A prioritization tool for cilia-associated genes and their in vivo resources unveils new avenues for ciliopathy research.

Defects in ciliary signaling or mutations in proteins that localize to primary cilia lead to a class of human diseases known as ciliopathies. Approximately 10% of mammalian genes encode cilia-associated proteins, and a major gap in the cilia research field is knowing which genes to prioritize to study and finding the in vivo vertebrate mutant alleles and reagents available for their study. Here, we present a unified resource listing the cilia-associated human genes cross referenced to available mouse and zebrafish mutant alleles, and their associated phenotypes, as well as expression data in the kidney and functional data for vertebrate Hedgehog signaling. This resource empowers researchers to easily sort and filter genes based on their own expertise and priorities, cross reference with newly generated -omics datasets, and quickly find in vivo resources and phenotypes associated with a gene of interest.

Ccrk-Mak/Ick signaling is a ciliary transport regulator essential for retinal photoreceptor survival.

Primary cilia are microtubule-based sensory organelles whose dysfunction causes ciliopathies in humans. The formation, function, and maintenance of primary cilia depend crucially on intraflagellar transport (IFT); however, the regulatory mechanisms of IFT at ciliary tips are poorly understood. Here, we identified that the ciliopathy kinase Mak is a ciliary tip-localized IFT regulator that cooperatively acts with the ciliopathy kinase Ick, an IFT regulator. Simultaneous disruption of Mak and Ick resulted in loss of photoreceptor ciliary axonemes and severe retinal degeneration. Gene delivery of Ick and pharmacological inhibition of FGF receptors, Ick negative regulators, ameliorated retinal degeneration in Mak -/- mice. We also identified that Ccrk kinase is an upstream activator of Mak and Ick in retinal photoreceptor cells. Furthermore, the overexpression of Mak, Ick, and Ccrk and pharmacological inhibition of FGF receptors suppressed ciliopathy-related phenotypes caused by cytoplasmic dynein inhibition in cultured cells. Collectively, our results show that the Ccrk-Mak/Ick axis is an IFT regulator essential for retinal photoreceptor maintenance and present activation of Ick as a potential therapeutic approach for retinitis pigmentosa caused by MAK mutations.

CC2D1A causes ciliopathy, intellectual disability, heterotaxy, renal dysplasia, and abnormal CSF flow.

Intellectual and developmental disabilities result from abnormal nervous system development. Over a 1,000 genes have been associated with intellectual and developmental disabilities, driving continued efforts toward dissecting variant functionality to enhance our understanding of the disease mechanism. This report identified two novel variants in CC2D1A in a cohort of four patients from two unrelated families. We used multiple model systems for functional analysis, including Xenopus, Drosophila, and patient-derived fibroblasts. Our experiments revealed that cc2d1a is expressed explicitly in a spectrum of ciliated tissues, including the left-right organizer, epidermis, pronephric duct, nephrostomes, and ventricular zone of the brain. In line with this expression pattern, loss of cc2d1a led to cardiac heterotaxy, cystic kidneys, and abnormal CSF circulation via defective ciliogenesis. Interestingly, when we analyzed brain development, mutant tadpoles showed abnormal CSF circulation only in the midbrain region, suggesting abnormal local CSF flow. Furthermore, our analysis of the patient-derived fibroblasts confirmed defective ciliogenesis, further supporting our observations. In summary, we revealed novel insight into the role of CC2D1A by establishing its new critical role in ciliogenesis and CSF circulation.

A class I PI3K signalling network regulates primary cilia disassembly in normal physiology and disease.

Primary cilia are antenna-like organelles which sense extracellular cues and act as signalling hubs. Cilia dysfunction causes a heterogeneous group of disorders known as ciliopathy syndromes affecting most organs. Cilia disassembly, the process by which cells lose their cilium, is poorly understood but frequently observed in disease and upon cell transformation. Here, we uncover a role for the PI3Kα signalling enzyme in cilia disassembly. Genetic PI3Kα-hyperactivation, as observed in PIK3CA-related overgrowth spectrum (PROS) and cancer, induced a ciliopathy-like phenotype during mouse development. Mechanistically, PI3Kα and PI3Kß produce the PIP3 lipid at the cilia transition zone upon disassembly stimulation. PI3Kα activation initiates cilia disassembly through a kinase signalling axis via the PDK1/PKCι kinases, the CEP170 centrosomal protein and the KIF2A microtubule-depolymerising kinesin. Our data suggest diseases caused by PI3Kα-activation may be considered 'Disorders with Ciliary Contributions', a recently-defined subset of ciliopathies in which some, but not all, of the clinical manifestations result from cilia dysfunction.

