Defects In Primary Cilia Research Articles

Craniofacial ciliopathies: A new classification for craniofacial disorders.

Craniofacial anomalies are some of the most variable and common defects affecting the population. Herein, we examine a group of craniofacial disorders that are the result of defects in primary cilia; ubiquitous, microtubule-based organelles that transduce molecular signals and facilitate the interactions between the cell and its environment. Based on the frequent appearance of craniofacial phenotypes in diseases born from defective primary cilia (ciliopathies) we propose a new class of craniofacial disorders referred to as craniofacial ciliopathies. We explore the most frequent phenotypes associated with ciliopathic conditions and the ciliary gene mutations responsible for craniofacial defects. Finally, we propose that some non-classified disorders may now be classified as craniofacial ciliopathies.

Lights on for aminopeptidases in cystic kidney disease

While erudite cell biologists have for many decades described singular immotile appendages known as primary cilia to be present on most cells in our bodies, cilial function(s) long remained an enigma. Driven largely by an ever increasing number of discoveries of genetic defects in primary cilia during the past decade, cilia were catapulted from a long lasting existence in obscurity into the bright spotlight in cell biology and medicine. The study by O'Toole et al. in this issue of the JCI adds a novel "enzymatic" facet to the rapidly growing information about these little cellular tails, by demonstrating that defects in the XPNPEP3 gene, which encodes mitochondrial and cytosolic splice variants of X-prolyl aminopeptidase 3, can cause nephronophthisis-like ciliopathy. Future studies are in order now to elucidate the cystogenic pathways affected by disrupted enzymatic function of XPNPEP3 in cilia-related cystogenic diseases.

New roles for an ancient organelle: ciliary defects cause cystic kidney disease and blindness

The discovery that the mutation in the Tg737orpk mouse model of polycystic kidney disease affected assembly of cilia in kidney tubules has fueled new interest in primary cilia. These organelles were first described in 1898 and over the next one hundred years, microscopists documented the presence of these organelles on nearly every cell type in vertebrate organisms. Prior to the work on the Tg737orpk mouse, the function of the primary cilium was controversial with many cell biologists believing them to be vestigial organelles with no function in the modern organism. The idea that primary cilia are vestigial has been soundly disproved as severe defects in primary cilia cause embryonic lethality and milder defects lead to a syndrome of phenotypes including cystic kidney disease, liver cysts and fibrosis, polydactyly, and defects of the mammary gland, heart, pancreas, brain and eye. It is now widely believed that these organelles are cellular antennae that monitor the extracellular environment and feed this information back to the cell to control many aspects of cell physiology including proliferation and differentiation. Recent evidence indicates that cilia are key players in hedgehog and Wnt signaling and so may integrate extracellular signals in the control of these and other signaling pathways.

TheTalpid3gene (KIAA0586) encodes a centrosomal protein that is essential for primary cilia formation

The chicken talpid(3) mutant, with polydactyly and defects in other embryonic regions that depend on hedgehog (Hh) signalling (e.g. the neural tube), has a mutation in KIAA0568. Similar phenotypes are seen in mice and in human syndromes with mutations in genes that encode centrosomal or intraflagella transport proteins. Such mutations lead to defects in primary cilia, sites where Hh signalling occurs. Here, we show that cells of talpid(3) mutant embryos lack primary cilia and that primary cilia can be rescued with constructs encoding Talpid3. talpid(3) mutant embryos also develop polycystic kidneys, consistent with widespread failure of ciliogenesis. Ultrastructural studies of talpid(3) mutant neural tube show that basal bodies mature but fail to dock with the apical cell membrane, are misorientated and almost completely lack ciliary axonemes. We also detected marked changes in actin organisation in talpid(3) mutant cells, which may explain misorientation of basal bodies. KIAA0586 was identified in the human centrosomal proteome and, using an antibody against chicken Talpid3, we detected Talpid3 in the centrosome of wild-type chicken cells but not in mutant cells. Cloning and bioinformatic analysis of the Talpid3 homolog from the sea anemone Nematostella vectensis identified a highly conserved region in the Talpid3 protein, including a predicted coiled-coil domain. We show that this region is required to rescue primary cilia formation and neural tube patterning in talpid(3) mutant embryos, and is sufficient for centrosomal localisation. Thus, Talpid3 is one of a growing number of centrosomal proteins that affect both ciliogenesis and Hh signalling.

