The mitochondrial calcium uniporter (MCU) activates mitochondrial respiration and enhances mobility by regulating mitochondrial redox state.

Regulation of mitochondrial redox balance is emerging as a key event for cell signaling in both physiological and pathological conditions. However, the link between the mitochondrial redox state and the modulation of these conditions remains poorly defined. Here, we discovered that activation of the evolutionary conserved mitochondrial calcium uniporter (MCU) modulates mitochondrial redox state.

A bioluminescent probe for longitudinal monitoring of mitochondrial membrane potential.

Mitochondrial membrane potential (ΔΨ) is a universal selective indicator of mitochondrial function and is known to play a central role in many human pathologies, such as diabetes mellitus, cancer and Alzheimer's and Parkinson's diseases. Here, we report the design, synthesis and several applications of mitochondria-activatable luciferin (MAL), a bioluminescent probe sensitive to ΔΨ, and partially to plasma membrane potential (ΔΨ), for non-invasive, longitudinal monitoring of ΔΨ in vitro and in vivo. We applied this new technology to evaluate the aging-related change of ΔΨ in mice and showed that nicotinamide riboside (NR) reverts aging-related mitochondrial depolarization, revealing another important aspect of the mechanism of action of this potent biomolecule.

Targeting Mitochondrial Calcium Uptake with the Natural Flavonol Kaempferol, to Promote Metabolism/Secretion Coupling in Pancreatic β-cells.

Pancreatic β-cells secrete insulin to lower blood glucose, following a meal. Maintenance of β-cell function is essential to preventing type 2 diabetes. In pancreatic β-cells, mitochondrial matrix calcium is an activating signal for insulin secretion.

The plant product quinic acid activates Ca -dependent mitochondrial function and promotes insulin secretion from pancreatic beta cells.

Background And Purpose: Quinic acid (QA) is an abundant natural compound from plant sources which may improve metabolic health. However, little attention has been paid to its effects on pancreatic beta-cell functions, which contribute to the control of metabolic health by lowering blood glucose. Strategies targeting beta-cell signal transduction are a new approach for diabetes treatment.

Mitochondrial lysine deacetylation promotes energy metabolism and calcium signaling in insulin-secreting cells.

In pancreatic β-cells, mitochondria generate signals that promote insulin granule exocytosis. Here we study how lysine acetylation of mitochondrial proteins mechanistically affects metabolism-secretion coupling in insulin-secreting cells. Using mass spectrometry-based proteomics, we identified lysine acetylation sites in rat insulinoma cell line clone 1E cells.

Modulation of estrogen related receptor alpha activity by the kinesin KIF17.

Estrogen-related receptor alpha (ERR1) is an orphan nuclear receptor that can bind transcriptional co-activators constitutively. ERR1 expression correlates with poor patient outcomes in breast cancer, heightening interest in this nuclear receptor as a therapeutic target. Because ERR1 has no known regulatory ligand, a major challenge in targeting its activity is to find cellular or synthetic modulators of its function.

A novel ATP-synthase-independent mechanism coupling mitochondrial activation to exocytosis in insulin-secreting cells.

Pancreatic β-cells sense glucose, promoting insulin secretion. Glucose sensing requires the sequential stimulation of glycolysis, mitochondrial metabolism and Ca entry. To elucidate how mitochondrial activation in β-cells contributes to insulin secretion, we compared the effects of glucose and the mitochondrial substrate methylsuccinate in the INS-1E insulin-secreting cell line at the respective concentrations at which they maximally activate mitochondrial respiration.

Encapsulation of Insulin-Secreting Cells Expressing a Genetically Encoded Fluorescent Calcium Indicator for Cell-Based Sensing In Vivo.

The development of cell-based biosensors that give insight into cell and tissue function in vivo is an attractive technology for biomedical research. Here, the development of a cell line expressing a fluorescent calcium sensor for the study of beta-cell function in vivo is reported. The bioresponsive cell model is based on INS-1E pancreatic beta-cells, stably expressing the genetically encoded cameleon-based fluorescent sensor YC3.

Characterization and Genetic Analyses of New Genes Coding for NOD2 Interacting Proteins.

NOD2 contributes to the innate immune response and to the homeostasis of the intestinal mucosa. In response to its bacterial ligand, NOD2 interacts with RICK and activates the NF-κB and MAPK pathways, inducing gene transcription and synthesis of proteins required to initiate a balanced immune response. Mutations in NOD2 have been associated with an increased risk of Crohn's Disease (CD), a disabling inflammatory bowel disease (IBD).

Calcium co-regulates oxidative metabolism and ATP synthase-dependent respiration in pancreatic beta cells.

Mitochondrial energy metabolism is essential for glucose-induced calcium signaling and, therefore, insulin granule exocytosis in pancreatic beta cells. Calcium signals are sensed by mitochondria acting in concert with mitochondrial substrates for the full activation of the organelle. Here we have studied glucose-induced calcium signaling and energy metabolism in INS-1E insulinoma cells and human islet beta cells.

