How important are Rho GTPases in neurosecretion?

International audienc

18F-FDOPA PET/CT Uptake Parameters Correlate with Catecholamine Secretion in Human Pheochromocytomas

International audienceBackground: 18 F-FDOPA positron emission tomography/ computed tomography (PET/CT) is a sensitive nuclear imaging technology for the diagnosis of pheochromocytomas (PHEO). However, its utility in determining predictive factors for the secretion of catecholamines remains poorly studied. Methods: Thirty-nine histologically confirmed PHEO were included in this retrospective single-center study. Patients underwent 18 F-FDOPA PET/CT before surgery, with an evaluation of several uptake parameters (standardized uptake values [SUV max and SUV mean ] and the metabolic burden [MB] calculated as follows: MB = SUV mean × tumor volume) and measurement of plasma and/or urinary metanephrine (MN), normetanephrine (NM), and chromogranin A. Thirty-five patients were screened for germline mutations in the RET, SDHx, and VHL genes. Once resected, primary cultures of 5 PHEO were used for real-time measurement of catechol-amine release by carbon fiber amperometry. Results: The MB of the PHEO positively correlated with 24-h urinary excre-tion of NM (r = 0.64, p < 0.0001), MN (r = 0.49, p = 0.002), combined MN and NM (r = 0.75, p < 0.0001), and eventually plasma free levels of NM (r = 0.55, p = 0.006). In the mutated patients (3 SDHD, 2 SDHB, 3 NF1, 1 VHL, and 3 RET), a similar correlation was observed between MB and 24-h urinary combined MN and NM (r = 0.86, p = 0.0012). For the first time, we demonstrate a positive correlation between the PHEO-to-liver SUV max ratio and the mean number of secretory granule fusion events of the corresponding PHEO cells revealed by amperometric spikes (p = 0.01). Conclusion: While the 18 F-FDOPA PET/CT MB of PHEO strongly correlates with the concentration of MN, amperometric recordings suggest that 18 F-FDOPA uptake could be enhanced by overactivity of cat-echolamine exocytosis

Exocytosis and Endocytosis in Neuroendocrine Cells: Inseparable Membranes!

International audienceAlthough much has been learned concerning the mechanisms of secretory vesicle formation and fusion at donor and acceptor membrane compartments, relatively little attention has been paid toward understanding how cells maintain a homeostatic membrane balance through vesicular trafficking. In neurons and neuroendocrine cells, release of neurotrans-mitters, neuropeptides, and hormones occurs through calcium-regulated exocytosis at the plasma membrane. To allow recycling of secretory vesicle components and to preserve organelles integrity, cells must initiate and regulate compensatory membrane uptake. This review relates the fate of secretory granule membranes after full fusion exocytosis in neuroendocrine cells. In particular, we focus on the potential role of lipids in preserving and sorting secretory granule membranes after exocytosis and we discuss the potential mechanisms of membrane retrieval

RhoGTPase Regulators Orchestrate Distinct Stages of Synaptic Development

Small RhoGTPases regulate changes in post-synaptic spine morphology and density that support learning and memory. They are also major targets of synaptic disorders, including Autism. Here we sought to determine whether upstream RhoGTPase regulators, including GEFs, GAPs, and GDIs, sculpt specific stages of synaptic development. The majority of examined molecules uniquely regulate either early spine precursor formation or later matura- tion. Specifically, an activator of actin polymerization, the Rac1 GEF β-PIX, drives spine pre- cursor formation, whereas both FRABIN, a Cdc42 GEF, and OLIGOPHRENIN-1, a RhoA GAP, regulate spine precursor elongation. However, in later development, a novel Rac1 GAP, ARHGAP23, and RhoGDIs inactivate actomyosin dynamics to stabilize mature synap- ses. Our observations demonstrate that specific combinations of RhoGTPase regulatory pro- teins temporally balance RhoGTPase activity during post-synaptic spine development

Rho GTPases and exocytosis: What are the molecular links?

International audienceDelivery of proteins or lipids to the plasma membrane or into the extracellular space occurs through exocytosis, a process that requires tethering, docking, priming and fusion of vesicles, as well as F-actin rearrangements in response to specific extracellular cues. GTPases of the Rho family have been implicated as important regulators of exocytosis, but how Rho proteins control this process is an open question. In this review, we focus on molecular connections that drive Rho-dependent exocytosis in polarized and regulated exocytosis. Specifically, we present data showing that Rho proteins interaction with the exocyst complex and IQGAP mediates polarized exocytosis, whereas interaction with actin-binding proteins like N-WASP mediates regulated exocytosis

The regulated secretory pathway in neuroendocrine cells

The regulated secretory pathway is a hallmark of neuroendocrine cells. This process comprises many sequential steps, which include ER-associated protein synthesis, post-translational modification of proteins in the Golgi complex, sorting and packing of secretory proteins into carrier granules, cytoskeleton-based granule transport towards the plasma membrane and tethering, docking and fusion of granules with specialized releasing zones. Each stage is subjected to a rigorous regulation by a plethora of factors that function in a spatially and temporarily coordinated fashion. Much effort has been devoted to characterize the precise role of the regulatory proteins participating in the different steps of this process and to identify new factors in order to obtain a unifying picture of the secretory pathway. In spite of this and given the enormous complexity of the process, certain stages are not fully understood yet and many players remain to be identified. The aim of this Research Topic is to gather review articles and original research papers on the molecular mechanisms that govern and ensure the correct release of neuropeptides

A lipidomics approach to measure phosphatidic acid species in subcellular membrane fractions obtained from cultured cells

Over the last decade, lipids have emerged as possessing an ever-increasing number of key functions, especially in membrane trafficking. For instance, phosphatidic acid (PA) has been proposed to play a critical role in different steps along the secretory pathway or during phagocytosis. To further investigate in detail the precise nature of PA activities, we need to identify the organelles in which PA is synthesized and the PA subspecies involved in these biological functions. Indeed, PA, like all phospholipids, has a large variety based on its fatty acid composition. The recent development of PA sensors has helped us to follow intracellular PA dynamics but has failed to provide information on individual PA species. Here, we describe a method for the subcellular fractionation of RAW264.7 macrophages that allows us to obtain membrane fractions enriched in specific organelles based on their density. Lipids from these membrane fractions are precipitated and subsequently processed by advanced mass spectrometry-based lipidomics analysis to measure the levels of different PA species based on their fatty acyl chain composition. This approach revealed the presence of up to 50 different species of PA in cellular membranes, opening up the possibility that a single class of phospholipid could play multiple functions in any given organelle. This protocol can be adapted or modified and used for the evaluation of other intracellular membrane compartments or cell types of interest