Exploring the diverse clinical and variant spectrum of CEP78-associated syndrome: Novel pathogenic variants identified in a case series.

Dual sensory impairment, commonly referred to as combined hearing and vision loss, can stem from a diverse spectrum of conditions, each presenting with its unique set of clinical characteristics. Our understanding of dual sensory impairment has expanded significantly in the past decade, broadening the scope of genetic differential diagnoses, including genes such as CEP250, ARSG, TUBB4B, CEP78, and ABHD12. A case series including three patients from two families with genetically diagnosed CEP78-associated cone-rod dystrophy was identified. We collected and reviewed their clinical records, imaging data, and genetic testing results. In addition, a comprehensive literature review was conducted on the phenotype and the genetic testing modality employed in all published CEP78 cases through a PubMed search using the keyword "CEP78." A retinal dystrophy panel detected a novel homozygous CEP78 pathogenic variant (c.1447C>T, p.Arg483*) in siblings-Cases 1 and 2-from Family 1. Both teenagers have a clinical diagnosis of cone-rod dystrophy with presumed normal hearing. Case 3 from Family 2, diagnosed with cone-rod dystrophy and early-onset hearing loss, was found to carry a CEP78 pathogenic variant (c.1206-2A>C) and a likely pathogenic variant (c.856_857del, p.Leu286Glyfs*12) also through panel-based genetic testing. Intriguingly, neither of these variants was reported in an affected sibling's clinical whole-exome sequencing (WES) report when performed in 2015. A review of CEP78-related literature unveiled that the initial report linking CEP78 to cone-rod dystrophy and hearing loss was published in September 2016. Any pathogenic variant found in CEP78 before 2016 would have been categorized as a "clearly disruptive variant in a gene of uncertain significance (GUS)" and might not have been reported in the WES report. It is important to acknowledge that our understanding of genotype-phenotype associations is undergoing rapid expansion. It is also crucial to recognize that repeat genetic testing may represent a fundamentally different approach, given the technological advancements not only in the coverage of the sequencing but also in the more comprehensive understanding of genotype-phenotype associations. This case series also enriches the existing CEP78 literature by providing phenotypic details of the youngest case of CEP78-associated retinopathy reported in the literature (Case 2), which expands our perspective on the natural history of disease in this disorder.

Functional characterization of the first missense variant in CEP78, a founder allele associated with cone-rod dystrophy, hearing loss, and reduced male fertility.

Inactivating variants in the centrosomal CEP78 gene have been found in cone-rod dystrophy with hearing loss (CRDHL), a particular phenotype distinct from Usher syndrome. Here, we identified and functionally characterized the first CEP78 missense variant c.449T>C, p.(Leu150Ser) in three CRDHL families. The variant was found in a biallelic state in two Belgian families and in a compound heterozygous state-in trans with c.1462-1G>T-in a third German family. Haplotype reconstruction showed a founder effect. Homology modeling revealed a detrimental effect of p.(Leu150Ser) on protein stability, which was corroborated in patients' fibroblasts. Elongated primary cilia without clear ultrastructural abnormalities in sperm or nasal brushes suggest impaired cilia assembly. Two affected males from different families displayed sperm abnormalities causing infertility. One of these is a heterozygous carrier of a complex allele in SPAG17, a ciliary gene previously associated with autosomal recessive male infertility. Taken together, our data indicate that a missense founder allele in CEP78 underlies the same sensorineural CRDHL phenotype previously associated with inactivating variants. Interestingly, the CEP78 phenotype has been possibly expanded with male infertility. Finally, CEP78 loss-of-function variants may have an underestimated role in misdiagnosed Usher syndrome, with or without sperm abnormalities.

HIV-1 Vpr hijacks EDD-DYRK2-DDB1DCAF1 to disrupt centrosome homeostasis.

