Functional dissection of osteoprotegerin and its interaction with receptor activator of NF-kappaB ligand

The receptor activator of NF-kappaB (RANK) belongs to the neuregulin/tumor necrosis factor (TNF) receptor superfamily and is activated by RANK ligand (RANK-L), a homotrimeric, TNF-like cytokine. RANK is present on the surface of osteoclast cell precursors, where its interaction with RANK-L induces their terminal differentiation into osteoclasts, thus increasing bone breakdown. The secreted, soluble receptor osteoprotegerin (OPG) interrupts this activation by binding directly to RANK-L. Therefore, osteoclast maturation (and bone homeostasis) is regulated in vivo by OPG levels of expression. We have studied the assembly state and affinity of OPG for RANK-L by sedimentation analyses and surface plasmon resonance (Biacore). Full-length, homodimeric OPG binds to RANK-L with a KD of 10 nM. OPG is also a member of the TNF receptor superfamily and contains four disulfide-rich ligand-binding domains, yet lacks a transmembrane region separating the ligand-binding region from the two death domains, as observed for other receptor family members. We showed that dimerization of OPG results from noncovalent interactions mediated by the death domains and to a lesser extent by a C-terminal heparin-binding region. In contrast, a C-terminal intermolecular disulfide bond does not contribute to the formation or stability of OPG dimers. A truncate of osteoprotegerin, containing the ligand-binding domains but lacking the dimerization domains, bound RANK-L with a KD of approximately 3 microM, indicating that monomer oligomerization for the OPG provides an increase of 3 orders of magnitude in the affinity for RANK-L. Therefore, OPG dimer formation is required for the mechanism of inhibition of the RANK-L/RANK receptor interaction.     

Exome Sequencing Identifies Mutations in CCDC114 as a Cause of Primary Ciliary Dyskinesia

Primary ciliary dyskinesia (PCD) is a genetically heterogeneous, autosomal-recessive disorder, characterized by oto-sino-pulmonary disease and situs abnormalities. PCD-causing mutations have been identified in 14 genes, but they collectively account for only ∼60% of all PCD. To identify mutations that cause PCD, we performed exome sequencing on six unrelated probands with ciliary outer dynein arm (ODA) defects. Mutations in CCDC114, an ortholog of the Chlamydomonas reinhardtii motility gene DCC2, were identified in a family with two affected siblings. Sanger sequencing of 67 additional individuals with PCD with ODA defects from 58 families revealed CCDC114 mutations in 4 individuals in 3 families. All 6 individuals with CCDC114 mutations had characteristic oto-sino-pulmonary disease, but none had situs abnormalities. In the remaining 5 individuals with PCD who underwent exome sequencing, we identified mutations in two genes (DNAI2, DNAH5) known to cause PCD, including an Ashkenazi Jewish founder mutation in DNAI2. These results revealed that mutations in CCDC114 are a cause of ciliary dysmotility and PCD and further demonstrate the utility of exome sequencing to identify genetic causes in heterogeneous recessive disorders.     

Endotoxemia is associated with an adverse metabolic profile

Our aim was to analyze whether endotoxemia, i.e. translocation of LPS to circulation, is reflected in the serum metabolic profile in a general population and in participants with cardiometabolic disorders. We investigated three Finnish cohorts separately and in a meta-analysis (n = 7178), namely population-based FINRISK97, FinnTwin16 consisting of young adult twins, and Parogene, a random cohort of cardiac patients. Endotoxemia was determined as serum LPS activity and metabolome by an NMR platform. Potential effects of body mass index (BMI), smoking, metabolic syndrome (MetS), and coronary heart disease (CHD) status were considered. Endotoxemia was directly associated with concentrations of VLDL, IDL, LDL, and small HDL lipoproteins, VLDL particle diameter, total fatty acids (FA), glycoprotein acetyls (GlycA), aromatic and branched-chain amino acids, and Glc, and inversely associated with concentration of large HDL, diameters of LDL and HDL, as well as unsaturation degree of FAs. Some of these disadvantageous associations were significantly stronger in smokers and subjects with high BMI, but did not differ between participants with different CHD status. In participants with MetS, however, the associations of endotoxemia with FA parameters and GlycA were particularly strong. The metabolic profile in endotoxemia appears highly adverse, involving several inflammatory characters and risk factors for cardiometabolic disorders.     