Obesity-Related Ciliopathies: Focus on Advances of Biomarkers.

Obesity-related ciliopathies, as a group of ciliopathies including Alström Syndrome and Bardet-Biedl Syndrome, exhibit distinct genetic and phenotypic variability. The understanding of these diseases is highly significant for understanding the functions of primary cilia in the human body, particularly regarding the relationship between obesity and primary cilia. The diagnosis of these diseases primarily relies on clinical presentation and genetic testing. However, there is a significant lack of research on biomarkers to elucidate the variability in clinical manifestations, disease progression, prognosis, and treatment responses. Through an extensive literature review, the paper focuses on obesity-related ciliopathies, reviewing the advancements in the field and highlighting the potential roles of biomarkers in the clinical presentation, diagnosis, and prognosis of these diseases.

Pleiotropy in FOXC1-attributable phenotypes involves altered ciliation and cilia-dependent signaling.

Alterations to cilia are responsible for a wide range of severe disease; however, understanding of the transcriptional control of ciliogenesis remains incomplete. In this study we investigated whether altered cilia-mediated signaling contributes to the pleiotropic phenotypes caused by the Forkhead transcription factor FOXC1. Here, we show that patients with FOXC1-attributable Axenfeld-Rieger Syndrome (ARS) have a prevalence of ciliopathy-associated phenotypes comparable to syndromic ciliopathies. We demonstrate that altering the level of Foxc1 protein, via shRNA mediated inhibition, CRISPR/Cas9 mutagenesis and overexpression, modifies cilia length in vitro. These structural changes were associated with substantially perturbed cilia-dependent signaling [Hedgehog (Hh) and PDGFRα], and altered ciliary compartmentalization of the Hh pathway transcription factor, Gli2. Consistent with these data, in primary cultures of murine embryonic meninges, cilia length was significantly reduced in heterozygous and homozygous Foxc1 mutants compared to controls. Meningeal expression of the core Hh signaling components Gli1, Gli3 and Sufu was dysregulated, with comparable dysregulation of Pdgfrα signaling evident from significantly altered Pdgfrα and phosphorylated Pdgfrα expression. On the basis of these clinical and experimental findings, we propose a model that altered cilia-mediated signaling contributes to some FOXC1-induced phenotypes.

Spectrum and frequencies of extraocular features reported in CEP290-associated ciliopathy - A systematic review.

Pathogenic variants in the CEP290 gene may result in a broad spectrum of diseases, ranging from lethal neonatal syndromes to isolated retinopathy. A detailed review of the clinical spectrum with the incidence of affected extraocular systems has not yet been published. A review of published papers was carried out to provide a comprehensive report on systemic signs and symptoms associated with CEP290 ciliopathies and to explore the genotype-phenotype correlation. Genetic and clinical data were collected on patients with biallelic variants in the CEP290 gene and the extraocular tissues affected. Genotype-phenotype analysis was performed. Two hundred thirty-five patients were included in the analysis. The most frequently reported organs affected, after the eye, were the central nervous system (82.6%, 194/235), followed by the kidney (53.2%, 125/235), skeletal system (15.3% 36/235), and a large spectrum of other, less frequently reported clinical manifestations. Patients with two variants that together predictably resulted in a low amount of CEP290 protein showed a significant association with having two or more extraocular organ systems affected. This is the most extensive report to date on patients with CEP290-ciliopathy and affected extraocular tissues. Based on these findings and previous publications, systemic screening is proposed, together with a clinical pathway for patients with CEP290-related ciliopathy.

Expanding the phenotypic spectrum of CC2D2A-related ciliopathies: a rare homozygous nonsense variant in a patient with suspected nephronophthisis.

