limited resources. Biodiversity hotspots have been used as surrogates for spatial conservation importance; however, as many protected areas have been established opportunistically and under differing criteria, it is unclear how well they actually protect hotspots. We evaluated how well the current protected area network and priority areas selected through previous systematic conservation planning exercises preserve biodiversity hotspots in the Gulf of California, Mexico. We also determined spatial congruence between biodiversity hotspots based on different criteria, which may determine their ability to be used as surrogates for each other. We focus on the Gulf of California because it is a megadiverse system where limited information regarding species diversity and distribution has constrained development of strategies for conservation and management. We developed a species occurrence database and identified biodiversity hotspots using four different criteria: species richness, rarity, endemism, and threatened species. We interpolated species occurrence, while accounting for heterogeneous sampling efforts. We then assessed overlap of hotspots with existing protected areas and priority areas, and between hotspots derived by distinct criteria. We gathered 286,533 occurrence records belonging to 12,105 unique species, including 6388 species identified as rare, 642 as endemic, and 386 as threatened. We found that biodiversity hotspots showed little spatial overlap with areas currently under protection and previously identified priority areas. Our results highlight the importance of distinct spatial areas of biodiversity and suggest that different ecological mechanisms sustain different aspects of diversity and multiple criteria should be used when defining conservation areas.     

Phosphoproteins analysis in plants: a proteomic approach

The study of phosphoproteome on a global scale represents one of the challenges in the post-genomic era. Here, we propose an integrated procedure starting from the crude protein extract, that consists of sequential purification steps, and ending up in the identification of phosphorylation sites. This involves (i) an enrichment in phosphoproteins with a commercially available chromatography matrix, (ii) a 2-D gel analysis of the enriched fraction followed by the selective staining with the phosphospecific fluorescent dye Pro-Q Diamond, (iii) a phosphopeptide capture, from the tryptic lysate of 2-D spots, using IMAC micro-columns. In the end, the identification of the phosphoproteins and their corresponding phosphorylation sites were achieved by MALDI-TOF-TOF spectrometry. The method was applied to contrasting samples prepared from cell suspension cultures of Arabidopsis thaliana and roots of Medicago truncatula. The results obtained, demonstrated the robustness of the combination of two enrichment stages, sequentially at the protein and at the peptide levels, to analyse phosphoproteins in plants.     

Antifungal activity of soybean and chickpea isoflavones and their reduced derivatives

The fungicidal activity of the isoflavones from soybean (Glycine max) and chickpea (Cicer arietinum) has been studied on three food and forage contaminating fungi, Aspergillus ochraceus, Penicillium digitatum and Fusarium culmorum. The reduced derivatives of the corresponding isoflavones—the isoflavanones and the isoflavans—were also included in the investigation. For the first time in a comparative study it is shown that isoflavones and isoflavanones are variable in their activity whereas the isoflavans are moderately active inhibitors of fungal growth.     

Behavioural method to detect marginal neurotoxic effects of ivermectin in DA/Orl rats.

Ivermectin (22, 23-dihydroavermectin B 1) in subtoxic doses was administered subcutaneously to young adult DA rats. Prior to treatment the rats had been trained in a visual discrimination learning programme until their response pattern was stable. The behavioural response data were recorded during continued discrimination testing following the Ivermectin injection and compared with those of a control group. The results showed that the Ivermectin injection reduced the total number of lever presses and reinforcement collections. Further the treatment caused an increase in the total number of erroneous responses.     

Alterations in the red blood cell membrane proteome in alzheimer's subjects reflect disease-related changes and provide insight into altered cell morphology

Background  

Our earlier studies have shown that red blood cell (RBC) morphology in Alzheimer's disease (AD) subjects was altered (> 15% of the RBCs were elongated as compared to 5.9% in normal controls (p < 0.0001)). These results suggested alterations in the RBC membrane architecture in AD subjects, possibly due to RBC-β-amyloid interactions and/or changes in the expression of membrane proteins. We hypothesized that the observed changes could be due to changes in the level of the protein components of the cytoskeleton and those linked to the RBC membrane. To examine this, we performed a proteomic analysis of RBC membrane proteins of AD subjects, and their age-matched controls using one pool of samples from each group, following their separation by SDS-PAGE, in-gel Tryptic digestion, LC-MS-MS of peptides generated, and a label-free approach of semi-quantitative analysis of their relative MS spectral intensities.     

EBNA3C Augments Pim-1 Mediated Phosphorylation and Degradation of p21 to Promote B-Cell Proliferation

Epstein–Barr virus (EBV), a ubiquitous human herpesvirus, can latently infect the human population. EBV is associated with several types of malignancies originating from lymphoid and epithelial cell types. EBV latent antigen 3C (EBNA3C) is essential for EBV-induced immortalization of B-cells. The Moloney murine leukemia provirus integration site (PIM-1), which encodes an oncogenic serine/threonine kinase, is linked to several cellular functions involving cell survival, proliferation, differentiation, and apoptosis. Notably, enhanced expression of Pim-1 kinase is associated with numerous hematological and non-hematological malignancies. A higher expression level of Pim-1 kinase is associated with EBV infection, suggesting a crucial role for Pim-1 in EBV-induced tumorigenesis. We now demonstrate a molecular mechanism which reveals a direct role for EBNA3C in enhancing Pim-1 expression in EBV-infected primary B-cells. We also showed that EBNA3C is physically associated with Pim-1 through its amino-terminal domain, and also forms a molecular complex in B-cells. EBNA3C can stabilize Pim-1 through abrogation of the proteasome/Ubiquitin pathway. Our results demonstrate that EBNA3C enhances Pim-1 mediated phosphorylation of p21 at the Thr¹⁴⁵ residue. EBNA3C also facilitated the nuclear localization of Pim-1, and promoted EBV transformed cell proliferation by altering Pim-1 mediated regulation of the activity of the cell-cycle inhibitor p21/WAF1. Our study demonstrated that EBNA3C significantly induces Pim-1 mediated proteosomal degradation of p21. A significant reduction in cell proliferation of EBV-transformed LCLs was observed upon stable knockdown of Pim-1. This study describes a critical role for the oncoprotein Pim-1 in EBV-mediated oncogenesis, as well as provides novel insights into oncogenic kinase-targeted therapeutic intervention of EBV-associated cancers.