Spectrum of Clinical Diseases Caused By Disorders

of Primary Cilia
Stephanie M. Ware1, Meral Gunay-Aygun2, and Friedhelm Hildebrandt3
1
Department of Pediatrics, Cincinnati Children’s Hospital Medical Center, Cincinnati, Ohio; 2National Human Genome Research Institute, National
Institutes of Health, Bethesda, Maryland; and 3Howard Hughes Medical Institute and Departments of Pediatrics and of Human Genetics, University
of Michigan, Ann Arbor, Michigan

The ciliopathies are a category of diseases caused by disruption of function are better understood, further delineation of subtypes
the physiological functions of cilia. Ciliary dysfunction results in a may occur. For example, the rod and cone cells of the retina
broad range of phenotypes, including renal, hepatic, and pancreatic possess a very unique type of primary cilium (connecting cilium)
cyst formation; situs abnormalities; retinal degeneration; anosmia; equipped with a specialized expanded tip that forms the outer
cerebellar or other brain anomalies; postaxial polydactyly; bronchi- segment of these photoreceptor cells (2). The node or organizer,
ectasis; and infertility. The specific clinical features are dictated by a transient embryonic structure required for left-right patterning,
the subtype, structure, distribution, and function of the affected also possesses a unique type of primary cilium that lacks a central
cilia. This review highlights the clinical variability caused by dysfunc- pair of microtubules but is nevertheless motile. Dysfunction of
tion of motile and nonmotile primary cilia and emphasizes the
cilia at the node can result in situs inversus or heterotaxy, a phe-
genetic heterogeneity and phenotypic overlap that are character-
notype that is characteristic of PCD and primary, nonmotile cil-
istics of these disorders. There is a need for additional research to
iopathies. Understanding the biological function of motile and
understand the shared and unique functions of motile and non-
motile cilia and the pathophysiology resulting from mutations in nonmotile cilia will provide insight into the underlying patho-
cilia, basal bodies, or centrosomes. Increased understanding of physiology of ciliary disorders and the wide array of clinical phe-
ciliary biology will improve the diagnosis and management of pri- notypes resulting from their disruption. The number of diseases
mary ciliary dyskinesia, syndromic ciliopathies, and cilia-related known to be caused by ciliary pathology has increased dramati-
cystic diseases. cally in the last decade (summarized in Table 2). The diagnosis
and management of patients with ciliary disorders will benefit
Keywords: primary cilia; ciliopathy; heterotaxy; nephronophthisis; cyst from elucidating the shared and unique functions directed by
motile and nonmotile cilia in a tissue- and developmental-stage–
Ciliopathies are phenotypically and genetically heterogeneous specific context.
disorders that share ciliary dysfunction as a common patholog-
ical mechanism. Motile cilia are primarily found on epi-
thelial cells that line the respiratory tract, brain ventricles, and FUNCTIONS OF PRIMARY CILIA
oviducts. These cilia have nine microtubule doublets surrounding Primary cilia, both nonmotile cilia and motile cilia at the node,
a central pair, a “912” arrangement that is visible by electron function as sensory organelles and coordinators of signal transduc-
microscopy of cross-sectional views. Absence or dysfunction of tion pathways such as hedgehog (Hh), wingless (WNT), platelet-
motile cilia causes primary ciliary dyskinesia (PCD). In con- derived growth factor-a, and fibroblast growth factor pathways
trast, primary cilia lack the central microtubule doublet and (3, 4). In addition, primary cilia are important for the regulation
have a “910” arrangement of microtubules. Primary cilia ex- of intracellular calcium. Nonmotile cilia are sensory antennas of
tend from the surface of almost all cell types in the human body, the cell, and intact nonmotile cilia function is required for main-
including epithelial cells such as those lining the kidney tubules tenance of the balance between the canonical and noncanonical
and bile ducts as well as nonepithelial cells such as chondrocytes (planar cell polarity) parts of the WNT pathway. For example,
and neurons (1). Primary cilia are present as monocilia and exist disruption of planar cell polarity contributes to the pathogenesis
as a single extension per cell. Initially, cilia were described as of polycystic kidney disease (PKD) by resulting in abnormalities
motile or nonmotile, with the latter group encompassing pri- in multicellular activities that maintain tubular diameter, such
mary cilia or sensory cilia (Table 1). However, the discovery as mitotic spindle orientation and convergence and extension
of motile primary cilia at the embryonic node and the identifi- movements (5). Nonmotile cilia also sense mechanical stimuli
cation of sensory functions for motile cilia has rendered these such as fluid flow in the lumen of tubular structures (kidney
historic designations imperfect. Although these broad classifica- tubules and bile ducts) and convert these to alterations in calcium
tions have utility, as the complexities of cilia structure and fluxes within the cell (1). Proteins defective in human PKD are
required for this mechanosensory function of nonmotile cilia.
Hence, during embryologic development, primary cilia regulate
fundamental processes, such as determination of asymmetry of
(Received in original form March 15, 2011; accepted in final form April 25, 2011) internal organs and neural tube and limb patterning. Nonmotile
The conference was supported by a grant from the National Institutes of Health cilia are also required for the maintenance of differentiated tis-
(R13HL105073–01) and by the Primary Ciliary Dyskinesia Foundation. The sues, such as retina and renal tubules. In addition, they play a role
authors’ research is supported by the National Institutes of Health grants in the pathogenesis of certain types of obesity, probably through
HL088639 (S.M.W.) and DK068305 and DK064614 (F.H.) and Burroughs- their role in appetite control and adipocyte differentiation (6).
Wellcome Fund #1008496 (S.M.W.).
Correspondence and requests for reprints should be addressed to Stephanie
M. Ware, M.D., Ph.D., Cincinnati Children’s Hospital Medical Center, 240 Albert HETEROTAXY SPECTRUM DISORDERS
Sabin Way, MLC 7020, Cincinnati, OH 45244. E-mail: stephanie.ware@cchmc.
org
Heterotaxy, or situs ambiguus, is an abnormal arrangement of tho-
Proc Am Thorac Soc Vol 8. pp 444–450, 2011
racic and/or abdominal viscera. Situs inversus is another type of lat-
DOI: 10.1513/pats.201103-025SD erality disorder in which all internal organs are reversed. Typically,
Internet address: www.atsjournals.org patients with situs inversus do not have the congenital anomalies
Ware, Gunay-Aygun, and Hildebrandt: Non-PCD Ciliopathies 445