Viruses exploit the host cell machinery for their own profit. To evade innate immune sensing and promote viral replication, HIV type 1 (HIV-1) subverts DNA repair regulatory proteins and induces G2/M arrest. The preintegration complex of HIV-1 is known to traffic along microtubules and accumulate near the microtubule-organizing center. The centrosome is the major microtubule-organizing center in most eukaryotic cells, but precisely how HIV-1 impinges on centrosome biology remains poorly understood. We report here that the HIV-1 accessory protein viral protein R (Vpr) localized to the centrosome through binding to DCAF1, forming a complex with the ubiquitin ligase EDD-DYRK2-DDB1DCAF1 and Cep78, a resident centrosomal protein previously shown to inhibit EDD-DYRK2-DDB1DCAF1 Vpr did not affect ubiquitination of Cep78. Rather, it enhanced ubiquitination of an EDD-DYRK2-DDB1DCAF1 substrate, CP110, leading to its degradation, an effect that could be overcome by Cep78 expression. The down-regulation of CP110 and elongation of centrioles provoked by Vpr were independent of G2/M arrest. Infection of T lymphocytes with HIV-1, but not with HIV-1 lacking Vpr, promoted CP110 degradation and centriole elongation. Elongated centrioles recruited more γ-tubulin to the centrosome, resulting in increased microtubule nucleation. Our results suggest that Vpr is targeted to the centrosome where it hijacks a ubiquitin ligase, disrupting organelle homeostasis, which may contribute to HIV-1 pathogenesis.

Cep78 controls centrosome homeostasis by inhibiting EDD-DYRK2-DDB1VprBP.

The centrosome plays a critical role in various cellular processes including cell division and cilia formation, and deregulation of centrosome homeostasis is a hallmark feature of many human diseases. Here, we show that centrosomal protein of 78 kDa (Cep78) localizes to mature centrioles and directly interacts with viral protein R binding protein (VprBP). Although VprBP is a component of two distinct E3 ubiquitin ligases, EDD-DYRK2-DDB1VprBP and CRL4VprBP, Cep78 binds specifically to EDD-DYRK2-DDB1VprBP and inhibits its activity. A pool of EDD-DYRK2-DDB1VprBP is active at the centrosome and mediates ubiquitination of CP110, a novel centrosomal substrate. Deregulation of Cep78 or EDD-DYRK2-DDB1VprBP perturbs CP110 ubiquitination and protein stability, thereby affecting centriole length and cilia assembly. Mechanistically, ubiquitination of CP110 entails its phosphorylation by DYRK2 and binding to VprBP Cep78 specifically impedes the transfer of ubiquitin from EDD to CP110 without affecting CP110 phosphorylation and binding to VprBP Thus, we identify Cep78 as a new player that regulates centrosome homeostasis by inhibiting the final step of the enzymatic reaction catalyzed by EDD-DYRK2-DDB1VprBP.

Cep78 is a new centriolar protein involved in Plk4-induced centriole overduplication.

Centrioles are core components of centrosomes, the major microtubule-organizing centers of animal cells, and act as basal bodies for cilia formation. Control of centriole number is therefore crucial for genome stability and embryogenesis. Centriole duplication requires the serine/threonine protein kinase Plk4. Here, we identify Cep78 as a human centrosomal protein and a new interaction partner of Plk4. Cep78 is mainly a centriolar protein that localizes to the centriolar wall. Furthermore, we find that Plk4 binds to Cep78 through its N-terminal domain but that Cep78 is not an in vitro Plk4 substrate. Cep78 colocalizes with Plk4 at centrioles and is required for Plk4-induced centriole overduplication. Interestingly, upon depletion of Cep78, newly synthesized Plk4 is not localized to centrosomes. Our results suggest that the interaction between Cep78 and the N-terminal catalytic domain of Plk4 is a new and important element in the centrosome overduplication process.

CEP78 is mutated in a distinct type of Usher syndrome.

BACKGROUND: Usher syndrome is a genetically heterogeneous disorder featured by combined visual impairment and hearing loss. Despite a dozen of genes involved in Usher syndrome having been identified, the genetic basis remains unknown in 20-30% of patients. In this study, we aimed to identify the novel disease-causing gene of a distinct subtype of Usher syndrome. METHODS: Ophthalmic examinations and hearing tests were performed on patients with Usher syndrome in two consanguineous families. Target capture sequencing was initially performed to screen causative mutations in known retinal disease-causing loci. Whole exome sequencing (WES) and whole genome sequencing (WGS) were applied for identifying novel disease-causing genes. RT-PCR and Sanger sequencing were performed to evaluate the splicing-altering effect of identified CEP78 variants. RESULTS: Patients from the two independent families show a mild Usher syndrome phenotype featured by juvenile or adult-onset cone-rod dystrophy and sensorineural hearing loss. WES and WGS identified two homozygous rare variants that affect mRNA splicing of a ciliary gene CEP78. RT-PCR confirmed that the two variants indeed lead to abnormal splicing, resulting in premature stop of protein translation due to frameshift. CONCLUSIONS: Our results provide evidence that CEP78 is a novel disease-causing gene for Usher syndrome, demonstrating an additional link between ciliopathy and Usher protein network in photoreceptor cells and inner ear hair cells.

