Functional flower traits and their diversity drive pollinator visitation

Recent studies have shown that the diversity of flowering plants can enhance pollinator richness and visitation frequency and thereby increase the resilience of pollination. It is assumed that flower traits explain these effects, but it is still unclear which flower traits are responsible, and knowing that, if pollinator richness and visitation frequency are more driven by mass‐ratio effects (mean trait values) or by trait diversity. Here, we analyse a three‐year data set of pollinator observations collected in a European grassland plant diversity experiment (The Jena experiment). The data entail comprehensive flower trait measurements, including reward traits (nectar and pollen amount), morphological traits (height, symmetry, area, colour spectra) and chemical traits (nectar‐amino acid and nectar‐sugar concentration). We test if pollinator species richness and visitation frequency of flower communities depend on overall functional diversity combining all flower traits within a community, single trait diversities (within trait variation) and community‐weighted means of the single traits, using Bayesian inference. Overall functional diversity did not affect pollinator species richness, but reduced visitation frequency. When looking at individual flower traits separately, we found that single trait diversity of flower reflectance and flower morphology were important predictors of pollinator visitation frequency. Moreover, independent of total flower abundance, community‐weighted means of flower height, area, reflectance, nectar‐sugar concentration and nectar‐amino acid concentration strongly affected both pollinator species richness and visitation frequency. Our results, challenge the idea that functional diversity always positively affects ecosystem functions. Nonetheless, we demonstrate that both single trait diversity and mass‐ratio effects of flower traits play an important role for diverse and frequent flower visits, which underlines the functionality of flower traits for pollination services.     

α4β2 nicotinic acetylcholine receptors in the early postnatal mouse superior cervical ganglion

Heteropentameric nicotinic acetylcholine receptors (nAChR) mediate fast synaptic transmission in ganglia of the autonomic nervous system. It is undisputed that α3 and β4 are the predominant subunits in the superior cervical ganglion (SCG); however, reports on the presence of receptors that contain α4 have been controversial. Here, we have searched for the presence of α4-containing nAChRs in the postnatal rat and mouse SCG. We now show by immunoprecipitation combined with radioligand binding that α4-containing receptors constitute about 20% of hetero-oligomeric nAChRs in postnatal Day 3 (P3) mice. However, already by P9, the level of α4 approaches zero. In contrast, the number of α4-containing receptors is close to zero in the rat SCG at all times investigated. Deletion of the β2 subunit by using α5β2-double knockout (KO) mice removes all α4-containing receptors, suggesting that in the postnatal mouse SCG, α4 co-assembles only with β2 but not with β4. α4β2 receptors are, on the other hand, up-regulated in the SCG of P3 α5β4-double KO mice, where they make up about 50% of receptors that bind [(3) H]-epibatidine. Nonetheless, receptors on the surface of SCG neurons from α5β4-double KO mice maintained for one to two days in culture comprise <10% of α4β2 and >90% of α3β2, as determined by patch clamp recordings with α4β2- and α3β2-specific ligands. We propose that in the P3 SCG of wild type mice, α3β4 (±α5) represent about 62% of receptors, whereas 17% are α3β2β4, and 21% are α4β2 (±α5) receptors.     

Insights into the activation of brain serine racemase by the multi-PDZ domain glutamate receptor interacting protein, divalent cations and ATP

Brain serine racemase contains pyridoxal phosphate as a prosthetic group and is known to become activated by divalent cations such as Ca(2+) and Mg(2+), as well as by ATP and ADP. In vivo, brain serine racemase is also activated by a multi-PSD-95/discs large/ZO-1 (PDZ) domain glutamate receptor interacting protein (GRIP) that is usually coupled to the GluR2/3 subunits of the alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid Ca(2+) channel. In the present study, we analysed the mechanisms by which serine racemase becomes activated by GRIP, divalent cations and ATP. We show that binding of PDZ6 of GRIP to serine racemase does not result in increased d-serine production. However, full-length GRIP does augment significantly enzymatic activity. We expressed various GRIP shorter constructs to map down the regions within GRIP that are necessary for serine racemase activation. We observed that, whereas recombinant proteins containing PDZ4-PDZ5-PDZ6 are unable to activate serine racemase, other constructs containing PDZ4-PDZ5-PDZ6-GAP2-PDZ7 significantly augment its activity. Hence, activation of serine racemase by GRIP is not a direct consequence of the translocation towards the calcium channel but rather a likely conformational change induced by GRIP on serine racemase. On the other hand, the observed activation of serine racemase by divalent cations has been assumed to be a side-effect associated with ATP binding, which is known to form a complex with Mg(2+) ions. Because no mammalian serine racemase has yet been crystallized, we used molecular modelling based on yeast and bacterial homologs to demonstrate that the binding sites for Ca(2+), ATP and the PDZ6 domain of GRIP are spatially separated and modulate the enzyme through distinct mechanisms.     

