R-Spondin2 is a secreted activator of Wnt/beta-catenin signaling and is required for Xenopus myogenesis

We have carried out a small pool expression screen for modulators of the Wnt/beta-catenin pathway and identified Xenopus R-spondin2 (Rspo2) as a secreted activator of this cascade. Rspo2 is coexpressed with and positively regulated by Wnt signals and synergizes with Wnts to activate beta-catenin. Analyses of functional interaction with components of the Wnt/beta-catenin pathway suggest that Rspo2 functions extracellularly at the level of receptor ligand interaction. In addition to activating the Wnt/beta-catenin pathway, Rspo2 overexpression blocks Activin, Nodal, and BMP4 signaling in Xenopus, raising the possibility that it may negatively regulate the TGF-beta pathway. Antisense Morpholino experiments in Xenopus embryos and RNAi experiments in HeLa cells reveal that Rspo2 is required for Wnt/beta-catenin signaling. In Xenopus embryos depleted of Rspo2, the muscle markers myoD and myf5 fail to be activated and later muscle development is impaired. Thus, Rspo2 functions in a positive feedback loop to stimulate the Wnt/beta-catenin cascade.     

Demography and natural selection have shaped genetic variation in Drosophila melanogaster: a multi-locus approach

Demography and selection have been recognized for their important roles in shaping patterns of nucleotide variability. To investigate the relative effects of these forces in the genome of Drosophila melanogaster, we used a multi-locus scan (105 fragments) of X-linked DNA sequence variation in a putatively ancestral African and a derived European population. Surprisingly, we found evidence for a recent size expansion in the African population, i.e., a significant excess of singletons at a chromosome-wide level. In the European population, such an excess was not detected. In contrast to the African population, we found evidence for positive natural selection in the European sample: (i) a large number of loci with low levels of variation and (ii) a significant excess of derived variants at the low-variation loci that are fixed in the European sample but rare in the African population. These results are consistent with the hypothesis that the European population has experienced frequent selective sweeps in the recent past during its adaptation to new habitats. Our study shows the advantages of a genomic approach (over a locus-specific analysis) in disentangling demographic and selective forces.     

Dickkopf1 and the Spemann-Mangold head organizer

Work in amphibians indicates that inhibition of Wnt and BMP signals is essential for head development and that head induction by the Spemann-Mangold organizer may be mediated by secreted Wnt antagonists. Wnts are potent posteriorizing factors and antagonize the Spemann-Mangold organizer. Dickkopf1 (dkk1) encodes a secreted effector expressed in head organizing centers of Xenopus, mouse and zebrafish. It acts as a Wnt inhibitor and is able together with BMP inhibitors to induce the formation of ectopic embryonic heads in Xenopus. It anteriorizes both mesendoderm and neuroectoderm, promoting prechordal plate and forebrain fates. Injection of inhibitory antibodies leads to microcephaly and cyclopia. Dkk1 thus is an essential mediator of the vertebrate head organizer.     

Phenotypic effects in Xenopus and zebrafish suggest that one-eyed pinhead functions as antagonist of BMP signalling

Zebrafish one-eyed pinhead (oep) is essential for embryonic axis and dorsal midline formation by promoting Nodal signalling and is thought to act as a permissive factor. Here we describe that oep elicits profound phenotypic effects when overexpressed in Xenopus and zebrafish. In Xenopus, wild-type oep inhibits mesoderm induction, disrupts axis formation and neuralizes animal caps. A secreted Oep dorsoanteriorizes and neuralizes Xenopus embryos indicative of BMP inhibition. In zebrafish, misexpression of smad1 in oep mutant embryos also reveals an interaction of oep with BMP signalling. Furthermore, the phenotypic effect of nodal overexpression can be rescued by coexpression of oep both in Xenopus and zebrafish. Taken together, our results support an interaction between oep and nodal but they suggest also (1) that the role of oep in Nodal signalling may include negative as well as positive regulation, (2) that oep is able to function in an active fashion and (3) that oep exerts a regulatory effect on the BMP signalling pathway.     

The genomic structure, chromosome location, and analysis of the human DKK1 head inducer gene as a candidate for holoprosencephaly

Holoprosencephaly (HPE) is the most common developmental defect of the brain and face in humans. Here we report the analysis of the human ortholog of dkk-1 as a candidate gene for HPE. We determined the genomic structure of the human gene DKK1 and mapped it to chromosome 10q11.2. Functional analysis of four missense mutations identified in HPE patients revealed preserved activity in head induction assays in frogs suggesting a limited role for this gene in HPE pathogenesis.     