Absence of CEP78 causes photoreceptor and sperm flagella impairments in mice and a human individual.

Cone-rod dystrophy (CRD) is a genetically inherited retinal disease that can be associated with male infertility, while the specific genetic mechanisms are not well known. Here, we report CEP78 as a causative gene of a particular syndrome including CRD and male infertility with multiple morphological abnormalities of sperm flagella (MMAF) both in human and mouse. Cep78 knockout mice exhibited impaired function and morphology of photoreceptors, typified by reduced ERG amplitudes, disrupted translocation of cone arrestin, attenuated and disorganized photoreceptor outer segments (OS) disks and widen OS bases, as well as interrupted connecting cilia elongation and abnormal structures. Cep78 deletion also caused male infertility and MMAF, with disordered '9+2' structure and triplet microtubules in sperm flagella. Intraflagellar transport (IFT) proteins IFT20 and TTC21A are identified as interacting proteins of CEP78. Furthermore, CEP78 regulated the interaction, stability, and centriolar localization of its interacting protein. Insufficiency of CEP78 or its interacting protein causes abnormal centriole elongation and cilia shortening. Absence of CEP78 protein in human caused similar phenotypes in vision and MMAF as Cep78-/- mice. Collectively, our study supports the important roles of CEP78 defects in centriole and ciliary dysfunctions and molecular pathogenesis of such multi-system syndrome.

A novel frameshift variant in CEP78 associated with nonsyndromic retinitis pigmentosa, and a review of CEP78-related phenotypes.

BACKGROUND: Pathogenic variants in the CEP78 gene can present as atypical Usher syndrome or as retinitis pigmentosa. Here, we present a review of all reported cases of CEP78 variants in the literature to date and present a novel variant of CEP78, c.1261_1262delinsA, in a consanguineous northern Finnish family with two individuals. MATERIALS AND METHODS: Our patients were first discovered in a registry-based study. Later, they gave their written consent for this study. In order to describe the genotype and phenotype, their historic clinical patient data and genetic data were gathered, and a clinical ophthalmic examination and an audiogram were performed. For this review, a PubMed search using the keyword CEP78 was carried out. The first article on CEP78 was published in the year 2007, and the publications from the years 2007-2021 were included. RESULTS: A large gene panel identified a homozygous CEP78 c.1261_1262delinsA variant in two affected siblings. In addition to the classical signs of retinitis pigmentosa, both siblings had large round atrophic spots in the mid periphery, and hyperautofluorescence of the macula. Patient 1 had age-related hearing impairment; patient 2 had normal hearing. In total, 20 articles have been published about CEP78. Eight of these papers report patient data with the affected individuals typically having retinal dystrophy combined with sensorineural hearing impairment, classified as atypical Usher syndrome. CONCLUSIONS: Here, we present a comprehensive review of CEP78 and expand the knowledge of pathogenic CEP78 variants and the phenotypic variety.

Expanding the clinical phenotype in patients with disease causing variants associated with atypical Usher syndrome.

Atypical Usher syndrome (USH) is poorly defined with a broad clinical spectrum. Here, we characterize the clinical phenotype of disease caused by variants in CEP78, CEP250, ARSG, and ABHD12.Chart review evaluating demographic, clinical, imaging, and genetic findings of 19 patients from 18 families with a clinical diagnosis of retinal disease and confirmed disease-causing variants in CEP78, CEP250, ARSG, or ABHD12.CEP78-related disease included sensorineural hearing loss (SNHL) in 6/7 patients and demonstrated a broad phenotypic spectrum including: vascular attenuation, pallor of the optic disc, intraretinal pigment, retinal pigment epithelium mottling, areas of mid-peripheral hypo-autofluorescence, outer retinal atrophy, mild pigmentary changes in the macula, foveal hypo-autofluorescence, and granularity of the ellipsoid zone. Nonsense and frameshift variants in CEP250 showed mild retinal disease with progressive, non-congenital SNHL. ARSG variants resulted in a characteristic pericentral pattern of hypo-autofluorescence with one patient reporting non-congenital SNHL. ABHD12-related disease showed rod-cone dystrophy with macular involvement, early and severe decreased best corrected visual acuity, and non-congenital SNHL ranging from unreported to severe.This study serves to expand the clinical phenotypes of atypical USH. Given the variable findings, atypical USH should be considered in patients with peripheral and macular retinal disease even without the typical RP phenotype especially when SNHL is noted. Additionally, genetic screening may be useful in patients who have clinical symptoms and retinal findings even in the absence of known SNHL given the variability of atypical USH.

