Primary cilia project in a single copy from the surface of most vertebrate cell types; they detect and transmit extracellular cues to regulate diverse cellular processes during development and to maintain tissue homeostasis. The sensory capacity of primary cilia relies on the coordinated trafficking and temporal localization of specific receptors and associated signal transduction modules in the cilium. The canonical hedgehog (HH) pathway, for example, is a bona fide ciliary signalling system that regulates cell fate and self-renewal in development and tissue homeostasis. Specific receptors and associated signal transduction proteins can also localize to primary cilia in a cell type-dependent manner; available evidence suggests that the ciliary constellation of these proteins can temporally change to allow the cell to adapt to specific developmental and homeostatic cues. Consistent with important roles for primary cilia in signalling, mutations that lead to their dysfunction underlie a pleiotropic group of diseases and syndromic disorders termed ciliopathies, which affect many different tissues and organs of the body. In this review we highlight central mechanisms by which primary cilia coordinate HH, G-protein-coupled receptor, WNT, receptor tyrosine kinase and TGFβ/BMP signalling, and illustrate how defects in the balanced output of ciliary signalling events are coupled to developmental disorders and disease progression. Opening section The primary cilium is a microtubule-based, non-motile organelle that extends as a solitary unit from the basal body (derived from the centrosomal mother centriole of most cell types

Primary cilia are ubiquitous cellular appendages that provide important yet not well understood sensory and signaling functions. Ciliary dysfunction underlies numerous human genetic disorders. However, the precise defects in cilia function and the basis of disease pathophysiology remain unclear. Here, we report that the proteins disrupted in the human ciliary disorder Bardet-Biedl syndrome (BBS) are required for the localization of G proteincoupled receptors to primary cilia on central neurons. We demonstrate a lack of ciliary localization of somatostatin receptor type 3 (Sstr3) and melanin-concentrating hormone receptor 1 (Mchr1) in neurons from mice lacking the Bbs2 or Bbs4 gene. Because Mchr1 is involved in the regulation of feeding behavior and BBS is associated with hyperphagia-induced obesity, our results suggest that altered signaling caused by mislocalization of ciliary signaling proteins underlies the BBS phenotypes. Our results also provide a potential molecular mechanism to link cilia defects with obesity.melanin-concentrating hormone receptor 1 ͉ neuronal cilia ͉ obesity ͉ somatostatin receptor 3 ͉ type III adenylyl cyclase

Bardet-Biedl syndrome (BBS) is a heterogeneous, pleiotropic human disorder characterized by obesity, retinopathy, polydactyly, renal and cardiac malformations, learning disabilities, hypogenitalism, and an increased incidence of diabetes and hypertension. No information is available regarding the specific function of BBS2. We show that mice lacking Bbs2 gene expression have major components of the human phenotype, including obesity and retinopathy. In addition, these mice have phenotypes associated with cilia dysfunction, including retinopathy, renal cysts, male infertility, and a deficit in olfaction. With the exception of male infertility, these phenotypes are not caused by a complete absence of cilia. We demonstrate that BBS2 retinopathy involves normal retina development followed by apoptotic death of photoreceptors, the primary ciliated cells of the retina. Photoreceptor cell death is preceded by mislocalization of rhodopsin, indicating a defect in transport. We also demonstrate that Bbs2 ؊/؊ mice and a second BBS mouse model, Bbs4 ؊/؊ , have a defect in social function. The evaluation of Bbs2 ؊/؊ mice indicates additional phenotypes that should be evaluated in human patients, including deficits in social interaction and infertility.Bardet-Biedl syndrome ͉ mouse model ͉ obesity

