Contribution à l'étude du rôle de la Chromogranine A dans la sécrétion neuroendocrine : identification de ses partenaires moléculaires impliqués dans la biogenèse des granules de sécrétion

ROUEN-BU Sciences (764512102) / SudocSudocFranceF

Chromogranin A as a Crucial Factor in the Sorting of Peptide Hormones to Secretory Granules

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Skin biological responses to urban pollution in an ex vivo model

International audienceThe skin epidermis is continuously exposed to external aggressions, including environmental pollution. The cosmetic industry must be able to offer dedicated products to fight the effectsof pollutants on the skin. We set up an experimental model that exposed skin explantsmaintained in culture to a pollutant mixture. This mixture representing urban pollution wasdesigned on the basis of the French organization ‘Air Parif’ database. A chamber, calledPollubox®, was built to allow a controlled nebulization of P on the cultured human skinexplants. We investigated ultrastructural morphology by transmission electron microscopy ofhigh pressure frozen skin explants. A global transcriptomic analysis indicated that the pollutant mixture was able toinduce relevant xenobiotic and antioxidant responses. Modulated detoxifying genes werefurther investigated by laser micro-dissection coupled to qPCR, and immunochemistry. Bothapproaches showed that P exposure correlated with overexpression of detoxifying genes andprovoked skin physiological alterations down to the stratum basale. The model developedherein might be an efficient tool to study the effects of pollutants on skin as well as apowerful testing method to evaluate the efficacy of cosmetic products against pollution

Myosin 1b and F-actin are involved in the control of secretory granule biogenesis

Hormone secretion relies on secretory granules which store hormones in endocrine cells and release them upon cell stimulation. The molecular events leading to hormone sorting and secretory granule formation at the level of the TGN are still elusive. Our proteomic analysis of purified whole secretory granules or secretory granule membranes uncovered their association with the actomyosin components myosin 1b, actin and the actin nucleation complex Arp2/3. We found that myosin 1b controls the formation of secretory granules and the associated regulated secretion in both neuroendocrine cells and chromogranin A-expressing COS7 cells used as a simplified model of induced secretion. We show that F-actin is also involved in secretory granule biogenesis and that myosin 1b cooperates with Arp2/3 to recruit F-actin to the Golgi region where secretory granules bud. These results provide the first evidence that components of the actomyosin complex promote the biogenesis of secretory granules and thereby regulate hormone sorting and secretion.This work was supported by Institut National de la Santé et de la Recherche Médicale, the University of Rouen Normandy, the Conseil Régional de Normandie and the Ministère de l’Enseignement Supérieur et de la RecherchePeer Reviewe

Chromogranin A Induces the Biogenesis of Granules with Calcium- and Actin-Dependent Dynamics and Exocytosis in Constitutively Secreting Cells

International audienceChromogranins are a family of acidic glycoproteins that play an active role in hormone and neuro-peptide secretion through their crucial role in secretory granule biogenesis in neuroendocrine cells. However, the molecular mechanisms underlying their granulogenic activity are still not fully understood. Because we previously demonstrated that the expression of the major component of secretory granules, chromogranin A (CgA), is able to induce the formation of secretory granules in nonendocrine COS-7 cells, we decided to use this model to dissect the mechanisms triggered by CgA leading to the biogenesis and trafficking of such granules. Using quantitative live cell imaging, we first show that CgA-induced organelles exhibit a Ca 2ϩ-dependent trafficking, in contrast to native vesicle stomatitis virus G protein-containing constitutive vesicles. To identify the proteins that confer such properties to the newly formed granules, we developed CgA-stably-expressing COS-7 cells, purified their CgA-containing granules by subcellular fractionation, and analyzed the granule proteome by liquid chromatography tandem mass spectrometry. This analysis revealed the association of several cytosolic proteins to the granule membrane, including GTPases, cytoskeleton-based molecular motors, and other proteins with actin-and/or Ca 2ϩ-binding properties. Furthermore, disruption of cytoskeleton affects not only the distribution and the transport but also the Ca 2ϩ-evoked exocytosis of the CgA-containing granules, indicating that these granules interact with microtubules and cortical actin for the regulated release of their content. These data demonstrate for the first time that the neuroendocrine factor CgA induces the recruitment of cytoskeleton-, GTP-, and Ca 2ϩ-binding proteins in constitutively secreting COS-7 cells to generate vesicles endowed with typical dynamics and exocytotic properties of neuroen-docrine secretory granules. (Endocrinology 153: 0000-0000, 2012

Myosin 1b and F-actin are involved in the control of secretory granule biogenesis

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Chromogranin A preferential interaction with Golgi phosphatidic acid induces membrane deformation and contributes to secretory granule biogenesis

International audienceChromogranin A (CgA) is a key luminal actor of secretory granule biogenesis at the trans‐Golgi network (TGN) level but the molecular mechanisms involved remain obscure. Here, we investigated the possibility that CgA acts synergistically with specific membrane lipids to trigger secretory granule formation. We show that CgA preferentially interacts with the anionic glycerophospholipid phosphatidic acid (PA). In accordance, bioinformatic analysis predicted a PA‐binding domain (PABD) in CgA sequence that effectively bound PA (36:1) or PA (40:6) in membrane models. We identified PA (36:1) and PA (40:6) as predominant species in Golgi and granule membranes of secretory cells, and we found that CgA interaction with these PA species promotes artificial membrane deformation and remodeling. Furthermore, we demonstrated that disruption of either CgA PABD or phospholipase D (PLD) activity significantly alters secretory granule formation in secretory cells. Our findings show for the first time the ability of CgA to interact with PLD‐generated PA, which allows membrane remodeling and curvature, key processes necessary to initiate secretory granule budding