TABLE 1. CLASSIFICATION OF CILIA

Category Microtubule Arrangement Cilia per Cell Motility Function Cell Types

Motile cilia 912 Multiple Planar motion Mechanical; possible secondary Airways, spermatozoa, fallopian tubes,
chemosensory functions choroid plexus, and brain ventricles
Primary cilia 910 One Rotary motion Mechanosensory; chemosensory Embryonic node
Primary or sensory cilia 910 One Nonmotile Mechanosensory; chemosensory Renal tubules, pancreatic or bile ducts,
bone and cartilage, retina, cochlea,
developing heart, fibroblasts, etc.

that occur in heterotaxy syndrome. Embryologically, heterotaxy The nomenclature related to heterotaxy has traditionally
and situs inversus fall into the category of disorders of laterality be- been confusing, with both anatomic and clinical designations
cause they have their root in the abnormal development of the left– (e.g., right isomerism, asplenia syndrome) (9). Any internal or-
right body axis. Primary cilia present on the embryonic node are gan that is asymmetrically positioned can be abnormal in indi-
required for the generation of the left–right axis during embryonic viduals with heterotaxy. Improper symmetry, such as bilateral
development (7, 8). The ciliated embryonic node is unusual in that trilobed lungs, and failure to regress symmetric embryonic
it contains motile primary cilia that generate fluid flow and non- structures, such as persistent bilateral superior vena cava, are
motile primary cilia that act as mechanosensors. Abnormal situs is additional manifestations of heterotaxy. Midline defects occur
a hallmark feature of a number of ciliopathies. in approximately 40% of patients (10). The wide phenotypic

TABLE 2. SUMMARY OF CILIOPATHIES

Ciliopathy Features Gene(s) Pulmonary involvement

Primary ciliary dyskinesia Bronchiectasis, sinusitis, otitis media, DNAI1, DNAH5, DNAH11, Prominent
infertility, situs defects DNAI2, KTU, TXNDC3,
RPGR, RSPH4A, RSPH9,
LRRC50, CCDC40, CCDC39
Polycystic kidney disease Renal cysts; hepatic fibrosis; autosomal PKHD1, PKD1, PKD2 Reported: bronchiectasis
dominant and recessive forms
Nephronophthisis Renal cysts with or without extra-renal NPHP1-4, IQCB1, CEP290, Reported in conjunction with
symptoms GLIS2, RPGRIP1L, NEK8, allelic syndromes
SDCCAG8, TMEM67,
TTC21B
Senior-Loken syndrome Juvenile nephronophthisis, Leber amaurosis NPHP1-6, SDCCAG8 Not reported
Leber congenital Visual impairment in first year of life; GUCY2D, RPE65, LCA3-14 Reported: abnormal
amaurosis pigmentary retinopathy (including LCA10, CEP290) respiratory cilia
Meckel-Gruber syndrome Renal cysts, CNS anomalies (encephalocele), MKS1, TMEM216, TMEM67, Not reported
polydactyly, congenital heart defects CEP290, RPGRIP1L, CC2D2A
Joubert and related hypoplasia of the cerebellar vermis JBTS1, TMEM216, AHI1, Not reported: episodic
syndromes (molar tooth sign), dysregulated breathing NPHP1, CEP290, TMEM67, tachypnea 6 apnea described as
pattern, retinal dystrophy, renal anomalies RPGRIP1L, ARL13B, CC2D2A, central in origin
Bardet-Biedl syndrome Obesity, polydactyly, retinitis pigmentosa, BBS1, 2, ARL6, BBS4,5, MKKS, Not reported, but evidence for
anosmia, congenital heart defects BBS7, TTC8, BBS9, 10, TRIM32, airway ciliary dysfunction in animal
BBS12, MKS1, CEP290, C2ORF86; models
modifiers MKS1, MKS3, CCDC28B
Oral-facial-digital Oral cavity, face, and digit anomalies; OFD1 Reported: PCD phenotype
syndrome type I CNS abnormalities; cystic kidney disease;
X-linked with male lethality
Alstrom syndrome Dilated cardiomyopathy, obesity, ALMS1 Frequent: pneumonia, COPD,
sensorineural hearing loss, retinitis moderate to severe interstitial
pigmentosa, endocrine abnormalities, fibrosis
renal and hepatic disease
McKusick-Kaufman Urogenital anomalies including MKKS (BBS6) Not reported
syndrome hydrometrocolpos, postaxial polydactyly,
congenital heart defects
Ellis van Creveld Skeletal dysplasia; congenital heart EVC1, EVC2 Reported: respiratory distress
syndrome defects; polydactyly; ectodermal dysplasia due to narrow thorax
Jeunechondrodysplasia Skeletal dysplasia; thoracic deformities; IFT80 Frequent: pulmonary hypoplasia,
polydactyly; renal cysts; retinitis pigmentosa respiratory distress, restrictive lung
disease
Cranioectodermal dysplasia Cranioectodermal dysplasia; narrow thorax, IFT122, WDR35 Not reported
(Sensenbrenner syndrome) dental anomalies, hepatic and renal
involvement
Short rib polydactyly Lethal skeletal dysplasia, polydactyly, NEK1, DYNC2H1 Frequent: pulmonary hypoplasia,
multiple congenital anomalies respiratory distress, restrictive
lung disease
Heterotaxy (?) Visceral situs anomalies, asplenia or polysplenia, Frequent: situs anomalies;
congenital heart defects, biliary atresia, reported: PCD
midline defects

