scholarly journals Pcdp1 is a central apparatus protein that binds Ca2+-calmodulin and regulates ciliary motility

2010 ◽  
Vol 189 (3) ◽  
pp. 601-612 ◽  
Author(s):  
Christen G. DiPetrillo ◽  
Elizabeth F. Smith
Keyword(s):  
Primary Ciliary Dyskinesia ◽  
Calcium Concentration ◽  
Beat Frequency ◽  
Calcium Sensor ◽  
Central Pair ◽  
Ciliary Motility ◽  
Significant Impairment ◽  
Central Apparatus ◽  
Cilia And Flagella ◽  
Ciliary Dyskinesia

For all motile eukaryotic cilia and flagella, beating is regulated by changes in intraciliary calcium concentration. Although the mechanism for calcium regulation is not understood, numerous studies have shown that calmodulin (CaM) is a key axonemal calcium sensor. Using anti-CaM antibodies and Chlamydomonas reinhardtii axonemal extracts, we precipitated a complex that includes four polypeptides and that specifically interacts with CaM in high [Ca2+]. One of the complex members, FAP221, is an orthologue of mammalian Pcdp1 (primary ciliary dyskinesia protein 1). Both FAP221 and mammalian Pcdp1 specifically bind CaM in high [Ca2+]. Reduced expression of Pcdp1 complex members in C. reinhardtii results in failure of the C1d central pair projection to assemble and significant impairment of motility including uncoordinated bends, severely reduced beat frequency, and altered waveforms. These combined results reveal that the central pair Pcdp1 (FAP221) complex is essential for control of ciliary motility.

scholarly journals The Pcdp1 complex coordinates the activity of dynein isoforms to produce wild-type ciliary motility

2011 ◽  
Vol 22 (23) ◽  
pp. 4527-4538 ◽  
Author(s):  
Christen G. DiPetrillo ◽  
Elizabeth F. Smith
Keyword(s):  
Primary Ciliary Dyskinesia ◽  
Beat Frequency ◽  
Structural Approach ◽  
Wild Type ◽  
Unique Role ◽  
Ciliary Beating ◽  
Ciliary Motility ◽  
Central Apparatus ◽  
Dynein Arms ◽  
Ciliary Dyskinesia

Generating the complex waveforms characteristic of beating cilia requires the coordinated activity of multiple dynein isoforms anchored to the axoneme. We previously identified a complex associated with the C1d projection of the central apparatus that includes primary ciliary dyskinesia protein 1 (Pcdp1). Reduced expression of complex members results in severe motility defects, indicating that C1d is essential for wild-type ciliary beating. To define a mechanism for Pcdp1/C1d regulation of motility, we took a functional and structural approach combined with mutants lacking C1d and distinct subsets of dynein arms. Unlike mutants completely lacking the central apparatus, dynein-driven microtubule sliding velocities are wild type in C1d- defective mutants. However, coordination of dynein activity among microtubule doublets is severely disrupted. Remarkably, mutations in either outer or inner dynein arm restore motility to mutants lacking C1d, although waveforms and beat frequency differ depending on which isoform is mutated. These results define a unique role for C1d in coordinating the activity of specific dynein isoforms to control ciliary motility.

scholarly journals Deep phenotyping, including quantitative ciliary beating parameters, and extensive genotyping in primary ciliary dyskinesia

2019 ◽  
Vol 57 (4) ◽  
pp. 237-244 ◽  
Author(s):  
Sylvain Blanchon ◽  
Marie Legendre ◽  
Mathieu Bottier ◽  
Aline Tamalet ◽  
Guy Montantin ◽  
...  
Keyword(s):  
Primary Ciliary Dyskinesia ◽  
High Speed ◽  
Genetic Disorder ◽  
Beat Frequency ◽  
Ciliary Beat Frequency ◽  
Central Complex ◽  
Ciliary Beating ◽  
Recovery Stroke ◽  
Biallelic Mutations ◽  
Ciliary Dyskinesia

BackgroundPrimary ciliary dyskinesia (PCD) is a rare genetic disorder resulting in abnormal ciliary motility/structure, extremely heterogeneous at genetic and ultrastructural levels. We aimed, in light of extensive genotyping, to identify specific and quantitative ciliary beating anomalies, according to the ultrastructural phenotype.MethodsWe prospectively included 75 patients with PCD exhibiting the main five ultrastructural phenotypes (n=15/group), screened all corresponding PCD genes and measured quantitative beating parameters by high-speed video-microscopy (HSV).ResultsSixty-eight (91%) patients carried biallelic mutations. Combined outer/inner dynein arms (ODA/IDA) defect induces total ciliary immotility, regardless of the gene involved. ODA defect induces a residual beating with dramatically low ciliary beat frequency (CBF) related to increased recovery stroke and pause durations, especially in case of DNAI1 mutations. IDA defect with microtubular disorganisation induces a low percentage of beating cilia with decreased beating angle and, in case of CCDC39 mutations, a relatively conserved mean CBF with a high maximal CBF. Central complex defect induces nearly normal beating parameters, regardless of the gene involved, and a gyrating motion in a minority of ciliated edges, especially in case of RSPH1 mutations. PCD with normal ultrastructure exhibits heterogeneous HSV values, but mostly an increased CBF with an extremely high maximal CBF.ConclusionQuantitative HSV analysis in PCD objectives beating anomalies associated with specific ciliary ultrastructures and genotypes. It represents a promising approach to guide the molecular analyses towards the best candidate gene(s) to be analysed or to assess the pathogenicity of the numerous sequence variants identified by next-generation-sequencing.