CEP78 functions downstream of CEP350 to control biogenesis of primary cilia by negatively regulating CP110 levels.

CEP78 is a centrosomal protein implicated in ciliogenesis and ciliary length control, and mutations in the CEP78 gene cause retinal cone-rod dystrophy associated with hearing loss. However, the mechanism by which CEP78 affects cilia formation is unknown. Based on a recently discovered disease-causing CEP78 p.L150S mutation, we identified the disease-relevant interactome of CEP78. We confirmed that CEP78 interacts with the EDD1-DYRK2-DDB1VPRBP E3 ubiquitin ligase complex, which is involved in CP110 ubiquitination and degradation, and identified a novel interaction between CEP78 and CEP350 that is weakened by the CEP78L150S mutation. We show that CEP350 promotes centrosomal recruitment and stability of CEP78, which in turn leads to centrosomal recruitment of EDD1. Consistently, cells lacking CEP78 display significantly increased cellular and centrosomal levels of CP110, and depletion of CP110 in CEP78-deficient cells restored ciliation frequency to normal. We propose that CEP78 functions downstream of CEP350 to promote ciliogenesis by negatively regulating CP110 levels via an EDD1-dependent mechanism.

Long-Read Sequencing to Unravel Complex Structural Variants of CEP78 Leading to Cone-Rod Dystrophy and Hearing Loss.

Inactivating variants as well as a missense variant in the centrosomal CEP78 gene have been identified in autosomal recessive cone-rod dystrophy with hearing loss (CRDHL), a rare syndromic inherited retinal disease distinct from Usher syndrome. Apart from this, a complex structural variant (SV) implicating CEP78 has been reported in CRDHL. Here we aimed to expand the genetic architecture of typical CRDHL by the identification of complex SVs of the CEP78 region and characterization of their underlying mechanisms. Approaches used for the identification of the SVs are shallow whole-genome sequencing (sWGS) combined with quantitative polymerase chain reaction (PCR) and long-range PCR, or ExomeDepth analysis on whole-exome sequencing (WES) data. Targeted or whole-genome nanopore long-read sequencing (LRS) was used to delineate breakpoint junctions at the nucleotide level. For all SVs cases, the effect of the SVs on CEP78 expression was assessed using quantitative PCR on patient-derived RNA. Apart from two novel canonical CEP78 splice variants and a frameshifting single-nucleotide variant (SNV), two SVs affecting CEP78 were identified in three unrelated individuals with CRDHL: a heterozygous total gene deletion of 235 kb and a partial gene deletion of 15 kb in a heterozygous and homozygous state, respectively. Assessment of the molecular consequences of the SVs on patient's materials displayed a loss-of-function effect. Delineation and characterization of the 15-kb deletion using targeted LRS revealed the previously described complex CEP78 SV, suggestive of a recurrent genomic rearrangement. A founder haplotype was demonstrated for the latter SV in cases of Belgian and British origin, respectively. The novel 235-kb deletion was delineated using whole-genome LRS. Breakpoint analysis showed microhomology and pointed to a replication-based underlying mechanism. Moreover, data mining of bulk and single-cell human and mouse transcriptional datasets, together with CEP78 immunostaining on human retina, linked the CEP78 expression domain with its phenotypic manifestations. Overall, this study supports that the CEP78 locus is prone to distinct SVs and that SV analysis should be considered in a genetic workup of CRDHL. Finally, it demonstrated the power of sWGS and both targeted and whole-genome LRS in identifying and characterizing complex SVs in patients with ocular diseases.

Significance of CEP78 and WDR62 gene expressions in differentiated thyroid carcinoma: Possible predictors of lateral lymph node metastasis.