Biallelic pathogenic variants in the gene CC2D2A cause a spectrum of ciliopathies, including Joubert and Meckel syndrome, which frequently involve the kidney; however, no cases of isolated renal disease (i.e., nephronophthisis) have yet been reported. In an adult with a clinical presentation consistent with nephronophthisis, next-generation sequencing identified a rare homozygous nonsense variant in CC2D2A (c.100 C > T; p.(Arg34*)). Tissue-specific expression data and promoter activity analysis demonstrates that this variant primarily affects a transcript isoform predominant in the kidneys but does not affect the transcript isoforms predominant in other tissues typically involved in CC2D2A-related ciliopathies (e.g., cerebellum, liver). Expression analysis of patient-specific cDNA in MDCK cells demonstrates partial translation re-initiation downstream of p.(Arg34*) as a possible escape mechanism from nonsense mediated decay. These data provide mechanistic insights in support of this novel genotype-phenotype association.

Ciliopathies are responsible for short stature and insulin resistance: A systematic review of this clinical association regarding SOFT syndrome.

SOFT syndrome (Short stature-Onychodysplasia-Facial dysmorphism-hypoTrichosis) is a rare primordial dwarfism syndrome caused by biallelic variants in POC1A encoding a centriolar protein. To refine the phenotypic spectrum of SOFT syndrome, recently shown to include metabolic features, we conducted a systematic review of all published cases (19 studies, including 42 patients). The SOFT tetrad affected only 24 patients (57%), while all cases presented with short stature from birth (median height: -5.5SDS([-8.5]-[-2.8])/adult height: 132.5 cm(103.5-148)), which was most often disproportionate (90.5%), with relative macrocephaly. Bone involvement resulted in short hands and feet (100%), brachydactyly (92.5%), metaphyseal (92%) or epiphyseal (84%) anomalies, and/or sacrum/pelvis hypoplasia (58%). Serum IGF-I was increased (median IGF-I level: + 2 SDS ([-0.5]-[+ 3])). Recombinant human growth hormone (rhGH) therapy was stopped for absence/poor growth response (7/9 patients, 78%) and/or hyperglycemia (4/9 patients, 45%). Among 11 patients evaluated, 10 (91%) presented with central distribution of fat (73%), clinical (64%) and/or biological insulin resistance (IR) (100%, median HOMA-IR: 18), dyslipidemia (80%), and hepatic steatosis (100%). Glucose tolerance abnormalities affected 58% of patients aged over 10 years. Patients harbored biallelic missense (52.4%) or truncating (45.2%) POC1A variants. Biallelic null variants, affecting 36% of patients, were less frequently associated with the SOFT tetrad (33% vs 70% respectively, p = 0.027) as compared to other variants, without difference in the prevalence of metabolic abnormalities. POC1A should be sequenced in children with short stature, altered glucose/insulin homeostasis and/or centripetal fat distribution. In patients with SOFT syndrome, rhGH treatment is not indicated, and IR-related complications should be regularly screened and monitored.PROSPERO registration: CRD42023460876.

Expanding the Phenotypic Spectrum of Pathogenic KIAA0586 Variants: From Joubert Syndrome to Hydrolethalus Syndrome.

KIAA0586 variants have been associated with a wide range of ciliopathies, mainly Joubert syndrome (JS, OMIM #616490) and short-rib thoracic dysplasia syndrome (SRTD, OMIM #616546). However, the hypothesis that this gene is involved with hydrolethalus syndrome (HSL, OMIM #614120) and orofaciodigital syndrome IV (OMIM #258860) has already been raised. Ciliopathies' clinical features are often overlapped despite differing in phenotype severity. Besides KIAA0586, HYLS1 and KIF7 are also known for being causative of ciliopathies, indicating that all three genes may have similar or converging genomic pathways. Overall, the genotypic and phenotypic spectrum of ciliopathies becomes wider and conflicting while more and more new variants are added to this group of disorders' molecular pot. In this case report we discuss the first Brazilian individual clinically diagnosed with hydrolethalus syndrome and molecular findings that demonstrate the role of KIAA0586 as a causative gene of a group of genetic disorders. Also, recent reports on individuals with intronic and exonic variants combined leading to ciliopathies support our patient's molecular diagnosis. At the same time, we discuss variable expressivity and overlapping features in ciliopathies.