Primary cilia are sensory organelles present on most mammalian cells. The functions of cilia are defined by the signaling proteins localized to the ciliary membrane. Certain G protein-coupled receptors (GPCRs), including somatostatin receptor 3 (Sstr3) and serotonin receptor 6 (Htr6), localize to cilia. As Sstr3 and Htr6 are the only somatostatin and serotonin receptor subtypes that localize to cilia, we hypothesized they contain ciliary localization sequences. To test this hypothesis we expressed chimeric receptors containing fragments of Sstr3 and Htr6 in the nonciliary receptors Sstr5 and Htr7, respectively, in ciliated cells. We found the third intracellular loop of Sstr3 or Htr6 is sufficient for ciliary localization. Comparison of these loops revealed a loose consensus sequence. To determine whether this consensus sequence predicts ciliary localization of other GPCRs, we compared it with the third intracellular loop of all human GPCRs. We identified the consensus sequence in melanin-concentrating hormone receptor 1 (Mchr1) and confirmed Mchr1 localizes to primary cilia in vitro and in vivo. Thus, we have identified a putative GPCR ciliary localization sequence and used this sequence to identify a novel ciliary GPCR. As Mchr1 mediates feeding behavior and metabolism, our results implicate ciliary signaling in the regulation of body weight. INTRODUCTIONPrimary cilia are appendages that project from almost all human cell types (Wheatley et al., 1996). It is generally accepted that primary cilia serve important specialized signaling functions (Pazour and Witman, 2003;Marshall and Nonaka, 2006;Singla and Reiter, 2006). In the eye, photoreceptors, which are modified primary cilia, sense and respond to light. In the nose, specialized olfactory cilia detect odors and initiate signaling cascades in olfactory neurons. In the kidney, it is proposed that bending of cilia on epithelial cells by fluid flow triggers an increase in intracellular calcium mediated by an ion channel located on the cilium (Praetorius and Spring, 2001;Nauli et al., 2003). In each case, the function of the cilium is defined by the signaling proteins that are enriched in the ciliary membrane (i.e., light receptors, odorant receptors, and mechanoreceptors). Importantly, disruption of the signaling mediated by these receptors can cause disease and altered development (Davenport and Yoder, 2005;Hildebrandt and Otto, 2005;Pan et al., 2005;Bisgrove and Yost, 2006). Yet, the specific signaling proteins that localize to the vast majority of cilia in the mammalian body are unknown. Thus, the functions of primary cilia on most cell types in the body are unknown.Neuronal primary cilia are abundant throughout the rodent brain (Bishop et al., 2007). The functional importance of these cilia is suggested by the fact that several human ciliary disorders, including Bardet-Biedl syndrome (BBS), Joubert syndrome (JS), and Meckel syndrome (MKS), have prominent functional and structural CNS phenotypes (Badano et al., 2006). Although the specific functions of neuron...

The functions of the proteins encoded by the Bardet-Biedl syndrome (BBS) genes are unknown. Mutations in these genes lead to the pleiotropic human disorder BBS, which is characterized by obesity, retinopathy, polydactyly, renal and cardiac malformations, learning disabilities, and hypogenitalism. Secondary features include diabetes mellitus and hypertension. Recently, it has been suggested that the BBS phenotypes are the result of a lack of cilia formation or function. In this study, we show that mice lacking the Bbs4 protein have major components of the human phenotype, including obesity and retinal degeneration. We show that Bbs4-null mice develop both motile and primary cilia, demonstrating that Bbs4 is not required for global cilia formation. Interestingly, male Bbs4-null mice do not form spermatozoa flagella, and BBS4 retinopathy involves apoptotic death of photoreceptors, the primary ciliated cells of the retina. These mutation data demonstrate a connection between the function of a BBS protein and cilia. To further evaluate an association between cilia and BBS, we performed homology comparisons of BBS proteins in model organisms and find that BBS proteins are specifically conserved in ciliated organisms.B ardet-Biedl syndrome (BBS) is a genetically heterogeneous disorder with linkage to eight loci. Six BBS genes (BBS1, BBS2, BBS4, BBS6, BBS7, and BBS8) have been identified (1-7). With the exception of BBS6, which has similarity to type II chaperonins (8), BBS proteins do not show extensive homology with proteins of known function. BBS4 and BBS8 contain tetratricopeptide repeat domains, and a region of BBS8 shows similarity to the prokaryotic pilF domain. Based on the observations that BBS8 localizes to the basal body of ciliated cells and expression of the Caenorhabditis elegans orthologues of several BBS proteins are limited to ciliated cells, it has been hypothesized that BBS is the result of a defect in cilia assembly or function (7). We previously used positional cloning to identify the human BBS4 gene (3). To help elucidate the function of the BBS proteins, we have now targeted the Bbs4 gene in mice. Bbs4 Ϫ/Ϫ mice recapitulate aspects of the human phenotype: they become obese, fail to reproduce, and display retinal degeneration. We show the presence of both motile and primary cilia in these mice, demonstrating that Bbs4 deficiency does not prevent global ciliogenesis. Interestingly, male knockout mice have a complete lack of flagella, demonstrating that the Bbs4 protein is necessary for flagella formation during spermatogenesis. Furthermore, Bbs4-null mice undergo retinal degeneration due principally to photoreceptor cell loss associated with outer segment attenuation, suggesting a role for BBS proteins in maintenance of sensory cilia. These data demonstrate a role for BBS proteins in facets of cilia function. Materials and MethodsConstruction of the Bbs4 Gene Targeting Vector. BLAST analysis of the Celera mouse fragment database with the human BBS4 cDNA sequence identified sequences corresponding to...

