Primary and secondary dimer interfaces of the fibroblast growth factor receptor 3 transmembrane domain: characterization via multiscale molecular dynamics simulations.

Receptor tyrosine kinases are single-pass membrane proteins that form dimers within the membrane. The interactions of their transmembrane domains (TMDs) play a key role in dimerization and signaling. Fibroblast growth factor receptor 3 (FGFR3) is of interest as a G380R mutation in its TMD is the underlying cause of ~99% of the cases of achondroplasia, the most common form of human dwarfism. The structural consequences of this mutation remain uncertain: the mutation shifts the position of the TMD relative to the lipid bilayer but does not alter the association free energy. We have combined coarse-grained and all-atom molecular dynamics simulations to study the dimerization of wild-type, heterodimer, and mutant FGFR3 TMDs. The simulations reveal that the helices pack together in the dimer to form a flexible interface. The primary packing mode is mediated by a Gx3G motif. There is also a secondary dimer interface that is more highly populated in heterodimer and mutant configurations that may feature in the molecular mechanism of pathology. Both coarse-grained and atomistic simulations reveal a significant shift of the G380R mutant dimer TMD relative to the bilayer to allow interactions of the arginine side chain with lipid headgroup phosphates.

Radiation therapy after radical surgery in prostate cancer.

Radiation therapy plays a key role in the treatment of prostate cancer on its own. For higher risk diseases, the risk of recurrence following single modality therapy increases and a combination of treatment modalities may be necessary to achieve optimal results. We review clinical outcomes of adjuvant and salvage radiotherapy following radical prostatectomy, including disease-free survival, cancer-specific survival and overall survival. We also discuss when best to intervene with post-prostatectomy radiotherapy.

A variant in IL6ST with a selective IL-11 signaling defect in human and mouse.

The GP130 cytokine receptor subunit encoded by IL6ST is the shared receptor for ten cytokines of the IL-6 family. We describe a homozygous non-synonymous variant in IL6ST (p.R281Q) in a patient with craniosynostosis and retained deciduous teeth. We characterize the impact of the variant on cytokine signaling in vitro using transfected cell lines as well as primary patient-derived cells and support these findings using a mouse model with the corresponding genome-edited variant Il6st p.R279Q. We show that human GP130 p.R281Q is associated with selective loss of IL-11 signaling without affecting IL-6, IL-27, OSM, LIF, CT1, CLC, and CNTF signaling. In mice Il6st p.R279Q lowers litter size and causes facial synostosis and teeth abnormalities. The effect on IL-11 signaling caused by the GP130 variant shows incomplete penetrance but phenocopies aspects of IL11RA deficiency in humans and mice. Our data show that a genetic variant in a pleiotropic cytokine receptor can have remarkably selective defects.

Endocytic sorting motif interactions involved in Nef-mediated downmodulation of CD4 and CD3.

Lentiviral Nefs recruit assembly polypeptide complexes and target sorting motifs in cellular receptors to induce their internalization. While Nef-mediated CD4 downmodulation is conserved, the ability to internalize CD3 was lost in HIV-1 and its precursors. Although both functions play key roles in lentiviral replication and pathogenicity, the underlying structural requirements are poorly defined. Here, we determine the structure of SIVmac239 Nef bound to the ExxxLM motif of another Nef molecule at 2.5 Å resolution. This provides a basis for a structural model, where a hydrophobic crevice in simian immunodeficiency virus (SIV) Nef targets a dileucine motif in CD4 and a tyrosine-based motif in CD3. Introducing key residues into this crevice of HIV-1 Nef enables CD3 binding but an additional N-terminal tyrosine motif is required for internalization. Our resolution of the CD4/Nef/AP2 complex and generation of HIV-1 Nefs capable of CD3 downregulation provide insights into sorting motif interactions and target discrimination of Nef.HIV and simian immunodeficiency virus (SIV) Nef proteins both stimulate the clathrin-mediated endocytosis of CD4 but differ in downmodulation of the immune receptor CD3. Here, the authors present the structure of SIV Nef bound to the ExxxLM motif of another Nef molecule, which allows them to propose a model how Nef recognizes these motifs in CD3 and CD4.