Retinal primary cilia and their dysfunction in retinal neurodegenerative diseases: beyond ciliopathies.

Primary cilia are sensory organelles that extend from the cellular membrane and are found in a wide range of cell types. Cilia possess a plethora of vital components that enable the detection and transmission of several signaling pathways, including Wnt and Shh. In turn, the regulation of ciliogenesis and cilium length is influenced by various factors, including autophagy, organization of the actin cytoskeleton, and signaling inside the cilium. Irregularities in the development, maintenance, and function of this cellular component lead to a range of clinical manifestations known as ciliopathies. The majority of people with ciliopathies have a high prevalence of retinal degeneration. The most common theory is that retinal degeneration is primarily caused by functional and developmental problems within retinal photoreceptors. The contribution of other ciliated retinal cell types to retinal degeneration has not been explored to date. In this review, we examine the occurrence of primary cilia in various retinal cell types and their significance in pathology. Additionally, we explore potential therapeutic approaches targeting ciliopathies. By engaging in this endeavor, we present new ideas that elucidate innovative concepts for the future investigation and treatment of retinal ciliopathies.

CilioGenics: an integrated method and database for predicting novel ciliary genes.

Uncovering the full list of human ciliary genes holds enormous promise for the diagnosis of cilia-related human diseases, collectively known as ciliopathies. Currently, genetic diagnoses of many ciliopathies remain incomplete (1-3). While various independent approaches theoretically have the potential to reveal the entire list of ciliary genes, approximately 30% of the genes on the ciliary gene list still stand as ciliary candidates (4,5). These methods, however, have mainly relied on a single strategy to uncover ciliary candidate genes, making the categorization challenging due to variations in quality and distinct capabilities demonstrated by different methodologies. Here, we develop a method called CilioGenics that combines several methodologies (single-cell RNA sequencing, protein-protein interactions (PPIs), comparative genomics, transcription factor (TF) network analysis, and text mining) to predict the ciliary capacity of each human gene. Our combined approach provides a CilioGenics score for every human gene that represents the probability that it will become a ciliary gene. Compared to methods that rely on a single method, CilioGenics performs better in its capacity to predict ciliary genes. Our top 500 gene list includes 258 new ciliary candidates, with 31 validated experimentally by us and others. Users may explore the whole list of human genes and CilioGenics scores on the CilioGenics database (https://ciliogenics.com/).

A case report of autosomal recessive polycystic kidney disease with noncompaction of ventricular myocardium: coincidence or different manifestations of ciliopathy?

BACKGROUND: Autosomal recessive polycystic kidney disease (ARPKD) is a rare inherited cystic disease characterized by bilateral renal cyst formation and congenital liver fibrosis. Cardiovascular disorders such as noncompaction of ventricular myocardium (NVM) have not been reported with ARPKD. CASE PRESENTATION: A 5-month-old girl was examined after presenting with a fever and turbid urine for one day and was diagnosed as urinary tract infection. Urinary ultrasound showed multiple round, small cysts varying in size in both kidneys. Genetic testing revealed two heterozygous mutations and one exon deletion in the polycystic kidney and hepatic disease 1 gene, indicating a diagnosis of ARPKD. During hospitalization, she was found to have chronic heart failure after respiratory tract infection, with an ejection fraction of 29% and fraction shortening of 13%. When the patient was 15 months old, it was found that she had prominent trabeculations and deep intertrabecular recesses with the appearance of blood flow from the ventricular cavity into the intertrabecular recesses by echocardiography. The noncompaction myocardium was 0.716 cm and compaction myocardium was 0.221 cm (N/C = 3.27), indicating a diagnosis of NVM. Liver and kidney function remained normal during four-year follow-up. CONCLUSIONS: This is the first report of NVM in a patient with ARPKD. It is unsure if the coexistence of NVM and ARPKD is a coincidence or they are different manifestations of ciliary dysfunction in the heart and kidneys.