The role of osteopontin (OPN/SPP1) haplotypes in the susceptibility to Crohn's disease

Background

Osteopontin represents a multifunctional molecule playing a pivotal role in chronic inflammatory and autoimmune diseases. Its expression is increased in inflammatory bowel disease (IBD). The aim of our study was to analyze the association of osteopontin (OPN/SPP1) gene variants in a large cohort of IBD patients.

Methodology/Principal Findings

Genomic DNA from 2819 Caucasian individuals (n = 841 patients with Crohn''s disease (CD), n = 473 patients with ulcerative colitis (UC), and n = 1505 healthy unrelated controls) was analyzed for nine OPN SNPs (rs2728127, rs2853744, rs11730582, rs11739060, rs28357094, rs4754 = p.Asp80Asp, rs1126616 = p.Ala236Ala, rs1126772 and rs9138). Considering the important role of osteopontin in Th17-mediated diseases, we performed analysis for epistasis with IBD-associated IL23R variants and analyzed serum levels of the Th17 cytokine IL-22. For four OPN SNPs (rs4754, rs1126616, rs1126772 and rs9138), we observed significantly different distributions between male and female CD patients. rs4754 was protective in male CD patients (p = 0.0004, OR = 0.69). None of the other investigated OPN SNPs was associated with CD or UC susceptibility. However, several OPN haplotypes showed significant associations with CD susceptibility. The strongest association was found for a haplotype consisting of the 8 OPN SNPs rs2728127-rs2853744-rs11730582-rs11439060-rs28357094-rs112661-rs1126772-rs9138 (omnibus p-value = 2.07×10⁻⁸). Overall, the mean IL-22 secretion in the combined group of OPN minor allele carriers with CD was significantly lower than that of CD patients with OPN wildtype alleles (p = 3.66×10⁻⁵). There was evidence for weak epistasis between the OPN SNP rs28357094 with the IL23R SNP rs10489629 (p = 4.18×10⁻²) and between OPN SNP rs1126616 and IL23R SNP rs2201841 (p = 4.18×10⁻²) but none of these associations remained significant after Bonferroni correction.

Conclusions/Significance

Our study identified OPN haplotypes as modifiers of CD susceptibility, while the combined effects of certain OPN variants may modulate IL-22 secretion.     

Mutation Analysis of BRCA1, BRCA2, PALB2 and BRD7 in a Hospital-Based Series of German Patients with Triple-Negative Breast Cancer

Triple-negative breast cancer (TNBC) is an aggressive form of breast carcinoma with a poor prognosis. Recent evidence suggests that some patients with TNBC harbour germ-line mutations in DNA repair genes which may render their tumours susceptible to novel therapies such as treatment with PARP inhibitors. In the present study, we have investigated a hospital-based series of 40 German patients with TNBC for the presence of germ-line mutations in BRCA1, BRCA2, PALB2, and BRD7 genes. Microfluidic array PCR and next-generation sequencing was used for BRCA1 and BRCA2 analysis while conventional high-resolution melting and Sanger sequencing was applied to study the coding regions of PALB2 and BRD7, respectively. Truncating mutations in BRCA1 were found in six patients, and truncating mutations in BRCA2 and PALB2 were detected in one patient each, whereas no truncating mutation was identified in BRD7. One patient was a double heterozygote for the PALB2 mutation, c.758insT, and a BRCA1 mutation, c.927delA. Our results confirm in a hospital-based setting that a substantial proportion of German TNBC patients (17.5%) harbour germ-line mutations in genes involved in homology-directed DNA repair, with a preponderance of BRCA1 mutations. Triple-negative breast cancer should be considered as an additional criterion for future genetic counselling and diagnostic sequencing.     

Phylogenetic Quantification of Intra-tumour Heterogeneity

Intra-tumour genetic heterogeneity is the result of ongoing evolutionary change within each cancer. The expansion of genetically distinct sub-clonal populations may explain the emergence of drug resistance, and if so, would have prognostic and predictive utility. However, methods for objectively quantifying tumour heterogeneity have been missing and are particularly difficult to establish in cancers where predominant copy number variation prevents accurate phylogenetic reconstruction owing to horizontal dependencies caused by long and cascading genomic rearrangements. To address these challenges, we present MEDICC, a method for phylogenetic reconstruction and heterogeneity quantification based on a Minimum Event Distance for Intra-tumour Copy-number Comparisons. Using a transducer-based pairwise comparison function, we determine optimal phasing of major and minor alleles, as well as evolutionary distances between samples, and are able to reconstruct ancestral genomes. Rigorous simulations and an extensive clinical study show the power of our method, which outperforms state-of-the-art competitors in reconstruction accuracy, and additionally allows unbiased numerical quantification of tumour heterogeneity. Accurate quantification and evolutionary inference are essential to understand the functional consequences of tumour heterogeneity. The MEDICC algorithms are independent of the experimental techniques used and are applicable to both next-generation sequencing and array CGH data.