Definition of abbreviations: CNS ¼ central nervous system; PCD ¼ primary ciliary dyskinesia.
446 PROCEEDINGS OF THE AMERICAN THORACIC SOCIETY VOL 8 2011

spectrum and range of congenital anomalies that can be seen in cause heterotaxy (17–19). Gene mutations identified in human
heterotaxy syndrome has hindered clinical care and research laterality disorders have been reviewed recently (13). The ma-
progress. jority of the mutations identified have reduced penetrance and
“Classic” heterotaxy, in which a characteristic cardiovascular variable expressivity. Although most cases of heterotaxy are
malformation is identified in conjunction with visceral situs anom- single occurrences in a pedigree, approximately 10% of infants
alies, comprises approximately 3% of congenital heart defect with heterotaxy have a family history of a close relative with
cases and has an estimated prevalence of 1 in 10,000 live births. a congenital heart defect. Epidemiologic data strongly indicate
However, this most likely underestimates the true incidence of the important role of genetic factors in heterotaxy, but there is
heterotaxy spectrum disorders. Heterotaxy is a clinically and a fundamental lack of information about the genetic basis of
genetically heterogeneous condition, and patients with a combi- disease in the vast majority of cases.
nation of cardiovascular malformations and visceral situs anom-
alies represent the severe end of this spectrum. Isolated
cardiovascular malformations may be caused by mutations in OVERVIEW OF SYNDROMIC CILIOPATHIES
the same genes that cause heterotaxy, and therefore the true Diseases of the nonmotile cilia are commonly associated with
incidence of these defects of laterality is not known with cer- a spectrum of fibrocystic changes in the kidneys and the liver,
tainty (11). Overall, male patients with heterotaxy outnumber including PKD, nephronophthisis (NPHP) and renal cystic dys-
female patients 2:1, although within the polysplenia subtype the plasia, congenital hepatic fibrosis, Caroli’s disease, and polycys-
gender distribution is equal. Known environmental risk factors tic liver disease (20, 21). Other features include developmental
for heterotaxy include twin gestation, maternal diabetes, and anomalies of the cerebellum and midbrain, retinal degenera-
maternal cocaine use (12). tion, colobomas of the retina and iris, polydactyly, abnormal
The clinical evaluation for patients with heterotaxy syndrome bone growth, obesity, anosmia and other sensory defects, situs
is focused on delineating the anatomy and managing the congen- defects, and pancreatic cysts or dysplasia (Table 2). Autosomal
ital anomalies present (13). Most frequently, patients with het- dominant (ADPKD) and recessive (ARPKD) polycystic kidney
erotaxy present in the newborn period with cyanotic congenital diseases and NPHP are the most common hepatorenal fibrocystic
heart disease. Echocardiography is important for defining car- diseases. Pleiotropic syndromic ciliopathies with multisystem in-
diac anatomy. In some cases, chest MRA is useful for better volvement include Joubert syndrome (JS) and related cerebel-
visualization of the vasculature. Abdominal ultrasound is neces- lar disorders, Bardet-Biedl (BBS), Alstrom syndrome, Meckel-
sary to determine the position of the liver and stomach, spleen Gruber (MKS), Senior-Loken syndrome, and oral-facial-digital
position and number, and renal anatomy. Other imaging includes type 1 (OFD1) syndromes. Skeletal ciliopathies include Jeune
head ultrasound or brain MRI, upper gastrointestinal series to chondrodysplasia, cranioectodermal dysplasia, short-rib poly-
evaluate malrotation, chest radiograph and spine films, and con- dactyly, and Ellis-Van Creveld (EVC) syndromes.
sideration of hebatobiliaryscintigraphy. A blood smear for Liver disease in ciliopathies is not a primary disease of the
Howell-Jolly bodies is an initial evaluation for splenic function. hepatocytes but rather is a developmental defect of the portobili-
Chromosome microarray analysis is indicated due to the as- ary system, termed ductal plate malformation (DPM) (21, 22).
sociation with chromosome abnormalities. In addition, molecular DPM is defective remodeling of the biliary system characterized
testing for ZIC3, NODAL, CFC1, and FOXH1, four genes known by retention of excessive numbers of embryonic bile duct rem-
to cause heterotaxy, is available on a clinical basis. nants, abnormal portal veins, and periportal fibrosis. DPM of
Consideration should be given to evaluation for PCD in patients the peripheral biliary system causes congenital hepatic fibrosis,
with heterotaxy. The prevalence of PCD in this population is un- whereas DPM of the central biliary tree results in Caroli’s dis-
known, but recent clinical research and animal models suggest that ease. The combination of the two is referred to as Caroli’s
the underlying etiology may be identical for a subset of patients in syndrome. The characteristics of liver disease in ARPKD and
these groups. Neonatal respiratory distress (not related to cardio- ADPKD are different. ARPKD is characterized by congenital
vascular malformations) is a frequent manifestation of PCD. hepatic fibrosis and Caroli’s disease and is often complicated by
Chronic wet, productive cough; recurrent pneumonia; and persis- portal hypertension, whereas ADPKD patients develop poly-
tent rhinitis are important features that should trigger evaluation. cystic liver disease and typically do not have portal hyperten-
Bronchiectasis, chronic otitis media with conductive hearing loss, sion (20, 21). In contrast to Caroli’s disease/Caroli’s syndrome,
hydrocephalus, or retinitis pigmentosa are features of motile cil- in which liver cysts are dilated parts of the bile tree itself, poly-
iary dysfunction that should be recognized as symptoms unrelated cystic liver disease is characterized by isolated cysts that are not
to the cardiovascular malformations in heterotaxy and should in continuity with the bile tree.
prompt evaluation by the appropriate specialists, including pul- JS and related cerebellar disorders are a heterogeneous group
monologists. Further clinical research is necessary to better de- of disorders in which the defining common pathology is the
fine the degree of overlap between PCD and heterotaxy and to so-called “molar tooth sign” on axial brain MRI that is caused
provide better management of comorbidities. by a combination of mid- and hindbrain anomalies, including
Heterotaxy is highly heritable, indicating a strong genetic influ- cerebellar vermis hypoplasia and abnormal superior cerebellar
ence, although complex or multifactorial causation may predom- peduncles (23). Oculomotor apraxia and speech apraxia are
inate. X-linked heterotaxy is caused by mutations or deletions of typical. Variable features in JS include retinal degeneration,
the zinc-finger transcription factor ZIC3. We have investigated colobomas, renal disease (commonly in the form of NPHP),
the mechanistic basis of X-linked heterotaxy in mouse congenital hepatic fibrosis/Caroli’s syndrome, and polydactyly.
and Xenopus models (14–16). The results indicate that Zic3 acts BBS is characterized by retinal degeneration, cognitive impair-
early in development upstream of Nodal expression at the node. ment, obesity, and hypothalamic hypogonadism in male patients;
Further identification of the developmental pathways disrupted variable features include renal disease, congenital hepatic fibro-
by Zic3 loss of function will elucidate important candidate genes sis, polydactyly, cardiac defects, situs inversus, and abnormalities
for human heterotaxy and will further delineate shared and di- of internal genital organs in female patients (24). Älstrom syn-
vergent functions with genes causing PCD and other ciliopathies. drome is also characterized by retinal dystrophy, obesity, cardio-
Mutations in genes important in the generation of left–right myopathy, progressive sensorineural hearing impairment, and
patterning, such as NODAL and CFC1, have been shown to insulin resistance. MKS represents the most severe end of the
Ware, Gunay-Aygun, and Hildebrandt: Non-PCD Ciliopathies 447

