In vivo imaging of germinal centres reveals a dynamic open structure

Germinal centres are specialized structures wherein B lymphocytes undergo clonal expansion, class switch recombination, antibody gene diversification and affinity maturation. Three to four antigen-specific B cells colonize a follicle to establish a germinal centre and become rapidly dividing germinal-centre centroblasts that give rise to dark zones. Centroblasts produce non-proliferating centrocytes that are thought to migrate to the light zone of the germinal centre, which is rich in antigen-trapping follicular dendritic cells and CD4+ T cells. It has been proposed that centrocytes are selected in the light zone on the basis of their ability to bind cognate antigen. However, there have been no studies of germinal-centre dynamics or the migratory behaviour of germinal-centre cells in vivo. Here we report the direct visualization of B cells in lymph node germinal centres by two-photon laser-scanning microscopy in mice. Nearly all antigen-specific B cells participating in a germinal-centre reaction were motile and physically restricted to the germinal centre but migrated bi-directionally between dark and light zones. Notably, follicular B cells were frequent visitors to the germinal-centre compartment, suggesting that all B cells scan antigen trapped in germinal centres. Consistent with this observation, we found that high-affinity antigen-specific B cells can be recruited to an ongoing germinal-centre reaction. We conclude that the open structure of germinal centres enhances competition and ensures that rare high-affinity B cells can participate in antibody responses.     

The genome of Theobroma cacao

We sequenced and assembled the draft genome of Theobroma cacao, an economically important tropical-fruit tree crop that is the source of chocolate. This assembly corresponds to 76% of the estimated genome size and contains almost all previously described genes, with 82% of these genes anchored on the 10 T. cacao chromosomes. Analysis of this sequence information highlighted specific expansion of some gene families during evolution, for example, flavonoid-related genes. It also provides a major source of candidate genes for T. cacao improvement. Based on the inferred paleohistory of the T. cacao genome, we propose an evolutionary scenario whereby the ten T. cacao chromosomes were shaped from an ancestor through eleven chromosome fusions.     

Modelling a ciliopathy: Ahi1 knockdown in model systems reveals an essential role in brain,retinal, and renal development

Joubert syndrome and related diseases (JSRD) are cerebello-oculo-renal syndromes with phenotypes including cerebellar hypoplasia, retinal dystrophy, and nephronophthisis (a cystic kidney disease). Mutations in AHI1 are the most common genetic cause of JSRD, with developmental hindbrain anomalies and retinal degeneration being prominent features. We demonstrate that Ahi1, a WD40 domain-containing protein, is highly conserved throughout evolution and its expression associates with ciliated organisms. In zebrafish ahi1 morphants, the phenotypic spectrum of JSRD is modeled, with embryos showing brain, eye, and ear abnormalities, together with renal cysts and cloacal dilatation. Following ahi1 knockdown in zebrafish, we demonstrate loss of cilia at Kupffer’s vesicle and subsequently defects in cardiac left–right asymmetry. Finally, using siRNA in renal epithelial cells we demonstrate a role for Ahi1 in both ciliogenesis and cell–cell junction formation. These data support a role for Ahi1 in epithelial cell organization and ciliary formation and explain the ciliopathy phenotype of AHI1 mutations in man.     

Short‐term forecasts of euro area real GDP growth: an assessment of real‐time performance based on vintage data

Forecasters commonly predict real gross domestic product growth from monthly indicators such as industrial production, retail sales and surveys, and therefore require an assessment of the reliability of such tools. While forecast errors related to model specification and unavailability of data in real time have been assessed, the impact of data revisions on forecast accuracy has seldom been evaluated, especially for the euro area. This paper proposes to evaluate the contributions of these three sources of forecast error using a set of data vintages for the euro area. The results show that gains in accuracy of forecasts achieved by using monthly data on actual activity rather than surveys or financial indicators are offset by the fact that the former set of monthly data is harder to forecast and less timely than the latter set. These results provide a benchmark which future research may build on as more vintage datasets become available. Copyright © 2008 John Wiley & Sons, Ltd.     

