Mutations in Outer Dynein Arm Heavy Chain DNAH9 Cause Motile Cilia Defects and Situs Inversus

International audienceMotile cilia move body fluids and gametes and the beating of cilia lining the airway epithelial surfaces ensures that they are kept clear and protected from inhaled pathogens and consequent respiratory infections. Dynein motor proteins provide mechanical force for cilia beating. Dynein mutations are a common cause of primary ciliary dyskinesia (PCD), an inherited condition characterized by deficient mucociliary clearance and chronic respiratory disease coupled with laterality disturbances and subfertility. Using next-generation sequencing, we detected mutations in the ciliary outer dynein arm (ODA) heavy chain gene DNAH9 in individuals from PCD clinics with situs inversus and in one case male infertility. DNAH9 and its partner heavy chain DNAH5 localize to type 2 ODAs of the distal cilium and in DNAH9-mutated nasal respiratory epithelial cilia we found a loss of DNAH9/DNAH5-containing type 2 ODAs that was restricted to the distal cilia region. This confers a reduced beating frequency with a subtle beating pattern defect affecting the motility of the distal cilia portion. 3D electron tomography ultrastructural studies confirmed regional loss of ODAs from the distal cilium, manifesting as either loss of whole ODA or partial loss of ODA volume. Paramecium DNAH9 knockdown confirms an evolutionarily conserved function for DNAH9 in cilia motility and ODA stability. We find that DNAH9 is widely expressed in the airways, despite DNAH9 mutations appearing to confer symptoms restricted to the upper respiratory tract. In summary, DNAH9 mutations reduce cilia function but some respiratory mucociliary clearance potential may be retained, widening the PCD disease spectrum

ParameciumParamecium, a model to study ciliary beating and ciliogenesis: insights from cutting-edge approaches

International audienceCilia are ubiquitous and highly conserved extensions that endow the cell with motility and sensory functions. They were present in the first eukaryotes and conserved throughout evolution ( Carvalho-Santos et al., 2011 ). Paramecium has around 4,000 motile cilia on its surface arranged in longitudinal rows, beating in waves to ensure movement and feeding. As with cilia in other model organisms, direction and speed of Paramecium ciliary beating is under bioelectric control of ciliary ion channels. In multiciliated cells of metazoans as well as paramecia, the cilia become physically entrained to beat in metachronal waves. This ciliated organism, Paramecium , is an attractive model for multidisciplinary approaches to dissect the location, structure and function of ciliary ion channels and other proteins involved in ciliary beating. Swimming behavior also can be a read-out of the role of cilia in sensory signal transduction. A cilium emanates from a BB, structurally equivalent to the centriole anchored at the cell surface, and elongates an axoneme composed of microtubule doublets enclosed in a ciliary membrane contiguous with the plasma membrane. The connection between the BB and the axoneme constitutes the transition zone, which serves as a diffusion barrier between the intracellular space and the cilium, defining the ciliary compartment. Human pathologies affecting cilia structure or function, are called ciliopathies, which are caused by gene mutations. For that reason, the molecular mechanisms and structural aspects of cilia assembly and function are actively studied using a variety of model systems, ranging from unicellular organisms to metazoa. In this review, we will highlight the use of Paramecium as a model to decipher ciliary beating mechanisms as well as high resolution insights into BB structure and anchoring. We will show that study of cilia in Paramecium promotes our understanding of cilia formation and function. In addition, we demonstrate that Paramecium could be a useful tool to validate candidate genes for ciliopathies

The centrosome is an actin-organizing centre.

International audienceMicrotubules and actin filaments are the two main cytoskeleton networks supporting intracellular architecture and cell polarity. The centrosome nucleates and anchors microtubules and is therefore considered to be the main microtubule-organizing centre. However, recurring, yet unexplained, observations have pointed towards a connection between the centrosome and actin filaments. Here we have used isolated centrosomes to demonstrate that the centrosome can directly promote actin-filament assembly. A cloud of centrosome-associated actin filaments could be identified in living cells as well. Actin-filament nucleation at the centrosome was mediated by the nucleation-promoting factor WASH in combination with the Arp2/3 complex. Pericentriolar material 1 (PCM1) seemed to modulate the centrosomal actin network by regulating Arp2/3 complex and WASH recruitment to the centrosome. Hence, our results reveal an additional facet of the centrosome as an intracellular organizer and provide mechanistic insights into how the centrosome can function as an actin-filament-organizing centre