A Mouse Model for Meckel Syndrome Type 3

Meckel-Gruber syndrome type 3 (MKS3; OMIM 607361) is a severe autosomal recessive disorder characterized by bilateral polycystic kidney disease. Other malformations associated with MKS3 include cystic changes in the liver, polydactyly, and brain abnormalities (occipital encephalocele, hydrocephalus, and Dandy Walker-type cerebellar anomalies). The disorder is hypothesized to be caused by defects in primary cilia. In humans, the underlying mutated gene, TMEM67, encodes transmembrane protein 67, also called meckelin (OMIM 609884), which is an integral protein of the renal epithelial cell and membrane of the primary cilium. Here, we describe a spontaneous deletion of the mouse ortholog, Tmem67, which results in polycystic kidney disease and death by 3 wk after birth. Hydrocephalus also occurs in some mutants. We verified the mutated gene by transgenic rescue and characterized the phenotype with microcomputed tomography, histology, scanning electron microscopy, and immunohistochemistry. This mutant provides a mouse model for MKS3 and adds to the growing set of mammalian models essential for studying the role of the primary cilium in kidney function.

Phosphatase inhibitor 2 promotes acetylation of tubulin in the primary cilium of human retinal epithelial cells.

BackgroundPrimary cilia are flagella-like projections from the centriole of mammalian cells that have a key role in cell signaling. Human diseases are linked to defects in primary cilia. Microtubules make up the axoneme of cilia and are selectively acetylated and this is thought to contribute to the stability of the structure. However, mechanisms to regulate tubulin acetylation in cilia are poorly understood.ResultsEndogenous phosphatase inhibitor-2 (I-2) was found concentrated in cilia of human epithelial cells, and was localized to cilia early in the process of formation, prior to the full acetylation of microtubules. Knockdown of I-2 by siRNA significantly reduced the acetylation of microtubules in cilia, without a net decrease in whole cell tubulin acetylation. There was a reduction in the percentage of I-2 knockdown cells with a primary cilium, but no apparent alteration in the cilium length, suggesting no change in microtubule-based transport processes. Inhibition of either histone deacetylases with trichostatin A, or protein phosphatase-1 with calyculin A in I-2 knockdown cells partially rescued the acetylation of microtubules in cilia and the percentage of cells with a primary cilium.ConclusionThe regulatory protein I-2 localizes to the primary cilium where it affects both Ser/Thr phosphorylation and is required for full tubulin acetylation. Rescue of tubulin acetylation in I-2 knockdown cells by different chemical inhibitors shows that deacetylases and phosphatases are functionally interconnected to regulate microtubules. As a multifunctional protein, I-2 may link cell cycle progression to structure and stability of the primary cilium.

Deletion of IFT20 in the mouse kidney causes misorientation of the mitotic spindle and cystic kidney disease

Primary cilia project from the surface of most vertebrate cells and are thought to be sensory organelles. Defects in primary cilia lead to cystic kidney disease, although the ciliary mechanisms that promote and maintain normal renal function remain incompletely understood. In this work, we generated a floxed allele of the ciliary assembly gene Ift20. Deleting this gene specifically in kidney collecting duct cells prevents cilia formation and promotes rapid postnatal cystic expansion of the kidney. Dividing collecting duct cells in early stages of cyst formation fail to properly orient their mitotic spindles along the tubule, whereas nondividing cells improperly position their centrosomes. At later stages, cells lacking cilia have increased canonical Wnt signaling and increased rates of proliferation. Thus, IFT20 functions to couple extracellular events to cell proliferation and differentiation.

Restoration of renal function in zebrafish models of ciliopathies

The ciliopathies are a class of rare human genetic disease whose aetioligies lie in defective primary cilia. Typical ciliopathies include Bardet–Biedl syndrome (BBS), nephronophthisis (NPHP), Jeune, Joubert, oro-facial-digital (OFD1) and Meckel (MKS) syndromes. All ciliopathies have the common denominator of renal disease, often including tubular cysts. In this study, we have modelled a range of ciliopathies in zebrafish and shown in all cases that knocking down these genes causes cystic lesions in the kidney. We have identified two drugs, rapamycin and roscovitine, which ameliorate the renal phenotype, both morphologically and functionally. This is the first study in which zebrafish has been used to identify potential therapeutic modalities for ciliopathic renal disease, and the results pave the way for further investigations in mammalian models.

A genetic screen in zebrafish identifies cilia genes as a principal cause of cystic kidney.

Polycystic kidney disease (PKD) is a common human genetic illness. It is characterized by the formation of multiple kidney cysts that are thought to result from over-proliferation of epithelial cells. Zebrafish larvae can also develop kidney cysts. In an insertional mutagenesis screen in zebrafish, we identified 12 genes that can cause cysts in the glomerular-tubular region when mutated and we cloned 10 of these genes. Two of these genes, vhnf1 (tcf2) and pkd2, are already associated with human cystic kidney diseases. Recently, defects in primary cilia have been linked to PKD. Strikingly, three out of the 10 genes cloned in this screen are homologues of Chlamydomonas genes that encode components of intraflagellar transport (IFT) particles involved in cilia formation. Mutation in a fourth blocks ciliary assembly by an unknown mechanism. These results provide compelling support for the connection between cilia and cystogenesis. Our results also suggest that lesions in genes involved in cilia formation and function are the predominant cause of cystic kidney disease, and that the genes identified here are excellent candidates for novel human PKD genes.