nonmotile ciliopathy spectrum, with the typical triad of occipital GTPase regulator (RPGR). Recently, several signaling path-
encephalocele, cystic dysplastic kidneys, and postaxial polydac- ways have been implicated in the downstream signaling path-
tyly (25). Congenital hepatic fibrosis is an invariable feature in ways that connect ciliary and basal body function to the renal
MKS. OFD1 is an X-linked dominant, male-lethal, nonmotile cystic phenotype. These include the Wnt signaling and planar cell
ciliopathy characterized by prominent external features, includ- polarity pathways (41–43) and the hedgehog signaling pathway
ing oral clefts, hamartomas, or cysts of the tongue, and digital (44).
anomalies and visceral involvement, including PKD and biliary Recessive mutation of NPHP6/CEP290 was found to cause
and pancreatic cystic disease (26). Jeune chondrodysplasia, JS (45). NPHP6 encodes a centrosomal protein, nephrocystin-6,
which is also referred to as asphyxiating thoracic dysplasia, is which modulates the activity of ATF4/CREB2, a transcription
characterized by small thorax due to short ribs, short stature factor implicated in cAMP-dependent renal cyst formation.
that is mainly caused by short limbs, and polydactyly (27). Jeune We demonstrated that abrogation of nphp6 function in zebra-
patients who survive the small thorax-related respiratory dis- fish causes planar cell polarity (convergent extension) defects
tress develop NPHP, retinopathy, and fibrocystic disease of and recapitulates the human phenotype of JS. Hypomorphic
the pancreas. EVC is characterized by short stature associated mutations in NPHP6/CEP290 were shown to cause retinal de-
with short limbs, congenital cardiac defects, teeth abnormalities, generation (rds16) in mice (46) and represent a major cause of
congenital hepatic fibrosis, and situs anomalies in some cases Leber’s congenital amaurosis in humans (47). Identification of
(28). Cranioectodermal dysplasia is characterized by frontal the NPHP6 gene, therefore, established a link between cen-
predominance and widely spaced eyes; teeth, hair, and nail ab- trosome function and tissue architecture in the pathogenesis
normalities; congenital hepatic fibrosis; and a NPHP-like renal of cystic kidney disease and in central nervous system devel-
pathology (29). opment. Mutation of GLIS2/NPHP7 causes NPHP type 7 in
Genetic heterogeneity (multiple genes causing similar clinical humans and mice (48). Differential gene expression studies on
phenotype) and genotypic and phenotypic overlap are two char- Glis2 mutant kidneys demonstrated that genes promoting
acteristics of syndromic ciliopathies (30, 31). There are multiple epithelial-to-mesenchymal transition and fibrosis are up-regu-
genes identified for JS and related cerebellar disorders, BBS, and lated in the absence of Glis2, demonstrating that Glis2 is essen-
MKS, still not accounting for all patients. These disorders were tial for the maintenance of renal tissue architecture through
originally defined based on their most prominent clinical fea- prevention of apoptosis and fibrosis. Recently, mutations of
tures. As we understand these disorders better, we realize that RPGRIP1L/NPHP8 were shown to cause MKS and JS (49),
the borders between them are blurred at the phenotypic and ge- and it has been demonstrated that NEK8 mutations may rarely
notypic levels. cause NPHP (50).