Taxonomic relationships among Arachis sect. Arachis species as revealed by AFLP markers.

Cultivated peanut, Arachis hypogaea L., is a tetraploid (2n = 4x = 40) species thought to be of allopolyploid origin. Its closest relatives are the diploid (2n = 2x = 20) annual and perennial species included with it in Arachis sect. Arachis. Species in section Arachis represent an important source of novel alleles for improvement of cultivated peanut. A better understanding of the level of speciation and taxonomic relationships between taxa within section Arachis is a prerequisite to the effective use of this secondary gene pool in peanut breeding programs. The AFLP technique was used to determine intra- and interspecific relationships among and within 108 accessions of 26 species of this section. A total of 1328 fragments were generated with 8 primer combinations. From those, 239 bands ranging in size from 65 to 760 bp were scored as binary data. Genetic distances among accessions ranged from 0 to 0.50. Average distances among diploid species (0.30) were much higher than that detected between tetraploid species (0.05). Cluster analysis using different methods and principal component analysis were performed. The resulting grouping of accessions and species supports previous taxonomic classifications and genome designations. Based on genetic distances and cluster analysis, A-genome accessions KG 30029 (Arachis helodes) and KSSc 36009 (Arachis simpsonii) and B-genome accession KGBSPSc 30076 (A. ipaensis) were the most closely related to both Arachis hypogaea and Arachis monticola. This finding suggests their involvement in the evolution of the tetraploid peanut species.     

Ecological photodynamic therapy: New trend to disrupt the intricate networks within tumor ecosystem

As with natural ecosystems, species within the tumor microenvironment are connected by pairwise interactions (e.g. mutualism, predation) leading to a strong interdependence of different populations on each other. In this review we have identified the ecological roles played by each non-neoplastic population (macrophages, endothelial cells, fibroblasts) and other abiotic components (oxygen, extracellular matrix) directly involved with neoplastic development. A way to alter an ecosystem is to affect other species within the environment that are supporting the growth and survival of the species of interest, here the tumor cells; thus, some features of ecological systems could be exploited for cancer therapy. We propose a well-known antitumor therapy called photodynamic therapy (PDT) as a novel modulator of ecological interactions. We refer to this as “ecological photodynamic therapy.” The main goal of this new strategy is the improvement of therapeutic efficiency through the disruption of ecological networks with the aim of destroying the tumor ecosystem. It is therefore necessary to identify those interactions from which tumor cells get benefit and those by which it is impaired, and then design multitargeted combined photodynamic regimes in order to orchestrate non-neoplastic populations against their neoplastic counterpart. Thus, conceiving the tumor as an ecological system opens avenues for novel approaches on treatment strategies.     

Review of uses of network and graph theory concepts within proteomics

The size and nature of data collected on gene and protein interactions has led to a rapid growth of interest in graph theory and modern techniques for describing, characterizing and comparing networks. Simultaneously, this is a field of growth within mathematics and theoretical physics, where the global properties, and emergent behavior of networks, as a function of the local properties has long been studied. In this review, a number of approaches for exploiting modern network theory to help describe and analyze different data sets and problems associated with proteomic data are considered. This review aims to help biologists find their way towards useful ideas and references, yet may also help scientists from a mathematics and physics background to understand where they may apply their expertise.     

