MYO1C stabilizes actin and facilitates the arrival of transport carriers at the Golgi complex

In this study, we aimed to identify the myosin motor proteins that control trafficking at the Golgi complex. In addition to the known Golgi-associated myosins MYO6, MYO18A and MYH9 (myosin IIA), we identified MYO1C as a novel player at the Golgi in a human cell line. We demonstrate that depletion of MYO1C induces Golgi complex fragmentation and decompaction. MYO1C accumulates at dynamic structures around the Golgi complex that colocalize with Golgi-associated actin dots. MYO1C depletion leads to loss of cellular F-actin, and Golgi complex decompaction is also observed after inhibition or loss of the actin-related protein 2/3 complex, Arp2/3 (also known as ARPC). We show that the functional consequence of MYO1C depletion is a delay in the arrival of incoming transport carriers, both from the anterograde and retrograde routes. We propose that MYO1C stabilizes actin at the Golgi complex, facilitating the arrival of incoming transport carriers at the Golgi.This article has an associated First Person interview with the first author of the paper.Fil: Capmany, Anahi. Institute Curie; Francia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza; Argentina. Universidad Nacional de Cuyo. Facultad de Ciencias Medicas. Instituto de Inmunologia; ArgentinaFil: Yoshimura, Azumi. Institute Curie; FranciaFil: Kerdous, Rachid. Institute Curie; FranciaFil: Caorsi, Valentina. Abbelight; FranciaFil: Lescure, Aurianne. Institute Curie; FranciaFil: Nery, Elaine Del. Institute Curie; FranciaFil: Coudrier, Evelyne. Institute Curie; FranciaFil: Goud, Bruno. Institute Curie; FranciaFil: Schauer, Kristine. Institute Curie; Franci

Hierarchical regulation of the NikR-mediated nickel response in Helicobacter pylori

Nickel is an essential metal for Helicobacter pylori, as it is the co-factor of two enzymes crucial for colonization, urease and hydrogenase. Nickel is taken up by specific transporters and its intracellular homeostasis depends on nickel-binding proteins to avoid toxicity. Nickel trafficking is controlled by the Ni(II)-dependent transcriptional regulator NikR. In contrast to other NikR proteins, NikR from H. pylori is a pleiotropic regulator that depending on the target gene acts as an activator or a repressor. We systematically quantified the in vivo Ni2+-NikR response of 11 direct NikR targets that encode functions related to nickel metabolism, four activated and seven repressed genes. Among these, four targets were characterized for the first time (hpn, hpn-like, hydA and hspA) and NikR binding to their promoter regions was demonstrated by electrophoretic mobility shift assays. We found that NikR-dependent repression was generally set up at higher nickel concentrations than activation. Kinetics of the regulation revealed a gradual and temporal NikR-mediated response to nickel where activation of nickel-protection mechanisms takes place before repression of nickel uptake. Our in vivo study demonstrates, for the first time, a chronological hierarchy in the NikR-dependent transcriptional response to nickel that is coherent with the control of nickel homeostasis in H. pylori

Rab27a and Rab27b control different steps of the exosome secretion pathway

Exosomes are secreted membrane vesicles that share structural and biochemical characteristics with intraluminal vesicles of multivesicular endosomes (MVEs). Exosomes could be involved in intercellular communication and in the pathogenesis of infectious and degenerative diseases. The molecular mechanisms of exosome biogenesis and secretion are, however, poorly understood. Using an RNA interference (RNAi) screen, we identified five Rab GTPases that promote exosome secretion in HeLa cells. Among these, Rab27a and Rab27b were found to function in MVE docking at the plasma membrane. The size of MVEs was strongly increased by Rab27a silencing, whereas MVEs were redistributed towards the perinuclear region upon Rab27b silencing. Thus, the two Rab27 isoforms have different roles in the exosomal pathway. In addition, silencing two known Rab27 effectors, Slp4 (also known as SYTL4, synaptotagmin-like 4) and Slac2b (also known as EXPH5, exophilin 5), inhibited exosome secretion and phenocopied silencing of Rab27a and Rab27b, respectively. Our results therefore strengthen the link between MVEs and exosomes, and introduce ways of manipulating exosome secretion in vivo

Does the Actin Network Architecture Leverage Myosin-I Functions?

