Défaut d’exocytose des granules lytiques : Plusieurs causes, un même effet

Une réponse immune exagérée, incontrôlée et le plus souvent fatale, connue sous le nom de syndrome hémophagocytaire (SH), est associée à un défaut de la fonction cytotoxique des lymphocytes T et natural killer (NK). Les anomalies moléculaires responsables, qui sont multiples, mettent en cause dans la plupart des cas un effecteur indispensable au fonctionnement de la machinerie lytique des lymphocytes. L’étude des lymphocytes cytotoxiques déficients en l’un ou l’autre de ces effecteurs apporte des éléments nouveaux quant à l’agencement des étapes clés de la sécrétion du contenu des granules lytiques au contact de la cellule cible. Des mécanismes moléculaires proches semblent contrôler la sécrétion vésiculaire au niveau des synapses immunologique et neurologique. D’autres effecteurs de la cytotoxicité ou du contrôle de l’homéostasie lymphocytaire à l’origine de SH doivent encore être caractérisés. Quant aux mécanismes précis de l’intervention de cette voie cytotoxique dans le maintien de l’homéostasie lymphocytaire (terminaison d’une réponse immune), ils demeurent à élucider.An in vivo disturbance of lymphocyte homeostasis occurs during the course of the hemophagocytic syndrome (HS). HS is a severe and often fatal syndrome resulting from potent and uncontrolled activation and proliferation of T-lymphocytes, mainly polyclonal CD8 lymphocytes, leading to excessive macrophage activation, high level of proinflammatory cytokine production and multiple deleterious effects. The onset of HS characterizes several inherited disorders in humans. In most of these conditions, the molecular defect impairs the granule-dependent cytotoxic activity of lymphocytes, thus highlighting the determinant role of this function in driving back the immune system to a state of equilibrium following infection. Several lines of evidence suggest that an increase in the expansion phase rather than a decrease in the contraction phase of the CD8+ T cells population characterizes the HS. Failure to kill antigen presenting cells through a transaction mechanism of cytotoxic cells should favor a sustained response, although the mechanism may be more complex than simple decrease of antigen load. Defect in the granule dependent cytotoxic function of lymphocytes result from perforin mutation in familial hemophagocytic lymphohistiocytosis type 2, from Munc13-4 (UNC13D) mutation in familial hemophagocytic lymphohistiocytosis type 3, from Rab27a mutation in Griscelli syndrome type 2, and from CHS/LYST mutation in Chediak-Higashi syndrome. The characterization of the molecular causes leading to these conditions identified Rab27a and Munc13-4 as two critical effectors of the exocytic machinery, required for the terminal transport/docking or priming of the cytotoxic granules, respectively. Different members of the Rab and Munc13 family of proteins are also used in neurotransmitter release at the neurological synapse, highlighting the similarity of the mechanisms regulating both secretory pathways. Future investigations regarding HS will continue to elucidate this exocytic pathway machinery and improve our understanding of how it finely regulates the immune response, an area that is likely to be useful for therapeutic intervention

Rab27a controls HIV-1 assembly by regulating plasma membrane levels of phosphatidylinositol 4,5-bisphosphate

During the late stages of the HIV-1 replication cycle, the viral polyprotein Pr55Gag is recruited to the plasma membrane (PM), where it binds phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2) and directs HIV-1 assembly. We show that Rab27a controls the trafficking of late endosomes carrying phosphatidylinositol 4-kinase type 2 α (PI4KIIα) toward the PM of CD4+ T cells. Hence, Rab27a promotes high levels of PM phosphatidylinositol 4-phosphate and the localized production of PI(4,5)P2, therefore controlling Pr55Gag membrane association. Rab27a also controls PI(4,5)P2 levels at the virus-containing compartments of macrophages. By screening Rab27a effectors, we identified that Slp2a, Slp3, and Slac2b are required for the association of Pr55Gag with the PM and that Slp2a cooperates with Rab27a in the recruitment of PI4KIIα to the PM. We conclude that by directing the trafficking of PI4KIIα-positive endosomes toward the PM, Rab27a controls PI(4,5)P2 production and, consequently, HIV-1 replication.Fil: Pereyra Gerber, Federico Pehuén. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; ArgentinaFil: Cabrini, Mercedes. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; ArgentinaFil: Jancic, Carolina Cristina. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Medicina Experimental. Academia Nacional de Medicina de Buenos Aires. Instituto de Medicina Experimental; ArgentinaFil: Paoletti, Luciana Elisa. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Instituto de Biología Molecular y Celular de Rosario. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Instituto de Biología Molecular y Celular de Rosario; ArgentinaFil: Banchio, Claudia Elena. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Instituto de Biología Molecular y Celular de Rosario. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Instituto de Biología Molecular y Celular de Rosario; ArgentinaFil: Von Bilderling, Catalina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Física de Buenos Aires. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Física de Buenos Aires; ArgentinaFil: Sigaut, Lorena. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Centro de Microscopías Avanzadas; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Pietrasanta, Lia. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Centro de Microscopías Avanzadas; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Duette, Gabriel. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; ArgentinaFil: Freed, Eric O.. National Cancer Institute at Frederick; Estados UnidosFil: Basile, Genevieve de Saint. Institut National de la Santé et de la Recherche Médicale; FranciaFil: Moita, Catarina Ferreira. Instituto Gulbenkian de Ciencia; PortugalFil: Moita, Luis Ferreira. Instituto Gulbenkian de Ciencia; PortugalFil: Amigorena, Sebastian. Institute Curie; FranciaFil: Benaroch, Philippe. Institute Curie; FranciaFil: Geffner, Jorge Raúl. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; ArgentinaFil: Ostrowski, Matias. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas en Retrovirus y Sida. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas en Retrovirus y Sida; Argentin