The effect of land use change on the vascular plant species turnover in boreal lakes

The effects of different land use types on nutrient and chemical run-off have been widely researched, but the total effect of change in land use on the aquatic species community is not well-known. For this study we researched the effect of land use change on the vascular plant communities in lakes. The study was conducted with aerial photographs and GIS-techniques in the countryside of central Finland. We compared the change in vascular plant species community structure with the change in land use around 25 lakes, between the years 1933–1934 and 1996. The change in land use was analyzed in two different riparian zones. We found several indications of links between changes in land use and aquatic vascular plant species. According to our data a decrease in agricultural land use, in particular fields and meadows, appears to influence species turnover and increases the number of new vascular plant species. When the changes in fields and meadows were added together and compared to the change in species turnover, the correlation was even more evident. Changes in land use types in a zone 0–100 m from the lake shoreline correlated with a change in vascular plant species more often than changes in a larger, adjacent zone, 100–400 m from the shore. This indicates that changes in the few hundred meters nearest the lake have the most elemental consequences for aquatic vascular plant species, giving an indication of the breadth of the protection zones needed between freshwaters and agricultural lands.     

Neuronal localization of the TNFalpha converting enzyme (TACE) in brain tissue and its correlation to amyloid plaques

The tumor necrosis factor (TNF)-alpha converting enzyme (TACE) can cleave the cell-surface ectodomain of the amyloid-beta precursor protein (APP), thus decreasing the generation of amyloid-beta (Abeta) by cultured non-neuronal cells. While the amyloidogenic processing of APP in neurons is linked to the pathogenesis of Alzheimer's disease (AD), the expression of TACE in neurons has not yet been examined. Thus, we assessed TACE expression in a series of neuronal and non-neuronal cell types by Western blots. We found that TACE was present in neurons and was only faintly detectable in lysates of astrocytes, oligodendrocytes, and microglial cells. Immunohistochemical analysis was used to determine the cellular localization of TACE in the human brain, and its expression was detected in distinct neuronal populations, including pyramidal neurons of the cerebral cortex and granular cell layer neurons in the hippocampus. Very low levels of TACE were seen in the cerebellum, with Purkinje cells at the granular-molecular boundary staining faintly. Because TACE was localized predominantly in areas of the brain that are affected by amyloid plaques in AD, we examined its expression in a series of AD brains. We found that AD and control brains showed similar levels of TACE staining, as well as similar patterns of TACE expression. By double labeling for Abeta plaques and TACE, we found that TACE-positive neurons often colocalized with amyloid plaques in AD brains. These observations support a neuronal role for TACE and suggest a mechanism for its involvement in AD pathogenesis as an antagonist of Abeta formation.     

The tumor necrosis factor-alpha converting enzyme (TACE): a unique metalloproteinase with highly defined substrate selectivity

TNF alpha converting enzyme (TACE) processes precursor TNF alpha between Ala76 and Val77, yielding a correctly processed bioactive 17 kDa protein. Genetic evidence indicates that TACE may also be involved in the shedding of other ectodomains. Here we show that native and recombinant forms of TACE efficiently processed a synthetic substrate corresponding to the TNF alpha cleavage site only. For all other substrates, conversion occurred only at high enzyme concentrations and prolonged reaction times. Often, cleavage under those conditions was accompanied by nonspecific reactions. We also compared TNF alpha cleavage by TACE to cleavage by those members of the matrix metalloproteinase (MMP) family previously implied in TNF alpha release. The specificity constants for TNF alpha cleavage by the MMPs were approximately 100-1000-fold slower relative to TACE. MMP 7 also processed precursor TNF alpha at the correct cleavage site but did so with a 30-fold lower specificity constant relative to TACE. In contrast, MMP 1 processed precursor TNF alpha between Ala74 and Gln75, in addition to between Ala76 and Val77, while MMP 9 cleaved this natural substrate solely between Ala74 and Gln75. Additionally, the MMP substrate Dnp-PChaGC(Me)HK(NMA)-NH(2) was not cleaved at all by TACE, while collagenase (MMP 1), gelatinase (MMP 9), stromelysin 1 (MMP 3), and matrilysin (MMP 7) all processed this substrate efficiently. All of these results indicate that TACE is unique in terms of its specificity requirements for substrate cleavage.