Coexpression of Three Odorant-Binding Protein Genes in the Foreleg Gustatory Sensilla of Swallowtail Butterfly Visualized by Multicolor FISH Analysis

Lepidopteran insects are mostly monophagous or oligophagous. Female butterflies distinguish their host plants by detecting a combination of specific phytochemicals through the gustatory sensilla densely distributed on their foreleg tarsi, thereby ensuring oviposition on appropriate host plants. In this study, to gain insight into the molecular mechanism underlying host plant recognition by the gustatory sensilla, using Asian swallowtail, Papilio xuthus, we focused on a family of small soluble ligand-binding molecules, odorant-binding proteins (OBPs), and found that three OBP genes showed enriched expression in the foreleg tarsus. Multicolor fluorescence in situ hybridization analyses demonstrated the coexpression of these three OBP genes at the bases of the foreleg gustatory sensilla. Further analyses on other appendages revealed that PxutOBP3 was exclusively expressed in the tissues which could have direct contact with the leaf surface, suggesting that this OBP gene specifically plays an important role in phytochemicals perception.

Airway monocyte modulation relates to TNF dysregulation in neutrophilic asthma

BackgroundDysregulation of TNFα signalling is implicated in neutrophilic asthma. TNFα signalling involves membrane-bound and soluble ligand (TNFα) and receptors (TNFRs); however, little is known about how these factors are altered in asthma. We hypothesized that intercompartment-, immune cell- and/or asthma inflammatory phenotype-dependent regulation could relate to TNF factor dysregulation in neutrophilic asthma.MethodsMeasures were made in 45 adults with asthma (36 non-neutrophilic, 9 neutrophilic) and 8 non-asthma controls. Soluble TNFα, TNFR1 and TNFR2 were quantified in plasma and sputum supernatant by ELISA, and membrane-bound TNFα/TNFR1/TNFR2 measured on eosinophils, neutrophils, monocytes, and macrophages in blood and sputum by flow cytometry. Marker expression was compared between inflammatory phenotypes and compartments, and relationship of membrane-bound and soluble TNF markers and immune cell numbers tested by correlation.ResultsSoluble sputum TNFR1 and TNFR2 were increased in neutrophilic versus non-neutrophilic asthma (p=0.010 and p=0.029). Membrane-bound TNFα expression was upregulated on sputum versus blood monocytes, while TNFR1 and TNFR2 levels were reduced on airway versus blood monocytes and neutrophils. Soluble TNFR1 and TNFR2 in sputum significantly correlated with the number of airway monocytes (p=0.016, r=0.358 and p=0.029, r=0.327).ConclusionOur results imply that increased sputum soluble TNF receptor levels observed in neutrophilic asthma relate to the increased recruitment of monocytes and neutrophils into the airways and their subsequent receptor shedding. Monocytes also increase TNFα ligand expression in the airways. These results suggest an important contribution of airway monocytes to the altered inflammatory milieu in neutrophilic asthma.

Circulating calcification inhibitors are associated with arterial damage in pediatric patients with primary hypertension

Background Circulating calcification inhibitors: fetuin A (FA) and osteoprotegerin (OPG) together with soluble ligand of receptor activator of nuclear factor kappa-B (sRANKL) have been linked to vascular calcifications and arterial damage. This study aimed to evaluate relationships between FA, OPG, sRANKL, and arterial damage in children with primary hypertension (PH). Methods In this cross-sectional single-center study, calcification inhibitors (FA, OPG, sRANKL) levels were measured in blood samples of 60 children with PH (median age 15.8, IQR: [14.5–16.8] years) and 20 age-matched healthy volunteers. In each participant, peripheral and central blood pressure evaluation (BP) and ambulatory BP monitoring (ABPM) were performed. Arterial damage was measured using common carotid artery intima media thickness (cIMT), pulse wave velocity (PWV), augmentation index (AIx75HR), and local arterial stiffness (ECHO-tracking—ET) analysis. Results Children with PH had significantly higher peripheral and central BP, BP in ABPM, thicker cIMT, higher PWV, and AIx75HR. FA was significantly lower in patients with PH compared to healthy peers without differences in OPG, sRANKL, and OPG/sRANKL and OPG/FA ratios. In children with PH, FA level correlated negatively with cIMT Z-score and ET AIx; sRANKL level correlated negatively with ABPM systolic blood pressure (SBP), SBP load, diastolic BP load, and AIx75HR; OPG/sRANKL ratio correlated positively with SBP load, while OPG/FA ratio correlated positively with ET AIx. In multivariate analysis, FA was a significant determinant of cIMT (mm) and cIMT Z-score. Conclusions This study reveals that in children with primary hypertension, arterial damage is related to lower fetuin A concentrations. Graphical abstract