Rab27A and its effector MyRIP link secretory granules to F-actin and control their motion towards release sites

The GTPase Rab27A interacts with myosin-VIIa and myosin-Va via MyRIP or melanophilin and mediates melanosome binding to actin. Here we show that Rab27A and MyRIP are associated with secretory granules (SGs) in adrenal chromaffin cells and PC12 cells. Overexpression of Rab27A, GTPase-deficient Rab27A-Q78L, or MyRIP reduced secretory responses of PC12 cells. Amperometric recordings of single adrenal chromaffin cells revealed that Rab27A-Q78L and MyRIP reduced the sustained component of release. Moreover, these effects on secretion were partly suppressed by the actin-depolymerizing drug latrunculin but strengthened by jasplakinolide, which stabilizes the actin cortex. Finally, MyRIP and Rab27A-Q78L restricted the motion of SGs in the subplasmalemmal region of PC12 cells, as measured by evanescent-wave fluorescence microscopy. In contrast, the Rab27A-binding domain of MyRIP and a MyRIP construct that interacts with myosin-Va but not with actin increased the mobility of SGs. We propose that Rab27A and MyRIP link SGs to F-actin and control their motion toward release sites through the actin cortex

Premature birth, respiratory distress, intracerebral hemorrhage, and silvery-gray hair: differential diagnosis of the 3 types of Griscelli syndrome

A preterm neonate, born to consanguineous parents, presented with respiratory distress, intracerebral hemorrhage, and a silvery-gray sheen of the hair and eyelashes. Griscelli syndrome (GS) type 3 was diagnosed after the detection of a novel homozygous mutation of the melanophilin gene. Thus, only the hypopigmentation, but not the patient's other clinical features, were attributable to this form of GS. Differential diagnosis of the various forms of GS must be performed as early as possible as GS2 is associated with a life threatening but curable immune disorder

Actin dynamics regulation by TTC7A/PI4KIIIα limits DNA damage and cell death under confinement

Background: The actin cytoskeleton has a crucial role in the maintenance of the immune homeostasis by controlling various cellular processes, including cell migration. Mutations in TTC7A have been described as the cause of a primary immunodeficiency associated to different degrees of gut involvement and alterations in the actin cytoskeleton dynamics. Objectives: This study investigates the impact of TTC7A deficiency in immune homeostasis. In particular, the role of the TTC7A/phosphatidylinositol 4 kinase type III α pathway in the control of leukocyte migration and actin dynamics. Methods: Microfabricated devices were leveraged to study cell migration and actin dynamics of murine and patient-derived leukocytes under confinement at the single-cell level. Results: We show that TTC7A-deficient lymphocytes exhibit an altered cell migration and reduced capacity to deform through narrow gaps. Mechanistically, TTC7A-deficient phenotype resulted from impaired phosphoinositide signaling, leading to the downregulation of the phosphoinositide 3-kinase/AKT/RHOA regulatory axis and imbalanced actin cytoskeleton dynamics. TTC7A-associated phenotype resulted in impaired cell motility, accumulation of DNA damage, and increased cell death in dense 3-dimensional gels in the presence of chemokines. Conclusions: These results highlight a novel role of TTC7A as a critical regulator of lymphocyte migration. Impairment of this cellular function is likely to contribute to the pathophysiology underlying progressive immunodeficiency in patients.</p

