PhySIC_IST: cleaning source trees to infer more informative supertrees

Background  

Supertree methods combine phylogenies with overlapping sets of taxa into a larger one. Topological conflicts frequently arise among source trees for methodological or biological reasons, such as long branch attraction, lateral gene transfers, gene duplication/loss or deep gene coalescence. When topological conflicts occur among source trees, liberal methods infer supertrees containing the most frequent alternative, while veto methods infer supertrees not contradicting any source tree, i.e. discard all conflicting resolutions. When the source trees host a significant number of topological conflicts or have a small taxon overlap, supertree methods of both kinds can propose poorly resolved, hence uninformative, supertrees.     

From drought sensing to developmental control: evolution of cyclic AMP signaling in social amoebas

Amoebas and other protists commonly encyst when faced with environmental stress. Although little is known of the signaling pathways that mediate encystation, the analogous process of spore formation in dictyostelid social amoebas is better understood. In Dictyostelium discoideum, secreted cyclic AMP (cAMP) mediates the aggregation of starving amoebas and induces the differentiation of prespore cells. Intracellular cAMP acting on cAMP-dependent protein kinase (PKA) triggers the maturation of spores and prevents their germination under the prevalent conditions of high osmolality in the spore head. The osmolyte-activated adenylate cyclase, ACG, produces cAMP for prespore differentiation and inhibition of spore germination. To retrace the origin of ACG function, we investigated ACG gene conservation and function in species that span the dictyostelid phylogeny. ACG genes, osmolyte-activated ACG activity, and osmoregulation of spore germination were detected in species that represent the 4 major groups of Dictyostelia. Unlike the derived species D. discoideum, many basal Dictyostelia have retained the ancestral mechanism of encystation from solitary amoebas. In these species and in solitary amoebas, encystation is independently triggered by starvation or by high osmolality. Osmolyte-induced encystation was accompanied by an increase in cAMP and prevented by inhibition of PKA, indicating that ACG and PKA activation mediate this response. We propose that high osmolality signals drought in soil amoebas and that developmental cAMP signaling in the Dictyostelia has evolved from this stress response.     

Polyanxanthone A, B and C, three xanthones from the wood trunk of Garcinia polyantha Oliv

Three xanthones, polyanxanthone A (1), B (2) and C (3) have been isolated from the methanol extract of the wood trunk of Garcinia polyantha, along with five known xanthones: 1,3,5-trihydroxyxanthone (4); 1,5-dihydroxyxanthone (5); 1,3,6,7-tetrahydroxyxanthone (6); 1,6-dihydroxy-5-methoxyxanthone (7) and 1,3,5,6-tetrahydroxyxanthone (8). Their structures were determined by means of 1D- and 2D-NMR techniques. Some of the above compounds were screened for their anticholinesterase activity on acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) enzymes.     

Vitamin B12-Impaired Metabolism Produces Apoptosis and Parkinson Phenotype in Rats Expressing the Transcobalamin-Oleosin Chimera in Substantia Nigra

Background

Vitamin B12 is indispensable for proper brain functioning and cytosolic synthesis of S-adenosylmethionine. Whether its deficiency produces effects on viability and apoptosis of neurons remains unknown. There is a particular interest in investigating these effects in Parkinson disease where Levodopa treatment is known to increase the consumption of S-adenosylmethionine. To cause deprivation of vitamin B12, we have recently developed a cell model that produces decreased synthesis of S-adenosylmethionine by anchoring transcobalamin (TCII) to the reticulum through its fusion with Oleosin (OLEO).

Methodology

Gene constructs including transcobalamin-oleosin (TCII-OLEO) and control constructs, green fluorescent protein-transcobalamin-oleosin (GFP-TCII-OLEO), oleosin-transcobalamin (OLEO-TCII), TCII and OLEO were used for expression in N1E-115 cells (mouse neuroblastoma) and in substantia nigra of adult rats, using a targeted transfection with a Neurotensin polyplex system. We studied the viability and the apoptosis in the transfected cells and targeted tissue. The turning behavior was evaluated in the rats transfected with the different plasmids.

Principal Findings

The transfection of N1E-115 cells by the TCII-OLEO-expressing plasmid significantly affected cell viability and increased immunoreactivity of cleaved Caspase-3. No change in propidium iodide uptake (used as a necrosis marker) was observed. The transfected rats lost neurons immunoreactive to tyrosine hydroxylase. The expression of TCII-OLEO was observed in cells immunoreactive to tyrosine hydroxylase of the substantia nigra, with a superimposed expression of cleaved Caspase-3. These cellular and tissular effects were not observed with the control plasmids. Rats transfected with TCII-OLEO expressing plasmid presented with a significantly higher number of turns, compared with those transfected with the other plasmids.

Conclusions/Significance

In conclusion, the TCII-OLEO transfection was responsible for apoptosis in N1E-115 cells and rat substantia nigra and for Parkinson-like phenotype. This suggests evaluating whether vitamin B12 deficit could aggravate the PD in patients under Levodopa therapy by impairing S-adenosylmethionine synthesis in substantia nigra.     