Receptor Oligomerization and Its Relevance for Signaling by Receptors of the Tumor Necrosis Factor Receptor Superfamily

With the exception of a few signaling incompetent decoy receptors, the receptors of the tumor necrosis factor receptor superfamily (TNFRSF) are signaling competent and engage in signaling pathways resulting in inflammation, proliferation, differentiation, and cell migration and also in cell death induction. TNFRSF receptors (TNFRs) become activated by ligands of the TNF superfamily (TNFSF). TNFSF ligands (TNFLs) occur as trimeric type II transmembrane proteins but often also as soluble ligand trimers released from the membrane-bound form by proteolysis. The signaling competent TNFRs are efficiently activated by the membrane-bound TNFLs. The latter recruit three TNFR molecules, but there is growing evidence that this is not sufficient to trigger all aspects of TNFR signaling; rather, the formed trimeric TNFL–TNFR complexes have to cluster secondarily in the cell-to-cell contact zone for full TNFR activation. With respect to their response to soluble ligand trimers, the signaling competent TNFRs can be subdivided into two groups. TNFRs of one group, designated as category I TNFRs, are robustly activated by soluble ligand trimers. The receptors of a second group (category II TNFRs), however, failed to become properly activated by soluble ligand trimers despite high affinity binding. The limited responsiveness of category II TNFRs to soluble TNFLs can be overcome by physical linkage of two or more soluble ligand trimers or, alternatively, by anchoring the soluble ligand molecules to the cell surface or extracellular matrix. This suggests that category II TNFRs have a limited ability to promote clustering of trimeric TNFL–TNFR complexes outside the context of cell–cell contacts. In this review, we will focus on three aspects on the relevance of receptor oligomerization for TNFR signaling: (i) the structural factors which promote clustering of free and liganded TNFRs, (ii) the signaling pathway specificity of the receptor oligomerization requirement, and (iii) the consequences for the design and development of TNFR agonists.

Comparative analysis of the levels of soluble forms of receptor and ligand of the immunity control point PD-1 / PD-L1 in the blood serum of patients with typical bone osteosarcoma and chondrosarcoma

Results of ELISA investigation of the pretreatment sPD-1 and sPD-L1 content in blood serum of 133 bone neoplasms patients aged 6-70 years and 57 practically healthy control persons aged 12-70 years are described. In 14 patients the neoplasms were of a benign character, in 16 - borderline giant-cell bone tumor was diagnosed, and in 103 - malignant bone lesions including 39 osteosarcomas and 42 chondrosarcomas were revealed. The sPD-1 receptor concentrations in blood serum did not differ between control healthy persons and primary bone tumor patients, while serum sPD-L1 level in bone tumor patients was statistically significantly increased (p<0.0000001). By means of ROC curve construction a cut-off sPD-L1 level of 16.5 pg/ml was found that imposed 75,9% sensitivity and 75,4% specificity in relation to healthy control. However, the frequency of sPD-L1 levels exceeding 16.5 pg/ml was approximately similar in benign, borderline and malignant bone tumor patients. Analysis of the pattern of sPD-1 and sPD-L1 circulation in the peripheral blood of patients with the most prevalent malignant bone tumors - osteosarcoma and chondrosarcoma - demonstrated that in both sarcoma types sPD-L1 level was significantly higher than in control, but in patients with chondrogenic tumors the soluble ligand sPD-L1 dominates in the circulation, while in those with osteogenic tumors - sPD-1 receptor prevails. In particular, sPD-1 level is statistically significantly higher in patients with typical osteosarcoma than in those with typical chondrosarcoma (p=0.002437), and sPD-L1/sPD-1 concentration ratio in chondrosarcoma is highly significantly more than 2-fold higher than in osteosarcoma (0.81 and 0.35 respectively; p=0.000284). The sensitivity of sPD-L1 ≥16.5 pg/ml test in typical osteosarcoma patients’ group comprised only 70.2%, and in those with typical chondrosarcoma - 84.6%. Serum sPD-1 and sPD-L1 concentrations in osteosarcoma and chondrosarcoma patients were not associated with the indices of tumor advancement, its histological grade, localization in the osseous system, and type of affected bone. Thus, it can be concluded that the ratio between circulating soluble forms of the receptor and the ligand of PD-1/PD-L signaling pathway differs between patients with chondrogenic and those with osteogenic tumors, sPD-L1 being diagnostically valuable mostly for chondrogenic bone neoplasms.