Fine-scale detection of population-specific linkage disequilibrium using haplotype entropy in the human genome

<p>Abstract</p> <p>Background</p> <p>The creation of a coherent genomic map of recent selection is one of the greatest challenges towards a better understanding of human evolution and the identification of functional genetic variants. Several methods have been proposed to detect linkage disequilibrium (LD), which is indicative of natural selection, from genome-wide profiles of common genetic variations but are designed for large regions.</p> <p>Results</p> <p>To find population-specific LD within small regions, we have devised an entropy-based method that utilizes differences in haplotype frequency between populations. The method has the advantages of incorporating multilocus association, conciliation with low allele frequencies, and independence from allele polarity, which are ideal for short haplotype analysis. The comparison of HapMap SNPs data from African and Caucasian populations with a median resolution size of ~23 kb gave us novel candidates as well as known selection targets. Enrichment analysis for the yielded genes showed associations with diverse diseases such as cardiovascular, immunological, neurological, and skeletal and muscular diseases. A possible scenario for a selective force is discussed. In addition, we have developed a web interface (ENIGMA, available at <url>http://gibk21.bse.kyutech.ac.jp/ENIGMA/index.html</url>), which allows researchers to query their regions of interest for population-specific LD.</p> <p>Conclusion</p> <p>The haplotype entropy method is powerful for detecting population-specific LD embedded in short regions and should contribute to further studies aiming to decipher the evolutionary histories of modern humans.</p

Stepwise Maturation of Lytic Granules during Differentiation and Activation of Human CD8+ T Lymphocytes

During differentiation, cytotoxic T lymphocytes (CTL) acquire their killing potential through the biogenesis and maturation of lytic granules that are secreted upon target cell recognition. How lytic granule load in lytic molecules evolves during CTL differentiation and which subsets of lytic granules are secreted following activation remains to be investigated. We set up a flow cytometry approach to analyze single lytic granules isolated from primary human CTL according to their size and molecular content. During CTL in vitro differentiation, a relatively homogeneous population of lytic granules appeared through the progressive loading of Granzyme B, Perforin and Granzyme A within LAMP1+ lysosomes. PMA/ionomycin-induced lytic granule exocytosis was preceded by a rapid association of the docking molecule Rab27a to approximately half of the lytic granules. Activated CTL were found to limit exocytosis by sparing lytic granules including some associated to Rab27a. Our study provides a quantification of key steps of lytic granule biogenesis and highlights the potential of flow cytometry to study organelle composition and dynamics

Rab proteins and Rab-associated proteins: major actors in the mechanism of protein-trafficking disorders

Ras-associated binding (Rab) proteins and Rab-associated proteins are key regulators of vesicle transport, which is essential for the delivery of proteins to specific intracellular locations. More than 60 human Rab proteins have been identified, and their function has been shown to depend on their interaction with different Rab-associated proteins regulating Rab activation, post-translational modification and intracellular localization. The number of known inherited disorders of vesicle trafficking due to Rab cycle defects has increased substantially during the past decade. This review describes the important role played by Rab proteins in a number of rare monogenic diseases as well as common multifactorial human ones. Although the clinical phenotype in these monogenic inherited diseases is highly variable and dependent on the type of tissue in which the defective Rab or its associated protein is expressed, frequent features are hypopigmentation (Griscelli syndrome), eye defects (Choroideremia, Warburg Micro syndrome and Martsolf syndrome), disturbed immune function (Griscelli syndrome and Charcot–Marie–Tooth disease) and neurological dysfunction (X-linked non-specific mental retardation, Charcot–Marie–Tooth disease, Warburg Micro syndrome and Martsolf syndrome). There is also evidence that alterations in Rab function play an important role in the progression of multifactorial human diseases, such as infectious diseases and type 2 diabetes. Rab proteins must not only be bound to GTP, but they need also to be ‘prenylated’—i.e. bound to the cell membranes by isoprenes, which are intermediaries in the synthesis of cholesterol (e.g. geranyl geranyl or farnesyl compounds). This means that isoprenylation can be influenced by drugs such as statins, which inhibit isoprenylation, or biphosphonates, which inhibit that farnesyl pyrophosphate synthase necessary for Rab GTPase activity. Conclusion: Although protein-trafficking disorders are clinically heterogeneous and represented in almost every subspeciality of pediatrics, the identification of common pathogenic mechanisms may provide a better diagnosis and management of patients with still unknown Rab cycle defects and stimulate the development of therapeutic agents