Biallelic loss-of-function variants in the centriolar protein CCP110 leads to a ciliopathy-like phenotype.

CCP110 (centriolar coiled coil protein 110, also known as CP110) is one of the essential proteins localized in the centrosome that plays critical roles in the regulation of the cell cycle and also in the initiation of ciliogenesis. So far, no human congenital disorders have been identified to be associated with pathogenic variants of CCP110. Mice with biallelic loss-of-function variants of Ccp110 (Ccp110-/-) are known to manifest multiple organ defects, including a small body size, polydactyly, omphalocele, congenital heart defects, cleft palate, short ribs, and a small thoracic cage, a pattern of abnormalities closely resembling that in "ciliopathies" in humans. Herein, we report a 7-month-old male infant who presented with growth failure and skeletal abnormalities, including a narrow thorax and severe brachydactyly. Trio exome analysis of the genomic DNA of the patient and his parents showed that the patient was a compound heterozygote for truncating variants of CCP110, including a frameshift variant NM_001323572.2:c.856_857del, p.(Val286Leufs*5) inherited from the father, and a nonsense variant NM_001323572.2:c.1129C>T, p.(Arg377*) inherited from the mother. The strikingly similar pattern of malformations between Ccp110-/- mice and the 7-month-old male infant reported herein carrying unequivocal truncating CCP110 variants strongly supports the contention that CCP110 is a novel disease-causative gene.

CEP41, a ciliopathy-linked centrosomal protein, regulates microtubule assembly and cell proliferation.

Centrosomal proteins play pivotal roles in orchestrating microtubule dynamics, and their dysregulation leads to disorders, including cancer and ciliopathies. Understanding the multifaceted roles of centrosomal proteins is vital to comprehend their involvement in disease development. Here, we report novel cellular functions of CEP41, a centrosomal and ciliary protein implicated in Joubert syndrome. We show that CEP41 is an essential microtubule-associated protein with microtubule-stabilizing activity. Purified CEP41 binds to preformed microtubules, promotes microtubule nucleation and suppresses microtubule disassembly. When overexpressed in cultured cells, CEP41 localizes to microtubules and promotes microtubule bundling. Conversely, shRNA-mediated knockdown of CEP41 disrupts the interphase microtubule network and delays microtubule reassembly, emphasizing its role in microtubule organization. Further, we demonstrate that the association of CEP41 with microtubules relies on its conserved rhodanese homology domain (RHOD) and the N-terminal region. Interestingly, a disease-causing mutation in the RHOD domain impairs CEP41-microtubule interaction. Moreover, depletion of CEP41 inhibits cell proliferation and disrupts cell cycle progression, suggesting its potential involvement in cell cycle regulation. These insights into the cellular functions of CEP41 hold promise for unraveling the impact of its mutations in ciliopathies.

Biallelic Variants in MNS1 Are Associated with Laterality Defects and Respiratory Involvement.

Defects in motile cilia, termed motile ciliopathies, result in clinical manifestations affecting the respiratory and reproductive system, as well as laterality defects and hydrocephalus. We previously defined biallelic MNS1 variants causing situs inversus and male infertility, mirroring the findings in Mns1-/- mice. Here, we present clinical and genomic findings in five newly identified individuals from four unrelated families affected by MNS1-related disorder. Ciliopathy panel testing and whole exome sequencing identified one previously reported and two novel MNS1 variants extending the genotypic spectrum of disease. A broad spectrum of laterality defects including situs inversus totalis and heterotaxia was confirmed. Interestingly, a single affected six-year-old girl homozygous for an MNS1 nonsense variant presented with a history of neonatal respiratory distress syndrome, recurrent respiratory tract infections, chronic rhinitis, and wet cough. Accordingly, immunofluorescence analysis showed the absence of MNS1 from the respiratory epithelial cells of this individual. Two other individuals with hypomorphic variants showed laterality defects and mild respiratory phenotype. This study represents the first observation of heterotaxia and respiratory disease in individuals with biallelic MNS1 variants, an important extension of the phenotype associated with MNS1-related motile ciliopathy disorder.

Genome sequencing identifies biallelic variants in SCLT1 in a patient with syndromic nephronophthisis: Reflections on the SCLT1-related ciliopathy spectrum.