Mutual antagonism between dickkopf1 and dickkopf2 regulates Wnt/beta-catenin signalling

Wnts are secreted glycoproteins implicated in diverse processes during embryonic patterning in metazoans. They signal through seven-transmembrane receptors of the Frizzled (Fz) family [1] to stabilise beta-catenin [2]. Wnts are antagonised by several extracellular inhibitors including the product of the dickkopf1 (dkk1) gene, which was identified in Xenopus embryos and is a member of a multigene family. The dkk1 gene acts upstream of the Wnt pathway component dishevelled but its mechanism of action is unknown [3]. Although the function of Dkk1 as a Wnt inhibitor in vertebrates is well established [3-6], the effect of other Dkks on the Wnt/beta-catenin pathway is unclear. Here, we report that a related family member, Dkk2, activates rather than inhibits the Wnt/beta-catenin signalling pathway in Xenopus embryos. Dkk2 strongly synergised with Wnt receptors of the Fz family to induce Wnt signalling responses. The study identifies Dkk2 as a secreted molecule that is able to activate Wnt/beta-catenin signalling. The results suggest that a coordinated interplay between inhibiting dkk1 and activating dkk2 can modulate Fz signalling.     

<Emphasis Type="Italic">TRAF1/C5</Emphasis>polymorphism is not associated with increased mortality in rheumatoid arthritis: two large longitudinal studies

Introduction  

Recently an association between a genetic variation in TRAF1/C5 and mortality from sepsis or cancer was found in rheumatoid arthritis (RA). The most prevalent cause of death, cardiovascular disease, may have been missed in that study, since patients were enrolled at an advanced disease stage. Therefore, we used an inception cohort of RA patients to investigate the association between TRAF1/C5 and cardiovascular mortality, and replicate the findings on all-cause mortality. As TRAF1/C5 associated mortality may not be restricted to RA, we also studied a large cohort of non-RA patients.     

Mesenchymal stem cells in autoimmune diseases: hype or hope?

Intervention with mesenchymal stem cells (MSCs) represents a promising therapeutic tool in treatment-refractory autoimmune diseases. A new report by Schurgers and colleagues in a previous issue of Arthritis Research & Therapy sheds novel mechanistic insight into the pathways employed by MSCs to suppress T-cell proliferation in vitro, but, at the same time, indicates that MSCs do not influence T-cell reactivity and the disease course in an in vivo arthritis model. Such discrepancies between the in vitro and in vivo effects of potent cellular immune modulators should spark further research and should be interpreted as a sign of caution for the in vitro design of MSC-derived interventions in the setting of human autoimmune diseases.     

Pentacycloundecane lactam vs lactone norstatine type protease HIV inhibitors: binding energy calculations and DFT study

Background

Novel pentacycloundecane (PCU)-lactone-CO-EAIS peptide inhibitors were designed, synthesized, and evaluated against wild-type C-South African (C-SA) HIV-1 protease. Three compounds are reported herein, two of which displayed IC₅₀ values of less than 1.00 μM. A comparative MM-PB(GB)SA binding free energy of solvation values of PCU-lactam and lactone models and their enantiomers as well as the PCU-lactam-NH-EAIS and lactone-CO-EAIS peptide inhibitors and their corresponding diastereomers complexed with South African HIV protease (C-SA) was performed. This will enable us to rationalize the considerable difference between inhibitory concentration (IC₅₀) of PCU-lactam-NH-EAIS and PCU-lactone-CO-EAIS peptides.

Results

The PCU-lactam model exhibited more negative calculated binding free energies of solvation than the PCU-lactone model. The same trend was observed for the PCU-peptide inhibitors, which correspond to the experimental activities for the PCU-lactam-NH-EAIS peptide (IC₅₀ = 0.076 μM) and the PCU-lactone-CO-EAIS peptide inhibitors (IC₅₀ = 0.850 μM). Furthermore, a density functional theory (DFT) study on the natural atomic charges of the nitrogen and oxygen atoms of the three PCU-lactam, PCU-lactim and PCU-lactone models were performed using natural bond orbital (NBO) analysis. Electrostatic potential maps were also used to visualize the electron density around electron-rich regions. The asymmetry parameter (η) and quadrupole coupling constant (χ) values of the nitrogen and oxygen nuclei of the model compounds were calculated at the same level of theory. Electronic molecular properties including polarizability and electric dipole moments were also calculated and compared. The Gibbs theoretical free solvation energies of solvation (∆G_solv) were also considered.

Conclusions

A general trend is observed that the lactam species appears to have a larger negative charge distribution around the heteroatoms, larger quadrupole constant, dipole moment and better solvation energy, in comparison to the PCU-lactone model. It can be argued that these characteristics will ensure better eletronic interaction between the lactam and the receptor, corresponding to the observed HIV protease activities in terms of experimental IC₅₀ data.

Electronic supplementary material

The online version of this article (doi:10.1186/s12929-015-0115-5) contains supplementary material, which is available to authorized users.