Bardet-Biedl syndrome (BBS) is an inherited ciliopathy generally associated with severe obesity, but the underlying mechanism remains hypothetical and is generally proposed to be of neuroendocrine origin. In this study, we show that while the proliferating preadipocytes or mature adipocytes are nonciliated in culture, a typical primary cilium is present in differentiating preadipocytes. This transient cilium carries receptors for Wnt and Hedgehog pathways, linking this organelle to previously described regulatory pathways of adipogenesis. We also show that the BBS10 and BBS12 proteins are located within the basal body of this primary cilium and inhibition of their expression impairs ciliogenesis, activates the glycogen synthase kinase 3 pathway, and induces peroxisome proliferator-activated receptor nuclear accumulation, hence favoring adipogenesis. Moreover, adipocytes derived from BBSpatients' dermal fibroblasts in culture exhibit higher propensity for fat accumulation when compared to controls. This strongly suggests that a peripheral primary dysfunction of adipogenesis participates to the pathogenesis of obesity in BBS.adipogenesis ͉ primary cilium ͉ ciliopathy ͉ obesity

BackgroundNeutrophils polarize and migrate in response to chemokines. Different types of membrane microdomains (rafts) have been postulated to be present in rear and front of polarized leukocytes and disruption of rafts by cholesterol sequestration prevents leukocyte polarization. Reggie/flotillin-1 and -2 are two highly homologous proteins that are ubiquitously enriched in detergent resistant membranes and are thought to shape membrane microdomains by forming homo- and hetero-oligomers. It was the goal of this study to investigate dynamic membrane microdomain reorganization during neutrophil activation.Methodology/Principal FindingsWe show now, using immunofluorescence staining and co-immunoprecipitation, that endogenous flotillin-1 and -2 colocalize and associate in resting spherical and polarized primary neutrophils. Flotillins redistribute very early after chemoattractant stimulation, and form distinct caps in more than 90% of the neutrophils. At later time points flotillins accumulate in the uropod of polarized cells. Chemotactic peptide-induced redistribution and capping of flotillins requires integrity and dynamics of the actin cytoskeleton, but does not involve Rho-kinase dependent signaling related to formation of the uropod. Both flotillin isoforms are involved in the formation of this membrane domain, as uropod location of exogenously expressed flotillins is dramatically enhanced by co-overexpression of tagged flotillin-1 and -2 in differentiated HL-60 cells as compared to cells expressing only one tagged isoform.Flotillin-1 and -2 associate with P-selectin glycoprotein ligand 1 (PSGL-1) in resting and in stimulated neutrophils as shown by colocalization and co-immunoprecipitation. Neutrophils isolated from PSGL-1-deficient mice exhibit flotillin caps to the same extent as cells isolated from wild type animals, implying that PSGL-1 is not required for the formation of the flotillin caps. Finally we show that stimulus-dependent redistribution of other uropod-located proteins, CD43 and ezrin/radixin/moesin, occurs much slower than that of flotillins and PSGL-1.Conclusions/SignificanceThese results suggest that flotillin-rich actin-dependent membrane microdomains are importantly involved in neutrophil uropod formation and/or stabilization and organize uropod localization of PSGL-1.

Studies of the primary cilium, now known to be present in all cells, have undergone a revolution, in part, because mutation of many of its proteins causes a large number of diseases, including cystic kidney disease. Bardet-Biedl syndrome (BBS) is an inherited ciliopathy characterized, among other dysfunctions, by renal defects for which the precise role of the cilia in kidney function remains unclear. We studied a cohort of patients with BBS where we found that these patients had a urinary concentration defect even when kidney function was near normal and in the absence of major cyst formation. Subsequent in vitro analysis showed that renal cells in which a BBS gene was knocked down were unciliated, but did not exhibit cell cycle defects. As the vasopressin receptor 2 is located in the primary cilium, we studied BBS-derived unciliated renal epithelial cells and found that they were unable to respond to luminal arginine vasopressin treatment and activate their luminal aquaporin 2. The ability to reabsorb water was restored by treating these unciliated renal epithelial cells with forskolin, a receptor-independent adenylate cyclase activator, showing that the intracellular machinery for water absorption was present but not activated. These findings suggest that the luminal receptor located on the primary cilium may be important for efficient transepithelial water absorption.

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