CYSTIC KIDNEY DISEASES AND GENETIC AND PHENOTYPIC OVERLAP

NEPHRONOPHTHISIS-RELATED CILIOPATHIES OF CILIOPATHIES
Nephronophthisis (NPHP), an autosomal recessive cystic kidney Ciliary dysfunction leads to a wide range of phenotypes, many of
disease, represents the most common genetic cause of terminal which overlap. In recessive ciliopathies, the nature of the two re-
renal failure occurring in the first three decades of life. It may be cessive mutations determines the severity and extent of organ
associated with retinitis pigmentosa (RP) in the renal-retinal dis- involvement, leading to seemingly different disorders. In this con-
order Senior-Loken syndrome or with cerebellar vermis aplasia text, loss-of-function mutations cause severe, early-onset, dysplas-
and mental retardation in JS. Ten different recessive genes have tic, multiorgan disease, whereas reduced-function mutations cause
been identified as mutated in NPHP-related ciliopathies (NPHP- mild, late-onset, degenerative disease with limited organ involve-
RC) with and without extrarenal manifestations. ment. For example, recent data indicate that mutations in
NPHP1 is mutated in NPHP type 1 (32). Its gene product, TTC21B, which encodes the intraflagellar transport protein
nephrocystin-1, localizes to transition fibers of cilia and acts in IFT139, cause isolated NPHP or Jeune asphyxiating thoracic
focal adhesion signaling. Mutations in the NPHP2/inversin gene dystrophy depending on mutation type (51). Causal genes can
cause infantile NPHP (type 2) with the rare association of situs also act as modifier alleles in clinically distinct disorders. In the
inversus and RP. Demonstration of expression of NPHP1 and case of TTC21B, sequence-based interrogation of a large ciliop-
NPHP2 in primary cilia of renal epithelial cells (33) supported athy cohort identified pathogenic rare variants in approximately
the “ciliopathy theory” for the pathogenesis of cystic kidney dis- 5% of cases. These alleles are proposed to act in trans with
eases (34). This theory states that the products (“cystoproteins”) other disease causing genes to influence the range of ciliopathy
of all genes mutated in cystic kidney disease in humans, mice, or phenotypes.
zebrafish are expressed in primary cilia, basal bodies, or centro- In human ciliary disorders, motor and sensory functions
somes of renal epithelial cells (35). Identification of NPHP3 as have been considered distinct properties of motile and nonmo-
mutated in NPHP type 3 supported this theory and revealed an tile cilia, respectively. However, there are recent data that in-
Nphp3 mutation as causing the renal cystic disease in pcy mice dicate that motile cilia of the airway are important for sensory
(36). For this animal model, efficient treatment was demon- reception (52, 53). It is theoretically possible that defects in
strated (37). certain ciliary proteins whose functions are important for the
Some of the general features of renal cystic diseases are ex- motile and nonmotile cilia can result in dysfunction of both
emplified by studies on the NPHP gene products nephrocystins. types of cilia, thereby causing diverse phenotypes. For example,
Positional cloning of NPHP4 as mutated in Senior-Loken syn- recently seven patients with Leber’s congenital amarosis due to
drome (38) led to the demonstration that its gene product, CEP290 mutations were shown to have defective respiratory cilia
nephrocystin-4, is conserved in Caenorhabditis elegans and (54). In addition, patients with autosomal dominant polycystic
expressed together with nephrocystin-1 in ciliated head and tail kidney disease have been shown to have increased prevalence
neurons of the nematode (39). The role of primary cilia function of radiographic bronchiectasis (55). Additional examples of sub-
for retinal–renal syndromes was confirmed by identification of tle or subclinical phenotypic overlap may become apparent with
the novel NPHP5 gene (40), demonstrating interaction of the growing clinical appreciation of the spectrum of ciliopathy
nephrocystin-5 with calmodulin and the retinitis pigmentosa features.
448 PROCEEDINGS OF THE AMERICAN THORACIC SOCIETY VOL 8 2011