OBJECTIVES: This study aimed at investigating the clinical significance of CEP78 and WDR62 in differentiated thyroid carcinoma (DTC). This study also aimed at finding predictors that help in detecting patients with DTC who have high risk for lateral lymph node metastasis (LNM). METHODS: Quantitative real-time polymerase chain reaction (RT-qPCR) was performed to examine CEP78, and WDR62 mRNA expressions in 40 tissue specimens of DTC, and 40 goiter tissue specimens. Additionally, we reviewed clinical, ultrasound, laboratory, pathological data of patients to analyze the associations between these characteristics and lateral LNM. RESULTS: Our results demonstrated that relative CEP78 mRNA levels were significantly decreased in thyroid cancer tissues than goiter tissues (P = 0.002). ROC curve analysis confirmed the diagnostic value of CEP78 mRNA expression, providing an AUC equals to 0.698 (95% confidence intervals (CI), 0.583-0.813; P = 0.002). The relative WDR62 mRNA expression was not statistically different in DTC tissues and goiter tissues (P = 0.686). Furthermore, the DTC patients had been included to examine risk factors for lateral LNM. In multivariate analysis, the significant factors for predicting lateral LNM were low CEP78 mRNA expression (cut off value ≤0.54; P = 0.03; OR = 19.62; 95% CI, 1.3-296.23), central LNM (P = 0.011; OR = 33.6; 95% CI, 2.24-503.6) and calcifications (P = 0.023; OR = 27.187; 95% CI, 1.57-469.5). CONCLUSIONS: CEP78 can be used as a promising molecular biomarker for differentiation between DTC and goiter tissues, in addition it might serve as a predictor of lateral LNM in DTC along with central LNM and calcifications. Unlike CEP78, WDR62 mRNA expression was not statistically different in DTC and goiter.

Complex structural variants in Mendelian disorders: identification and breakpoint resolution using short- and long-read genome sequencing.

BACKGROUND: Studies have shown that complex structural variants (cxSVs) contribute to human genomic variation and can cause Mendelian disease. We aimed to identify cxSVs relevant to Mendelian disease using short-read whole-genome sequencing (WGS), resolve the precise variant configuration and investigate possible mechanisms of cxSV formation. METHODS: We performed short-read WGS and analysis of breakpoint junctions to identify cxSVs in a cohort of 1324 undiagnosed rare disease patients. Long-read WGS and gene expression analysis were used to resolve one case. RESULTS: We identified three pathogenic cxSVs: a de novo duplication-inversion-inversion-deletion affecting ARID1B, a de novo deletion-inversion-duplication affecting HNRNPU and a homozygous deletion-inversion-deletion affecting CEP78. Additionally, a de novo duplication-inversion-duplication overlapping CDKL5 was resolved by long-read WGS demonstrating the presence of both a disrupted and an intact copy of CDKL5 on the same allele, and gene expression analysis showed both parental alleles of CDKL5 were expressed. Breakpoint analysis in all the cxSVs revealed both microhomology and longer repetitive elements. CONCLUSIONS: Our results corroborate that cxSVs cause Mendelian disease, and we recommend their consideration during clinical investigations. We show that resolution of breakpoints can be critical to interpret pathogenicity and present evidence of replication-based mechanisms in cxSV formation.

Low expression of centrosomal protein 78 (CEP78) is associated with poor prognosis of colorectal cancer patients.

BACKGROUND: Centrosomal protein 78 (CEP78) has been characterized as a component of the centrosome required for the regulation of centrosome-related events during the cell cycle, but its role in human cancers remains unclear. This study aimed to investigate the role and the clinical value of CEP78 in colorectal cancer (CRC). METHODS: Quantitative real-time polymerase chain reaction (qRT-PCR) and immunohistochemistry were performed to examine CEP78 expression in CRC tissues and adjacent noncancerous tissues. The association between CEP78 expression and clinical outcomes of CRC patients was determined. The effect of CEP78 on cell growth was examined in vitro by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) assay, colony formation, and flow cytometry assays and in vivo using a nude mouse model. RESULTS: The expression level of CEP78 was significantly lower in tumor tissues than in the adjacent normal tissues (P < 0.01). Low CEP78 expression was significantly associated with poor differentiation (P = 0.003), large tumor size (P = 0.017), lymphatic metastasis (P = 0.034), distant metastasis (P = 0.029), and advanced stage (P = 0.011). Kaplan-Meier analysis indicated that patients with low CEP78 expression had shorter survival than those with high CEP78 expression (P < 0.01). Overexpression of CEP78 in CRC cells significantly reduced cell viability and colony formation in vitro and halted tumor growth in vivo. Further study showed that CEP78 reintroduction in CRC cells resulted in G2/M phase arrest rather than cell apoptosis. CONCLUSIONS: CEP78 might function as a tumor suppressor and serve as a novel prognostic marker in CRC.