Solitary primary cilia project from nearly every cell type in the human body. These organelles are considered to have important sensory and signaling functions. Although primary cilia have been detected throughout the mammalian brain, their functions are unknown. The study of primary cilia in the brain is constrained by the scarcity of specific markers for these organelles. We previously demonstrated that type III adenylyl cyclase (ACIII) is a marker for primary cilia on neonatal hippocampal neurons in vivo and in vitro. We further showed that ACIII localizes to cilia on cultured glial cells. Here, we report that ACIII is a marker for primary cilia throughout many regions of the adult mouse brain. Furthermore, we report that ACIII localizes to primary cilia on choroid plexus cells and some astrocytes in the brain, which to our knowledge is the first report of a marker for visualizing cilia on glia in vivo. Overall, our data indicate that ACIII is a prominent marker of primary cilia in the brain and will provide an important tool to facilitate further investigations into the functions of these organelles.

Primary cilia are nearly ubiquitous cellular appendages that provide important sensory and signaling functions. Ciliary dysfunction underlies numerous human diseases, collectively termed ciliopathies. Primary cilia have distinct functions on different cell types and these functions are defined by the signaling proteins that localize to the ciliary membrane. Neurons throughout the mammalian brain possess primary cilia upon which certain G protein-coupled receptors localize. Yet, the precise signaling proteins present on the vast majority of neuronal cilia are unknown. Here, we report that dopamine receptor 1 (D1) localizes to cilia on mouse central neurons, thereby implicating neuronal cilia in dopamine signaling. Interestingly, ciliary localization of D1 is dynamic and the receptor rapidly translocates to and from cilia in response to environmental cues. Notably, the translocation of D1 from cilia requires proteins mutated in the ciliopathy Bardet-Biedl syndrome (BBS) and we find that one of the BBS proteins, Bbs5, specifically interacts with D1.

Bardet-Biedl syndrome (BBS, OMIM 209900) is a genetic disorder with the primary features of obesity, pigmentary retinopathy, polydactyly, renal malformations, mental retardation and hypogenitalism. Individuals with BBS are also at increased risk for diabetes mellitus, hypertension and congenital heart disease. What was once thought to be a homogeneous autosomal recessive disorder is now known to map to at least six loci: 11q13 (BBS1), 16q21 (BBS2), 3p13 p12 (BBS3), 15q22.3 q23 (BBS4), 2q31 (BBS5) and 20p12 (BBS6). There has been considerable interest in identifying the genes that underlie BBS, because some components of the phenotype are common. Cases of BBS mapping ro BBS6 are caused by mutations in MKKS; mutations in this gene also cause McKusick-Kaufman syndrome (hydrometrocolpos, post-axial polydactyly and congenital heart defects). In addition, we recently used positional cloning to identify the genes underlying BBS2 (ref. 16) and BBS4 (ref. 17). The BBS6 protein has similarity to a Thermoplasma acidophilum chaperonin, whereas BBS2 and BBS4 have no significant similarity to chaperonins. It has recently been suggested that three mutated alleles (two at one locus, and a third at a second locus) may be required for manifestation of BBS (triallelic inheritance). Here we report the identification of the gene BBS1 and show that a missense mutation of this gene is a frequent cause of BBS. In addition, we provide data showing that this common mutation is not involved in triallelic inheritance.