TSG-6 inhibits neutrophil migration via direct interaction with the chemokine CXCL8.

TNF-stimulated gene/protein-6 (TSG-6) is expressed by many different cell types in response to proinflammatory cytokines and plays an important role in the protection of tissues from the damaging consequences of acute inflammation. Recently, TSG-6 was identified as being largely responsible for the beneficial effects of multipotent mesenchymal stem cells, for example in the treatment of animal models of myocardial infarction and corneal injury/allogenic transplant. The protective effect of TSG-6 is due in part to its inhibition of neutrophil migration, but the mechanisms underlying this activity remain unknown.

Chemokine receptor antagonist development.

This chapter describes assays that focus on the characterization of compounds identified in high--throughput screening campaigns, and the subsequent medicinal chemistry programs. They cover methods to determine potency in buffer, the effect of whole blood on the compounds' activity and finally the pharmacokinetic (PK)/pharmacodynamic (PD) -relationship of the compounds in a rodent species.

A high-throughput protein refolding screen in 96-well format combined with design of experiments to optimize the refolding conditions.

Production of correctly folded and biologically active proteins in Escherichiacoli can be a challenging process. Frequently, proteins are recovered as insoluble inclusion bodies and need to be denatured and refolded into the correct structure. To address this, a refolding screening process based on a 96-well assay format supported by design of experiments (DOE) was developed for identification of optimal refolding conditions.

Molecular mapping of the RNA Cap 2'-O-methyltransferase activation interface between severe acute respiratory syndrome coronavirus nsp10 and nsp16.

Several protein-protein interactions within the SARS-CoV proteome have been identified, one of them being between non-structural proteins nsp10 and nsp16. In this work, we have mapped key residues on the nsp10 surface involved in this interaction. Alanine-scanning mutagenesis, bioinformatics, and molecular modeling were used to identify several "hot spots," such as Val(42), Met(44), Ala(71), Lys(93), Gly(94), and Tyr(96), forming a continuous protein-protein surface of about 830 Å(2), bearing very conserved amino acids among coronaviruses.

MCC, a new interacting protein for Scrib, is required for cell migration in epithelial cells.

To further characterize the molecular events supporting the tumor suppressor activity of Scrib in mammals, we aim to identify new binding partners. We isolated MCC, a recently identified binding partner for beta-catenin, as a new interacting protein for Scrib. MCC interacts with both Scrib and the NHERF1/NHERF2/Ezrin complex in a PDZ-dependent manner.

Biochemical studies and molecular dynamics simulations of Smad3-Erbin interaction identify a non-classical Erbin PDZ binding.

In this work, we describe how the Erbin PDZ domain interacts with Smad3, a transductor of the Transforming Growth Factor-beta (TGFbeta) pathway, via its MH2 domain. This interaction was described as important for TGFbeta signaling as it could potentially repress the transcriptional activity of the growth factor. In order to clarify our preliminary experimental observations pointing this interaction, we built a 3D model of the Erbin PDZ/Smad3 MH2 complex and checked its stability using molecular dynamics simulations.

Identification of multi-SH3 domain-containing protein interactome in pancreatic cancer: a yeast two-hybrid approach.

Pancreatic ductal adenocarcinoma (PDAC) is a fatal disease that shows minimal response to chemotherapy. Genetic changes involved in the progression of PDAC concern genes that encode proteins related to signal transduction networks. This fact reveals the importance in identifying the role and the relations between multiple signaling cascades in PDAC.

Prevention of mantle lymphoma tumor establishment by routing transferrin receptor toward lysosomal compartments.

Mantle cell lymphoma (MCL) is one of the most frequent of the newly recognized non-Hodgkin's lymphomas. The major problem of MCL therapy is the occurrence of relapse and subsequent resistance to chemotherapy and immunotherapy in virtually all cases. Here, we show that one injection of anti-human transferrin receptor (TfR) monoclonal antibody A24 totally prevented xenografted MCL tumor establishment in nude mice.

The Salmonella effector protein PipB2 is a linker for kinesin-1.

Understanding the mechanisms of Salmonella virulence is an important challenge. The capacity of this intracellular bacterial pathogen to cause diseases depends on the expression of virulence factors including the second type III secretion system (TTSS-2), which is used to translocate into the eukaryotic cytosol a set of effector proteins that divert the biology of the host cell and shape the bacterial replicative niche. Yet little is known about the eukaryotic functions affected by individual Salmonella effectors.

Rapamycin inhibits growth and survival of D816V-mutated c-kit mast cells.

Two classes of oncogenic mutations of the c-kit tyrosine kinase have been described: the juxtamembrane domain V560G mutation, which is preferentially found in gastrointestinal stromal tumors (GISTs), and the kinase domain D816V mutation, which is highly representative of systemic mastocytosis (SM). Here we show that both mutations constitutively activate the mammalian target of rapamycin (mTOR) signaling pathway. Surprisingly, the mTOR inhibitor rapamycin induces only apoptosis in HMC-1 cells bearing the D816V but not the V560G mutation.