International audienceThe actin cytoskeleton plays crucial roles in cell morphogenesis and functions. The main partners of cortical actin are molecular motors of the myosin superfamily. Although our understanding of myosin functions is heavily based on myosin-II and its ability to dimerize, the largest and most ancient class is represented by myosin-I. Class 1 myosins are monomeric, actin-based motors that regulate a wide spectrum of functions, and whose dysregulation mediates multiple human diseases. We highlight the current challenges in identifying the “pantograph” for myosin-I motors: we need to reveal how conformational changes of myosin-I motors lead to diverse cellular as well as multicellular phenotypes. We review several mechanisms for scaling, and focus on the (re-) emerging function of class 1 myosins to remodel the actin network architecture, a higher-order dynamic scaffold that has potential to leverage molecular myosin-I functions. Undoubtfully, understanding the molecular functions of myosin-I motors will reveal unexpected stories about its big partner, the dynamic actin cytoskeleton

Study of the nickel metabolism of Helicobacter pylori

PARIS7-Bibliothèque centrale (751132105) / SudocSudocFranceF

Quantifying Spatiotemporal Parameters of Cellular Exocytosis in Micropatterned Cells

International audienc

Novel nickel transport mechanism across the bacterial outer membrane energized by the TonB/ExbB/ExbD machinery

International audienceNickel is a cofactor for various microbial enzymes, yet as a trace element, its scavenging is challenging. In the case of the pathogen Helicobacter pylori, nickel is essential for the survival in the human stomach, because it is the cofactor of the important virulence factor urease. While nickel transport across the cytoplasmic membrane is accomplished by the nickel permease NixA, the mechanism by which nickel traverses the outer membrane (OM) of this Gram‐negative bacterium is unknown. Import of iron‐siderophores and cobalamin through the bacterial OM is carried out by specific receptors energized by the TonB/ExbB/ExbD machinery. In this study, we show for the first time that H. pylori utilizes TonB/ExbB/ExbD for nickel uptake in addition to iron acquisition. We have identified the nickel‐regulated protein FrpB4, homologous to TonB‐dependent proteins, as an OM receptor involved in nickel uptake. We demonstrate that ExbB/ExbD/TonB and FrpB4 deficient bacteria are unable to efficiently scavenge nickel at low pH. This condition mimics those encountered by H. pylori during stomach colonization, under which nickel supply and full urease activity are essential to combat acidity. We anticipate that this nickel scavenging system is not restricted to H. pylori, but will be represented more largely among Gram‐negative bacteria

The Helicobacter pylori GroES Cochaperonin HspA Functions as a Specialized Nickel Chaperone and Sequestration Protein through Its Unique C-Terminal Extension ▿ †

The transition metal nickel plays a central role in the human gastric pathogen Helicobacter pylori because it is required for two enzymes indispensable for colonization, the nickel metalloenzyme urease and [NiFe] hydrogenase. To sustain nickel availability for these metalloenzymes while providing protection from the metal's harmful effects, H. pylori is equipped with several specific nickel-binding proteins. Among these, H. pylori possesses a particular chaperone, HspA, that is a homolog of the highly conserved and essential bacterial heat shock protein GroES. HspA contains a unique His-rich C-terminal extension and was demonstrated to bind nickel in vitro. To investigate the function of this extension in H. pylori, we constructed mutants carrying either a complete deletion or point mutations in critical residues of this domain. All mutants presented a decreased intracellular nickel content measured by inductively coupled plasma mass spectrometry (ICP-MS) and reduced nickel tolerance. While urease activity was unaffected in the mutants, [NiFe] hydrogenase activity was significantly diminished when the C-terminal extension of HspA was mutated. We conclude that H. pylori HspA is involved in intracellular nickel sequestration and detoxification and plays a novel role as a specialized nickel chaperone involved in nickel-dependent maturation of hydrogenase