NEK8 regulates DNA damage-induced RAD51 foci formation and replication fork protection

Proteins essential for homologous recombination play a pivotal role in the repair of DNA double strand breaks, DNA inter-strand crosslinks and replication fork stability. Defects in homologous recombination also play a critical role in the development of cancer and the sensitivity of these cancers to chemotherapy. RAD51, an essential factor for homologous recombination and replication fork protection, accumulates and forms immunocytochemically detectable nuclear foci at sites of DNA damage. To identify kinases that may regulate RAD51 localization to sites of DNA damage, we performed a human kinome siRNA library screen, using DNA damage-induced RAD51 foci formation as readout. We found that NEK8, a NIMA family kinase member, is required for efficient DNA damage-induced RAD51 foci formation. Interestingly, knockout of Nek8 in murine embryonic fibroblasts led to cellular sensitivity to the replication inhibitor, hydroxyurea, and inhibition of the ATR kinase. Furthermore, NEK8 was required for proper replication fork protection following replication stall with hydroxyurea. Loading of RAD51 to chromatin was decreased in NEK8-depleted cells and Nek8-knockout cells. Single-molecule DNA fiber analyses revealed that nascent DNA tracts were degraded in the absence of NEK8 following treatment with hydroxyurea. Consistent with this, Nek8-knockout cells showed increased chromosome breaks following treatment with hydroxyurea. Thus, NEK8 plays a critical role in replication fork stability through its regulation of the DNA repair and replication fork protection protein RAD51.     

Evaluating the influence of quality control decisions and software algorithms on SNP calling for the affymetrix 6.0 SNP array platform

Objective: Our goal was to evaluate the influence of quality control (QC) decisions using two genotype calling algorithms, CRLMM and Birdseed, designed for the Affymetrix SNP Array 6.0. Methods: Various QC options were tried using the two algorithms and comparisons were made on subject and call rate and on association results using two data sets. Results: For Birdseed, we recommend using the contrast QC instead of QC call rate for sample QC. For CRLMM, we recommend using the signal-to-noise rate ≥4 for sample QC and a posterior probability of 90% for genotype accuracy. For both algorithms, we recommend calling the genotype separately for each plate, and dropping SNPs with a lower call rate (<95%) before evaluating samples with lower call rates. To investigate whether the genotype calls from the two algorithms impacted the genome-wide association results, we performed association analysis using data from the GENOA cohort; we observed that the number of significant SNPs were similar using either CRLMM or Birdseed. Conclusions: Using our suggested workflow both algorithms performed similarly; however, fewer samples were removed and CRLMM took half the time to run our 854 study samples (4.2 h) compared to Birdseed (8.4 h).     

Suberoylanilide hydroxamic acid, a potent histone deacetylase inhibitor; its X-ray crystal structure and solid state and solution studies of its Zn(II), Ni(II), Cu(II) and Fe(III) complexes

Reaction of the potent hydroxamate-based histone deacetylase (HDAC) inhibitor, suberoylanilide hydroxamic acid (SAHA), with hydrated metal salts of Fe(III), Cu(II), Ni(II) and Zn(II) yielded a tris-hydroxamato complex in the case of Fe(III) and bis-hydroxamato complexes in the case of Cu(II), Ni(II) and Zn(II) both in the solid state and in solution. Reaction of the secondary hydroxamic acid, N-Me-SAHA, also yielded a tris-hydroxamato complex in the case of Fe(III) and bis-hydroxamato complexes in the case of Cu(II), Ni(II) and Zn(II) in solution. These metal complexes have the hydroxamato moiety coordinated in an O,O’-bidentate fashion. Stability constants of the metal complexes formed with SAHA and N-Me-SAHA in a DMSO/H₂O 70/30%(v/v) mixture are described. A novel crystal structure of SAHA together with a novel synthesis for N-Me-SAHA are also reported.     

Mechanism of GABAB receptor-induced BDNF secretion and promotion of GABAA receptor membrane expression

Recent studies have shown that GABA(B) receptors play more than a classical inhibitory role and can function as an important synaptic maturation signal early in life. In a previous study, we reported that GABA(B) receptor activation triggers secretion of brain-derived neurotrophic factor (BDNF) and promotes the functional maturation of GABAergic synapses in the developing rat hippocampus. To identify the signalling pathway linking GABA(B) receptor activation to BDNF secretion in these cells, we have now used the phosphorylated form of the cAMP response element-binding protein as a biological sensor for endogenous BDNF release. In the present study, we show that GABA(B) receptor-induced secretion of BDNF relies on the activation of phospholipase C, followed by the formation of diacylglycerol, activation of protein kinase C, and the opening of L-type voltage-dependent Ca(2+) channels. We further show that once released by GABA(B) receptor activation, BDNF increases the membrane expression of β(2/3) -containing GABA(A) receptors in neuronal cultures. These results reveal a novel function of GABA(B) receptors in regulating the expression of GABA(A) receptor through BDNF-tropomyosin-related kinase B receptor dependent signalling pathway.     

Hyperactivation of the G12-mediated signaling pathway in Caenorhabditis elegans induces a developmental growth arrest via protein kinase C

The G(12) type of heterotrimeric G-proteins play an important role in development and behave as potent oncogenes in cultured cells. However, little is known about the molecular nature of the components that act in the G(12)-signaling pathway in an organism. We characterized a C. elegans Galpha subunit gene, gpa-12, which is a homolog of mammalian G(12)/G(13)alpha, and found that animals defective in gpa-12 are viable. Expression of activated GPA-12 (G(12)QL) results in a developmental growth arrest caused by a feeding behavior defect that is due to a dramatic reduction in pharyngeal pumping. To elucidate the molecular nature of the signaling pathways in which G(12) participates, we screened for suppressors of the G(12)QL phenotype. We isolated 50 suppressors that contain mutations in tpa-1, which encodes two protein kinase C isoforms, TPA-1A and TPA-1B, most similar to PKCtheta/delta. TPA-1 mediates the action of the tumor promoter PMA. Expression of G(12)QL and treatment of wild-type animals with PMA induce an identical growth arrest caused by inhibition of larval feeding, which is dependent on TPA-1A and TPA-1B function. These results suggest that TPA-1 is a downstream target of both G(12) signaling and PMA in modulating feeding and growth in C. elegans. Taken together, our findings provide a potential molecular mechanism for the transforming capability of G(12) proteins.