Membrane traffic in the secretory pathway

Cholesterol, certain lipids, membrane-bound and soluble proteins, as well as viruses that are synthesized in the endoplasmic reticulum (ER), reach the plasma membrane (PM) via non-classical pathway(s) that remain poorly understood. Typical for this transport is (i) its insensitivity to brefeldin A (BFA), which dissociates selected coat complexes from membranes, resulting in the disassembly of the Golgi apparatus; (ii) its rapid kinetics as compared to the classical secretory pathway; and (iii) its role in the trafficking of lipid raft components. Based on results showing that the intermediate compartment (IC) at the ER-Golgi boundary constitutes a stable tubular network that maintains its dynamics in the presence of BFA, we propose that two bidirectional Golgi-bypass pathways to the PM exist, a direct route from early IC elements, and another, reminiscent of the yeast secretory pathway, from late IC elements via the endosomal system. These pathways have implications for the organization of the secretory processes in different cell types.     

Thromboxane synthase mutations in an increased bone density disorder (Ghosal syndrome)

Studying consanguineous families with Ghosal hematodiaphyseal dysplasia syndrome (GHDD), a disorder of increased bone density, we identified mutations in TBXAS1, which encodes thromboxane synthase (TXAS). TXAS, an enzyme of the arachidonic acid cascade, produces thromboxane A(2) (TXA(2)). Platelets from subjects with GHDD showed a specific deficit in arachidonic acid-produced aggregation. We also found that TXAS and TXA(2) modulated expression of TNFSF11 and TNFRSF11B (encoding RANKL and osteoprotegerin (OPG), respectively) in primary cultured osteoblasts.     

Handling Missing Values with Regularized Iterative Multiple Correspondence Analysis

A common approach to deal with missing values in multivariate exploratory data analysis consists in minimizing the loss function over all non-missing elements, which can be achieved by EM-type algorithms where an iterative imputation of the missing values is performed during the estimation of the axes and components. This paper proposes such an algorithm, named iterative multiple correspondence analysis, to handle missing values in multiple correspondence analysis (MCA). The algorithm, based on an iterative PCA algorithm, is described and its properties are studied. We point out the overfitting problem and propose a regularized version of the algorithm to overcome this major issue. Finally, performances of the regularized iterative MCA algorithm (implemented in the R-package named missMDA) are assessed from both simulations and a real dataset. Results are promising with respect to other methods such as the missing-data passive modified margin method, an adaptation of the missing passive method used in Gifi’s Homogeneity analysis framework.     

Dystrophin deficiency leads to disturbance of LAMP1-vesicle-associated protein secretion

Duchenne muscular dystrophy results from loss of the protein dystrophin, which links the intracellular cytoskeletal network with the extracellular matrix, but deficiency in this function does not fully explain the onset or progression of the disease. While some intracellular events involved in the degeneration of dystrophin-deficient muscle fibers have been well characterized, changes in their secretory profile are undescribed. To analyze the secretome profile of mdx myotubes independently of myonecrosis, we labeled the proteins of mdx and wild-type myotubes with stable isotope-labeled amino acids (SILAC), finding marked enrichment of vesicular markers in the mdx secretome. These included the lysosomal-associated membrane protein, LAMP1, that co-localized in vesicles with an over-secreted cytoskeletal protein, myosin light chain 1. These LAMP1/MLC1-3-positive vesicles accumulated in the cytosol of mdx myotubes and were secreted into the culture medium in a range of abnormal densities. Restitution of dystrophin expression, by exon skipping, to some 30 % of the control value, partially normalized the secretome profile and the excess LAMP1 accumulation. Together, our results suggest that a lack of dystrophin leads to a general dysregulation of vesicle trafficking. We hypothesize that disturbance of the export of proteins through vesicles occurs before, and then concurrently with, the myonecrotic cascade and contributes chronically to the pathophysiology of DMD, thereby presenting us with a range of new potential therapeutic targets.