Ciliopathies represent a major category of rare multisystem disease. Arriving at a specific diagnosis for a given patient is challenged by the significant genetic and clinical heterogeneity of these conditions. We report the outcome of the diagnostic odyssey of a child with obesity, renal, and retinal disease. Genome sequencing identified biallelic splice site variants in sodium channel and clathrin linker 1 (SCLT1), an emerging ciliopathy gene. We review the literature on all patients reported with biallelic SCLT1 variants highlighting a frequent clinical presentation that overlaps Bardet-Biedl and Senior-Loken syndromes. We also discuss current concepts in syndrome designation in light of these data.

Defects in diffusion barrier function of ciliary transition zone caused by ciliopathy variations of TMEM218.

Primary cilia are antenna-like structures protruding from the surface of various eukaryotic cells, and have distinct protein compositions in their membranes. This distinct protein composition is maintained by the presence of the transition zone (TZ) at the ciliary base, which acts as a diffusion barrier between the ciliary and plasma membranes. Defects in cilia and the TZ are known to cause a group of disorders collectively called the ciliopathies, which demonstrate a broad spectrum of clinical features, such as perinatally lethal Meckel syndrome (MKS), relatively mild Joubert syndrome (JBTS), and nonsyndromic nephronophthisis (NPHP). Proteins constituting the TZ can be grouped into the MKS and NPHP modules. The MKS module is composed of several transmembrane proteins and three soluble proteins. TMEM218 was recently reported to be mutated in individuals diagnosed as MKS and JBTS. However, little is known about how TMEM218 mutations found in MKS and JBTS affect the functions of cilia. In this study, we found that ciliary membrane proteins were not localized to cilia in TMEM218-knockout cells, indicating impaired barrier function of the TZ. Furthermore, the exogenous expression of JBTS-associated TMEM218 variants but not MKS-associated variants in TMEM218-knockout cells restored the localization of ciliary membrane proteins. In particular, when expressed in TMEM218-knockout cells, the TMEM218(R115H) variant found in JBTS was able to restore the barrier function of cells, whereas the MKS variant TMEM218(R115C) could not. Thus, the severity of symptoms of MKS and JBTS individuals appears to correlate with the degree of their ciliary defects at the cellular level.

ZFYVE19 deficiency: a ciliopathy involving failure of cell division, with cell death.

BACKGROUND AND AIMS: Variants in ZFYVE19 underlie a disorder characterised by progressive portal fibrosis, portal hypertension and eventual liver decompensation. We aim to create an animal model to elucidate the pathogenic mechanism. METHODS: Zfyve19 knockout (Zfyve19-/- ) mice were generated and exposed to different liver toxins. Their livers were characterised at the tissue, cellular and molecular levels. Findings were compared with those in wild-type mice and in ZFYVE19-deficient patients. ZFYVE19 knockout and knockdown retinal pigment epithelial-1 cells and mouse embryonic fibroblasts were generated to study cell division and cell death. RESULTS: The Zfyve19-/- mice were normal overall, particularly with respect to hepatobiliary features. However, when challenged with α-naphthyl isothiocyanate, Zfyve19-/- mice developed changes resembling those in ZFYVE19-deficient patients, including elevated serum liver injury markers, increased numbers of bile duct profiles with abnormal cholangiocyte polarity and biliary fibrosis. Failure of cell division, centriole and cilia abnormalities, and increased cell death were observed in knockdown/knockout cells. Increased cell death and altered mRNA expression of cell death-related signalling pathways was demonstrated in livers from Zfyve19-/- mice and patients. Transforming growth factor-ß (TGF-ß) and Janus kinase-Signal Transducer and Activator of Transcription 3 (JAK-STAT3) signalling pathways were upregulated in vivo, as were chemokines such as C-X-C motif ligands 1, 10 and 12. CONCLUSIONS: Our findings demonstrated that ZFYVE19 deficiency is a ciliopathy with novel histological features. Failure of cell division with ciliary abnormalities and cell death activates macrophages and may thus lead to biliary fibrosis via TGF-ß pathway in the disease.