Heterotaxy, PCD, and other ciliopathies that occasionally defects in three-dimensional renal cell cultures. Further gene
have abnormal laterality as features share a similar underlying identification in NPHP will result in the definition of functional
pathogenesis centering on the role of cilia. Heterotaxy may result networks of primary cilia, centrosomes, and planar cell polarity
from defects in any one of a series of signaling or morphogenetic as they pertain to the pathogenic mechanisms of NPHP-
processes that are required for the establishment of the left-right associated syndromes and other cystic kidney diseases.
axis. In contrast, situs inversus caused by PCD usually results
from specific defects in the structure and function of cilia at the
CONCLUSIONS
node. Recent data indicate that not less than 6% of patients
with PCD have heterotaxy, highlighting the overlap of these Primary cilia are ubiquitous organelles that serve a sensory func-
conditions (56). Situs defects are also observed in nonmotile tion. Disruption of the function of primary cilia results in clini-
ciliopathies, including RHPD, EVC, JC, and rare cases of cally heterogeneous disorders that may affect a single organ or
BBS, JS, and OFD1. There is a lack of clarity regarding the result in a syndromic phenotype. There is substantial clinical and
mechanisms underlying the development of situs inversus ver- genetic heterogeneity. There is some evidence suggesting that
sus heterotaxy as phenotypes. An improved mechanistic under- certain ciliary defects might affect primary and motile cilia asso-
standing of the steps leading to the development of abnormal ciated with PCD. Additional research is needed to better under-
laterality is critical to delineate the interrelatedness of these stand the biological functions of these diverse classes of cilia, the
disorders and to direct proper management and therapy. genetic basis of disease, and the management of clinical symp-
toms caused by functional disruption.
RESEARCH AND FUTURE DIRECTIONS Author Disclosure: None of the authors has a financial relationship with a com-
mercial entity that has an interest in the subject of this manuscript.
Within the last 5 years, the improved technology available for
genetic analyses has had a dramatic impact on the investigation Acknowledgments: The authors gratefully acknowledge the sponsorship and
of ciliopathies. This has facilitated a more rapid identification of meeting organization provided by the Primary Ciliary Dyskinesia Foundation.
causative genes, investigation of modifier alleles, expansion and
refinement of genotype–phentoype correlations, and improve-
ment of clinical diagnostics. References
The use of genome-wide copy number analysis coupled with 1. Fliegauf M, Benzing T, Omran H. When cilia go bad: cilia defects and
in vitro and in vivo analyses of gene function is a powerful ciliopathies. Nat Rev Mol Cell Biol 2007;8:880–893.
technique for the identification of novel genetic causes of cilio- 2. Adams NA, Awadein A, Toma HS. The retinal ciliopathies. Ophthalmic
pathies. Imbalances in chromosomal copy number resulting in Genet 2007;28:113–125.
deletion or duplication of a small number of genes are a known 3. Christensen ST, Pedersen LB, Schneider L, Satir P. Sensory cilia and
integration of signal transduction in human health and disease. Traffic
cause of congenital heart defects in a small number of patients
2007;8:97–109.
with genetic syndromes. Until recently, it has not been possible 4. Cardenas-Rodriguez M, Badano JL. Ciliary biology: understanding the cellular
to evaluate the entire genome for these genetic alterations. Re- and genetic basis of human ciliopathies. Am J Med Genet C Semin Med
cently, whole–genome, single-nucleotide polymorphism genotyp- Genet 2009;151C:263–280.
ing was used to evaluate chromosomal copy number imbalances 5. Singla V, Reiter JF. The primary cilium as the cell’s antenna: signaling at
in a well characterized cohort of patients with complex cardio- a sensory organelle. Science 2006;313:629–633.
vascular malformations and heterotaxy syndrome. The results 6. Mok CA, Heon E, Zhen M. Ciliary dysfunction and obesity. Clin Genet
indicate that 15 to 30% of patients have pathogenic copy num- 2010;77:18–27.
ber variation. Efforts are ongoing to identify dosage-sensitive 7. Hamada H. Breakthroughs and future challenges in left-right patterning.
genes within these regions of chromosomal imbalance that will Dev Growth Differ 2008;50:S71–S78.
8. Levin M. Left-right asymmetry in embryonic development: a compre-
allow for identification of novel genes required for cardiac de-
hensive review. Mech Dev 2005;122:3–25.
velopment and patterning (57). Identification of specific genes 9. Zhu L, Belmont JW, Ware SM. Genetics of human heterotaxias. Eur
underlying heterotaxy spectrum congenital heart disease will J Hum Genet 2006;14:17–25.
lead to the development of novel diagnostics and therapeutics 10. Ticho BS, Goldstein AM, Van Praagh R. Extracardiac anomalies in the
and improved management. heterotaxy syndromes with focus on anomalies of midline-associated
Sequence-based analysis is another strategy that has been en- structures. Am J Cardiol 2000;85:729–734.
hanced by the development of novel genomic technology and 11. Ware SM, Peng J, Zhu L, Fernbach S, Colicos S, Casey B, Towbin J,
subsequently has proven tremendously valuable for the study Belmont JW. Identification and functional analysis of zic3 mutations
of ciliopathies. Massively parallel resequencing improves the in heterotaxy and related congenital heart defects. Am J Hum Genet
2004;74:93–105.
ability to interrogate large numbers of disease-causing genes
12. Kuehl KS, Loffredo C. Risk factors for heart disease associated with
in well phenotyped cohorts, and as a result there is a wider ap- abnormal sidedness. Teratology 2002;66:242–248.
preciation of the genetic architecture and phenotypic spectrum 13. Sutherland MJ, Ware SM. Disorders of left-right asymmetry: heterotaxy
of the ciliopathies (58, 59). These methods are also translating and situs inversus. Am J Med Genet C Semin Med Genet 2009;151C:
into improved availability of testing on a clinical basis. How- 307–317.
ever, challenges remain in the interpretation of rare variants in 14. Purandare SM, Ware SM, Kwan KM, Gebbia M, Bassi MT, Deng JM,
a clinical setting. Exome capture technology allows unbiased Vogel H, Behringer RR, Belmont JW, Casey B. A complex syndrome
interrogation of all coding region of the genome, allowing iden- of left-right axis, central nervous system and axial skeleton defects in
tification of novel disease-causing genes. This approach is par- zic3 mutant mice. Development 2002;129:2293–2302.
ticularly useful for autosomal recessive conditions. Recently, 15. Ware SM, Harutyunyan KG, Belmont JW. Heart defects in x–linked
heterotaxy: evidence for a genetic interaction of zic3 with the nodal
exome capture identified 12 different truncating mutations of
signaling pathway. Dev Dyn 2006;235:1631–1637.
SDCCAG8 (serologically defined colon cancer antigen 8) in 20 16. Ware SM, Harutyunyan KG, Belmont JW. Zic3 is critical for early
individuals with Senior-Loken syndrome of 10 different families embryonic patterning during gastrulation. Dev Dyn 2006;235:776–785.
(60). SDCCAG8 is localized at both centrioles and interacts 17. Mohapatra B, Casey B, Li H, Ho-Dawson T, Smith L, Fernbach SD,
directly with OFD1. Depletion of sdccag8 causes kidney cysts Molinari L, Niesh SR, Jefferies JL, Craigen WJ, et al. Identification
and a body axis defect in zebrafish and induces cell polarity and functional characterization of nodal rare variants in heterotaxy
Ware, Gunay-Aygun, and Hildebrandt: Non-PCD Ciliopathies 449