A biallelic mutation in IL6ST encoding the GP130 co-receptor causes immunodeficiency and craniosynostosis.

Multiple cytokines, including interleukin 6 (IL-6), IL-11, IL-27, oncostatin M (OSM), and leukemia inhibitory factor (LIF), signal via the common GP130 cytokine receptor subunit. In this study, we describe a patient with a homozygous mutation of IL6ST (encoding GP130 p.N404Y) who presented with recurrent infections, eczema, bronchiectasis, high IgE, eosinophilia, defective B cell memory, and an impaired acute-phase response, as well as skeletal abnormalities including craniosynostosis. The p.N404Y missense substitution is associated with loss of IL-6, IL-11, IL-27, and OSM signaling but a largely intact LIF response. This study identifies a novel immunodeficiency with phenotypic similarities to STAT3 hyper-IgE syndrome caused by loss of function of GP130.

Tratamiento hipolipemiante y su efectividad en pacientes de cuatro ciudades colombianas / Lipid-lowering therapy and its effectiveness in patients from four Colombian cities

Resumen Introducción: la prevalencia de enfermedad cardiovascular ha aumentado, la hipercolesterolemia se ha asociado como factor de riesgo y se desconoce en algunas poblaciones cuántos pacientes logran el control metabólico. Objetivo: determinar la efectividad del tratamiento hipolipemiante en pacientes afiliados al sistema de salud colombiano. Métodos: estudio de corte transversal para determinar la efectividad del tratamiento hipolipemiante en una muestra aleatoria de 211 pacientes adultos tratados con estatinas, en cuatro ciudades colombianas entre 2012 y 2013. Se obtuvieron variables sociodemográficas, antropométricas, cardiovasculares, farmacológicas y paraclínicas (Colesterol Total (CT), LDL, HDL, triglicéridos). Resultados: se encontró predominio femenino (56,4%), promedio de edad 64±12 años, tratados en monoterapia con atorvastatina 78.6% y lovastatina 19.9%. El 43.4% de los casos de alto riesgo alcanzó la meta de C-LDL. El promedio de colesterol-LDL inicial en pacientes con atorvastatina fue de 103.7±48.2 mg/dL y final fue 81.9±31.2 mg/dL con una reducción promedio de 21.0% de los valores, esta diferencia fue estadísticamente significativa (p<0.001). Ser tratado en la ciudad de Manizales (OR: 2.44; IC95%: 1.029-5.789; p=0.043) y tener historia personal de infarto agudo de miocardio (OR:2.96; IC95%: 1.256-6.980; p=0.013) se asociaron con aumento de la probabilidad de controlar la dislipidemia, mientras que ser tratado con lovastatina (OR:0.03; IC95%: 0.855-4.614; p=0.02) se asoció con disminución en la probabilidad de lograr el control. Discusión: el tratamiento hipolipemiante fue realizado principalmente con atorvastatina seguido por una baja proporción de lovastatina; sin embargo, los pacientes de alto riesgo cardiovascular tratados con este último tuvieron menor probabilidad de llegar al control metabólico. (Acta Med Colomb 2016; 41: 181-186).

Abstract Introduction: The prevalence of cardiovascular disease has increased; hypercholesterolemia has been associated as a risk factor and the number of patients achieving metabolic control in some populations is unknown. Objective: To determine the effectiveness of lipid-lowering therapy in patients affiliated to the Colombian Health System. Methods: A cross-sectional study to determine the effectiveness of lipid-lowering therapy in a random sample of 211 adult patients treated with statins in four Colombian cities between 2012 and 2013. Sociodemographic, anthropometric, cardiovascular, pharmacological and paraclinical variables were obtained (total cholesterol (CT), LDL, HDL, triglycerides). Results: There was female predominance (56.4%), mean age 64 ± 12 years, 78.6% were treated with atorvastatin monotherapy and 19.9% with lovastatin. 43.4% of high-risk cases reached the target of C-LDL. The average initial LDL cholesterol in patients with atorvastatin was 103.7 ± 48.2 mg / dL and the final was 81.9 ± 31.2 mg / dL with an average reduction of 21.0% of the values. This difference was statistically significant (p <0.001). Being treated in the city of Manizales (OR: 2.44; 95% CI: 1029-5789; p = 0.043) and having personal history of acute myocardial infarction (OR: 2.96; 95% CI: 1256-6980; p = 0.013) were associated with increased likelihood of controlling dyslipidemia while being treated with lovastatin (OR: 0.03; 95% CI: 0855-4614; p = 0.02) was associated with decreased likelihood of achieving control. Discussion: lipid-lowering therapy was performed mainly with atorvastatin followed by a low proportion of lovastatin; however patients at high cardiovascular risk treated with the latter were less likely to get metabolic control. (Acta Med Colomb 2016; 41: 181-186).