The heteromeric PC-1/PC-2 polycystin complex is activated by the PC-1 N-terminus

Mutations in the polycystin proteins, PC-1 and PC-2, result in autosomal dominant polycystic kidney disease (ADPKD) and ultimately renal failure. PC-1 and PC-2 enrich on primary cilia, where they are thought to form a heteromeric ion channel complex. However, a functional understanding of the putative PC-1/PC-2 polycystin complex is lacking due to technical hurdles in reliably measuring its activity. Here we successfully reconstitute the PC-1/PC-2 complex in the plasma membrane of mammalian cells and show that it functions as an outwardly rectifying channel. Using both reconstituted and ciliary polycystin channels, we further show that a soluble fragment generated from the N-terminal extracellular domain of PC-1 functions as an intrinsic agonist that is necessary and sufficient for channel activation. We thus propose that autoproteolytic cleavage of the N-terminus of PC-1, a hotspot for ADPKD mutations, produces a soluble ligand in vivo. These findings establish a mechanistic framework for understanding the role of PC-1/PC-2 heteromers in ADPKD and suggest new therapeutic strategies that would expand upon the limited symptomatic treatments currently available for this progressive, terminal disease.

Quantification of Receptor Occupancy by Ligand—An Understudied Class of Potential Biomarkers

Molecular complexes, such as ligand–receptor complexes, are vital for both health and disease and can be shed into the circulation in soluble form. Relatively little is known about the biology of soluble ligand–receptor complexes. The functional importance of such complexes and their potential use as clinical biomarkers in diagnosis and therapy remains underappreciated. Most traditional technologies used to study ligand–receptor complexes measure the individual levels of soluble ligands or receptors rather than the complexes themselves. The fraction of receptors occupied by ligand, and the potential clinical relevance of such information, has been largely overlooked. Here, we review the biological significance of soluble ligand–receptor complexes with a specific focus on their potential as biomarkers of cancer and other inflammatory diseases. In addition, we discuss a novel RNA aptamer-based technology, designated ligand–receptor complex-binding aptamers (LIRECAP), that can provide precise measurement of the fraction of a soluble receptor occupied by its ligand. The potential applicability of the LIRECAP technology as a biomarker discovery platform is also described.

A systems-biology model of the tumor necrosis factor (TNF) interactions with TNF receptor 1 and 2

Motivation Clustering enables TNF receptors to stimulate intracellular signaling. The differential soluble ligand-induced clustering behavior of TNF receptor 1 (TNFR1) and TNFR2 was modeled. A structured, rule-based model implemented ligand-independent pre-ligand binding assembly domain (PLAD)-mediated homotypic low affinity interactions of unliganded and liganded TNF receptors. Results Soluble TNF initiates TNFR1 signaling but not TNFR2 signaling despite receptor binding unless it is secondarily oligomerized. We consider high affinity binding of TNF to signaling-incompetent pre-assembled dimeric TNFR1 and TNFR2 molecules and secondary clustering of liganded dimers to signaling competent ligand–receptor clusters. Published receptor numbers, affinities and measured different activities of clustered receptors validated model simulations for a large range of receptor and ligand concentrations. Different PLAD–PLAD affinities and different activities of receptor clusters explain the observed differences in the TNF receptor stimulating activities of soluble TNF. Availability and implementation All scripts and data are in manuscript and supplement at Bioinformatics online. Supplementary information Supplementary data are available at Bioinformatics online.

Autocrine Signaling of NRP1 Ligand Galectin-1 Elicits Resistance to BRAF-Targeted Therapy in Melanoma Cells

Melanoma cells addicted to mutated BRAF oncogene activity can be targeted by specific kinase inhibitors until they develop resistance to therapy. We observed that the expression of Galectin-1 (Gal-1), a soluble ligand of Neuropilin-1 (NRP1), is upregulated in melanoma tumor samples and melanoma cells resistant to BRAF-targeted therapy. We then demonstrated that Gal-1 is a novel driver of resistance to BRAF inhibitors in melanoma and that its activity is linked to the concomitant upregulation of the NRP1 receptor observed in drug-resistant cells. Mechanistically, Gal-1 sustains increased expression of NRP1 and EGFR in drug-resistant melanoma cells. Moreover, consistent with its role as a NRP1 ligand, Gal-1 negatively controls p27 levels, a mechanism previously found to enable EGFR upregulation in cancer cells. Finally, the combined treatment with a Gal-1 inhibitor and a NRP1 blocking drug enabled resistant melanoma cell resensitization to BRAF-targeted therapy. In summary, we found that the activation of Galectin-1/NRP1 autocrine signaling is a new mechanism conferring independence from BRAF kinase activity to oncogene-addicted melanoma cells.