and isolated cardiovascular malformations. Hum Mol Genet 2009;18: nephronophthisis and retinitis pigmentosa encodes a novel protein,
861–871. nephroretinin, conserved in evolution. Am J Hum Genet 2002;71:1161–
18. Goldmuntz E, Bamford R, Karkera JD, dela Cruz J, Roessler E, 1167.
Muenke M. Cfc1 mutations in patients with transposition of the great 39. Wolf MT, Lee J, Panther F, Otto EA, Guan KL, Hildebrandt F. Expression
arteries and double-outlet right ventricle. Am J Hum Genet 2002;70: and phenotype analysis of the nephrocystin-1 and nephrocystin-
776–780. 4 homologs in caenorhabditis elegans. J Am Soc Nephrol 2005;16:
19. Roessler E, Ouspenskaia MV, Karkera JD, Velez JI, Kantipong A, 676–687.
Lacbawan F, Bowers P, Belmont JW, Towbin JA, Goldmuntz E, et al. 40. Otto EA, Loeys B, Khanna H, Hellemans J, Sudbrak R, Fan S, Muerb
Reduced nodal signaling strength via mutation of several pathway U, O’Toole JF, Helou J, Attanasio M, et al. Nephrocystin-5, a ciliary
members including foxh1 is linked to human heart defects and hol- iq domain protein, is mutated in senior-loken syndrome and interacts
oprosencephaly. Am J Hum Genet 2008;83:18–29. with rpgr and calmodulin. Nat Genet 2005;37:282–288.
20. Gunay-Aygun M. Liver and kidney disease in ciliopathies. Am J Med 41. Ross AJ, May-Simera H, Eichers ER, Kai M, Hill J, Jagger DJ, Leitch
Genet C Semin Med Genet 2009;151C:296–306. CC, Chapple JP, Munro PM, Fisher S, et al. Disruption of bardet-biedl
21. Gunay-Aygun M, Heller T, Gahl WA. Congenital hepatic fibrosis syndrome ciliary proteins perturbs planar cell polarity in vertebrates.
overview. In: GeneReviews at GeneTests: Medical genetics in- Nat Genet 2005;37:1135–1140.
formation resource (database online) [accessed March 10, 2011]. 42. Simons M, Gloy J, Ganner A, Bullerkotte A, Bashkurov M, Kronig C,
Available from: http://wwwgenetestsorg. University of Washington, Schermer B, Benzing T, Cabello OA, Jenny A, et al. Inversin, the
Seattle, WA; 2008. gene product mutated in nephronophthisis type ii, functions as a mo-
22. Desmet VJ. Congenital diseases of intrahepatic bile ducts: variations on lecular switch between wnt signaling pathways. Nat Genet 2005;37:
the theme “ductal plate malformation”. Hepatology 1992;16:1069– 537–543.
1083. 43. Fischer E, Legue E, Doyen A, Nato F, Nicolas JF, Torres V, Yaniv M,
23. Parisi MA, Doherty D, Chance PF, Glass IA. Joubert syndrome (and Pontoglio M. Defective planar cell polarity in polycystic kidney dis-
related disorders) (omim 213300). Eur J Hum Genet 2007;15:511–521. ease. Nat Genet 2006;38:21–23.
24. Beales RA. Bardet-biedl syndrome [accessed March 10, 2011]. Gen- 44. Huangfu D, Liu A, Rakeman AS, Murcia NS, Niswander L, Anderson
eReviews; 2007. Available from: wwwgenereviewsorg KV. Hedgehog signalling in the mouse requires intraflagellar transport
25. Alexiev BA, Lin X, Sun CC, Brenner DS. Meckel-gruber syndrome: proteins. Nature 2003;426:83–87.
pathologic manifestations, minimal diagnostic criteria, and differen- 45. Sayer JA, Otto EA, O’Toole JF, Nurnberg G, Kennedy MA, Becker
tial diagnosis. Arch Pathol Lab Med 2006;130:1236–1238. C, Hennies HC, Helou J, Attanasio M, Fausett BV, et al. The
26. Chetty-John S, Piwnica-Worms K, Bryant J, Bernardini I, Fischer RE, centrosomal protein nephrocystin-6 is mutated in joubert syndrome
Heller T, Gahl WA, Gunay-Aygun M. Fibrocystic disease of liver and and activates transcription factor atf4. Nat Genet 2006;38:674–681.
pancreas; under-recognized features of the x–linked ciliopathy oral- 46. Chang B, Khanna H, Hawes N, Jimeno D, He S, Lillo C, Parapuram SK,
facial-digital syndrome type 1 (ofd i). Am J Med Genet A 2010;152A: Cheng H, Scott A, Hurd RE, et al. In-frame deletion in a novel
2640–2645. centrosomal/ciliary protein cep290/nphp6 perturbs its interaction with
27. Tuysuz B, Baris S, Aksoy F, Madazli R, Ungur S, Sever L. Clinical rpgr and results in early-onset retinal degeneration in the rd16 mouse.
variability of asphyxiating thoracic dystrophy (jeune) syndrome: Hum Mol Genet 2006;15:1847–1857.
evaluation and classification of 13 patients. Am J Med Genet A 2009; 47. den Hollander AI, Koenekoop RK, Yzer S, Lopez I, Arends ML,
149A:1727–1733. Voesenek KE, Zonneveld MN, Strom TM, Meitinger T, Brunner
28. Ruiz-Perez VL, Goodship JA. Ellis-van creveld syndrome and weyers HG, et al. Mutations in the cep290 (nphp6) gene are a frequent cause
acrodental dysostosis are caused by cilia-mediated diminished re- of leber congenital amaurosis. Am J Hum Genet 2006;79:556–561.
sponse to hedgehog ligands. Am J Med Genet C Semin Med Genet 48. Attanasio M, Uhlenhaut NH, Sousa VH, O’Toole JF, Otto E, Anlag K,
2009;151C:341–351. Klugmann C, Treier AC, Helou J, Sayer JA, et al. Loss of glis2 causes
29. Zaffanello M, Diomedi-Camassei F, Melzi ML, Torre G, Callea F, nephronophthisis in humans and mice by increased apoptosis and fi-
Emma F. Sensenbrenner syndrome: a new member of the hepatorenal brosis. Nat Genet 2007;39:1018–1024.
fibrocystic family. Am J Med Genet A 2006;140:2336–2340. 49. Delous M, Baala L, Salomon R, Laclef C, Vierkotten J, Tory K, Golzio
30. Tobin JL, Beales PL. The nonmotile ciliopathies. Genet Med 2009;11: C, Lacoste T, Besse L, Ozilou C, et al. The ciliary gene rpgrip1l is
386–402. mutated in cerebello-oculo-renal syndrome (joubert syndrome type b)
31. Gunay-Aygun M, Parisi MA, Doherty D, Tuchman M, Tsilou E, Kleiner and meckel syndrome. Nat Genet 2007;39:875–881.
DE, Huizing M, Turkbey B, Choyke P, Guay-Woodford L, et al. 50. Otto EA, Trapp ML, Schultheiss UT, Helou J, Quarmby LM,
Mks3-related ciliopathy with features of autosomal recessive poly- Hildebrandt F. Nek8 mutations affect ciliary and centrosomal locali-
cystic kidney disease, nephronophthisis, and joubert syndrome. zation and may cause nephronophthisis. J Am Soc Nephrol 2008;19:
J Pediatr 2009;155:386–392, e381. 587–592.
32. Hildebrandt F, Otto E, Rensing C, Nothwang HG, Vollmer M, Adolphs 51. Davis EE, Zhang Q, Liu Q, Diplas BH, Davey LM, Hartley J, Stoetzel
J, Hanusch H, Brandis M. A novel gene encoding an sh3 domain C, Szymanska K, Ramaswami G, Logan CV, et al. Ttc21b contributes
protein is mutated in nephronophthisis type 1. Nat Genet 1997;17:149– both causal and modifying alleles across the ciliopathy spectrum. Nat
153. Genet 2011;43:189–196.
33. Otto EA, Schermer B, Obara T, O’Toole JF, Hiller KS, Mueller AM, 52. Shah AS, Ben-Shahar Y, Moninger TO, Kline JN, Welsh MJ. Motile
Ruf RG, Hoefele J, Beekmann F, Landau D, et al. Mutations in invs cilia of human airway epithelia are chemosensory. Science 2009;325:
encoding inversin cause nephronophthisis type 2, linking renal cystic 1131–1134.
disease to the function of primary cilia and left-right axis de- 53. Bloodgood RA. Sensory reception is an attribute of both primary cilia
termination. Nat Genet 2003;34:413–420. and motile cilia. J Cell Sci 2010;123:505–509.
34. Watnick T, Germino G. From cilia to cyst. Nat Genet 2003;34:355–356. 54. Papon JF, Perrault I, Coste A, Louis B, Gerard X, Hanein S, Fares-Taie
35. Hildebrandt F, Otto E. Cilia and centrosomes: a unifying pathogenic L, Gerber S, Defoort-Dhellemmes S, Vojtek AM, et al. Abnormal
concept for cystic kidney disease? Nat Rev Genet 2005;6:928–940. respiratory cilia in non-syndromic leber congenital amaurosis with
36. Olbrich H, Fliegauf M, Hoefele J, Kispert A, Otto E, Volz A, Wolf MT, cep290 mutations. J Med Genet 2010;47:829–834.
Sasmaz G, Trauer U, Reinhardt R, et al. Mutations in a novel gene, 55. Driscoll JA, Bhalla S, Liapis H, Ibricevic A, Brody SL. Autosomal
nphp3, cause adolescent nephronophthisis, tapeto-retinal degeneration dominant polycystic kidney disease is associated with an increased
and hepatic fibrosis. Nat Genet 2003;34:455–459. prevalence of radiographic bronchiectasis. Chest 2008;133:1181–1188.
37. Gattone VH II, Wang X, Harris PC, Torres VE. Inhibition of renal cystic 56. Kennedy MP, Omran H, Leigh MW, Dell S, Morgan L, Molina PL,
disease development and progression by a vasopressin v2 receptor Robinson BV, Minnix SL, Olbrich H, Severin T, et al. Congenital
antagonist. Nat Med 2003;9:1323–1326. heart disease and other heterotaxic defects in a large cohort of
38. Otto E, Hoefele J, Ruf R, Mueller AM, Hiller KS, Wolf MT, Schuermann patients with primary ciliary dyskinesia. Circulation 2007;115:2814–
MJ, Becker A, Birkenhager R, Sudbrak R, et al. A gene mutated in 2821.
450 PROCEEDINGS OF THE AMERICAN THORACIC SOCIETY VOL 8 2011