HIV-1 Nef induces proinflammatory state in macrophages through its acidic cluster domain: involvement of TNF alpha receptor associated factor 2.

BACKGROUND: HIV-1 Nef is a virulence factor that plays multiple roles during HIV replication. Recently, it has been described that Nef intersects the CD40 signalling in macrophages, leading to modification in the pattern of secreted factors that appear able to recruit, activate and render T lymphocytes susceptible to HIV infection. The engagement of CD40 by CD40L induces the activation of different signalling cascades that require the recruitment of specific tumor necrosis factor receptor-associated factors (i.e. TRAFs). We hypothesized that TRAFs might be involved in the rapid activation of NF-κB, MAPKs and IRF-3 that were previously described in Nef-treated macrophages to induce the synthesis and secretion of proinflammatory cytokines, chemokines and IFNß to activate STAT1, -2 and -3. METHODOLOGY/PRINCIPAL FINDINGS: Searching for possible TRAF binding sites on Nef, we found a TRAF2 consensus binding site in the AQEEEE sequence encompassing the conserved four-glutamate acidic cluster. Here we show that all the signalling effects we observed in Nef treated macrophages depend on the integrity of the acidic cluster. In addition, Nef was able to interact in vitro with TRAF2, but not TRAF6, and this interaction involved the acidic cluster. Finally silencing experiments in THP-1 monocytic cells indicate that both TRAF2 and, surprisingly, TRAF6 are required for the Nef-induced tyrosine phosphorylation of STAT1 and STAT2. CONCLUSIONS: Results reported here revealed TRAF2 as a new possible cellular interactor of Nef and highlighted that in monocytes/macrophages this viral protein is able to manipulate both the TRAF/NF-κB and TRAF/IRF-3 signalling axes, thereby inducing the synthesis of proinflammatory cytokines and chemokines as well as IFNß.

In vitro treatment of human monocytes/macrophages with myristoylated recombinant Nef of human immunodeficiency virus type 1 leads to the activation of mitogen-activated protein kinases, IkappaB kinases, and interferon regulatory factor 3 and to the release of beta interferon.

The viral protein Nef is a virulence factor that plays multiple roles during the early and late phases of human immunodeficiency virus (HIV) replication. Nef regulates the cell surface expression of critical proteins (including down-regulation of CD4 and major histocompatibility complex class I), T-cell receptor signaling, and apoptosis, inducing proapoptotic effects in uninfected bystander cells and antiapoptotic effects in infected cells. It has been proposed that Nef intersects the CD40 ligand signaling pathway in macrophages, leading to modification in the pattern of secreted factors that appear able to recruit and activate T lymphocytes, rendering them susceptible to HIV infection. There is also increasing evidence that in vitro cell treatment with Nef induces signaling effects. Exogenous Nef treatment is able to induce apoptosis in uninfected T cells, maturation in dendritic cells, and suppression of CD40-dependent immunoglobulin class switching in B cells. Previously, we reported that Nef treatment of primary human monocyte-derived macrophages (MDMs) induces a cycloheximide-independent activation of NF-kappaB and the synthesis and secretion of a set of chemokines/cytokines that activate STAT1 and STAT3. Here, we show that Nef treatment is capable of hijacking cellular signaling pathways, inducing a very rapid regulatory response in MDMs that is characterized by the rapid and transient phosphorylation of the alpha and beta subunits of the IkappaB kinase complex and of JNK, ERK1/2, and p38 mitogen-activated protein kinase family members. In addition, we have observed the activation of interferon regulatory factor 3, leading to the synthesis of beta interferon mRNA and protein, which in turn induces STAT2 phosphorylation. All of these effects require Nef myristoylation.