57. Fakhro KA, Choi M, Ware SM, Belmont JW, Towbin JA, Lifton RP, 59. Otto EA, Ramaswami G, Janssen S, Chaki M, Allen SJ, Zhou W, Airik
Khokha MK, Brueckner M. Rare copy number variations in con- R, Hurd TW, Ghosh AK, Wolf MT, et al. Mutation analysis of 18
genital heart disease patients identify unique genes in left-right pat- nephronophthisis associated ciliopathy disease genes using a DNA
terning. Proc Natl Acad Sci USA 2011;108:2915–2920. pooling and next generation sequencing strategy. J Med Genet 2011;
58. Janssen S, Ramaswami G, Davis EE, Hurd T, Airik R, Kasanuki JM, 48:105–116.
60. Otto EA, Hurd TW, Airik R, Chaki M, Zhou W, Stoetzel C, Patil SB,
Van Der Kraak L, Allen SJ, Beales PL, Katsanis N, et al. Mutation
Levy S, Ghosh AK, Murga-Zamalloa CA, et al. Candidate exome
analysis in bardet-biedl syndrome by DNA pooling and massively
capture identifies mutation of sdccag8 as the cause of a retinal-renal
parallel resequencing in 105 individuals. Hum Genet 2011;129:79–90. ciliopathy. Nat Genet 2010;42:840–850.