Incidence of Primary Ciliary Dyskinesia in Children with Recurrent Respiratory Diseases

1997 ◽  
Vol 106 (10) ◽  
pp. 854-858 ◽  
Author(s):  
André Coste ◽  
Marie-Claude Millepied ◽  
Catherine Chapelin ◽  
Philippe Reinert ◽  
Françoise Poron ◽  
...  
Keyword(s):  
Primary Ciliary Dyskinesia ◽  
Respiratory Tract Infections ◽  
Beat Frequency ◽  
Ciliary Beat Frequency ◽  
Systematic Investigation ◽  
Ciliated Cells ◽  
Tract Infections ◽  
Ciliary Dyskinesia ◽  
Ciliary Ultrastructure ◽  
Recurrent Respiratory Tract Infections

The goal of the study was to evaluate the incidence of primary ciliary dyskinesia (PCD) in children suffering from recurrent respiratory tract infections (RRIs) by means of a noninvasive method. Respiratory ciliated cells were collected by nasal brushing in 118 children (4.6 ± 2.5 years) with RRIs. The ciliary beat frequency (CBF) was measured with a stroboscopic method, and when the CBF was abnormal, the ciliary ultrastructure was analyzed by a quantitative method. The CBF could be measured in 106 patients (90%) and was abnormal in 15 patients. The ciliary ultrastructure was found to be abnormal in 11 of 15 patients: PCD was diagnosed in 6 cases, and acquired ciliary defects were observed in the remaining 5 patients. Our conclusion, that PCD is rare but not exceptional (5.6%) in children with RRIs, justifies the systematic investigation of ciliated cells in such patients. For this purpose, nasal brushing can be used to sample ciliated cells even in young children.

Mutation of serine/threonine protein kinase 36 ( STK36 ) causes primary ciliary dyskinesia with a central pair defect

2017 ◽  
Vol 38 (8) ◽  
pp. 964-969 ◽  
Author(s):  
Christine Edelbusch ◽  
Sandra Cindrić ◽  
Gerard W. Dougherty ◽  
Niki T. Loges ◽  
Heike Olbrich ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Primary Ciliary Dyskinesia ◽  
Central Pair ◽  
Pair Defect ◽  
Ciliary Dyskinesia

scholarly journals A novel hypomorphic allele of Spag17 causes primary ciliary dyskinesia phenotypes in mice

2020 ◽  
Vol 13 (10) ◽  
pp. dmm045344
Author(s):  
Zakia Abdelhamed ◽  
Marshall Lukacs ◽  
Sandra Cindric ◽  
Heymut Omran ◽  
Rolf W. Stottmann
Keyword(s):  
Cerebrospinal Fluid ◽  
Fluid Flow ◽  
Primary Ciliary Dyskinesia ◽  
Human Condition ◽  
Central Pair ◽  
Cerebrospinal Fluid Flow ◽  
Hypomorphic Allele ◽  
Motile Cilia ◽  
The Brain ◽  
Ciliary Dyskinesia

ABSTRACTPrimary ciliary dyskinesia (PCD) is a human condition of dysfunctional motile cilia characterized by recurrent lung infection, infertility, organ laterality defects and partially penetrant hydrocephalus. We recovered a mouse mutant from a forward genetic screen that developed many of the hallmark phenotypes of PCD. Whole-exome sequencing identified this primary ciliary dyskinesia only (Pcdo) allele to be a nonsense mutation (c.5236A>T) in the Spag17 coding sequence creating a premature stop codon (K1746*). The Pcdo variant abolished several isoforms of SPAG17 in the Pcdo mutant testis but not in the brain. Our data indicate differential requirements for SPAG17 in different types of motile cilia. SPAG17 is essential for proper development of the sperm flagellum and is required for either development or stability of the C1 microtubule structure within the central pair apparatus of the respiratory motile cilia, but not the brain ependymal cilia. We identified changes in ependymal ciliary beating frequency, but these did not appear to alter lateral ventricle cerebrospinal fluid flow. Aqueductal stenosis resulted in significantly slower and abnormally directed cerebrospinal fluid flow, and we suggest that this is the root cause of the hydrocephalus. The Spag17Pcdo homozygous mutant mice are generally viable to adulthood but have a significantly shortened lifespan, with chronic morbidity. Our data indicate that the c.5236A>T Pcdo variant is a hypomorphic allele of Spag17 that causes phenotypes related to motile, but not primary, cilia. Spag17Pcdo is a useful new model for elucidating the molecular mechanisms underlying central pair PCD pathogenesis in the mouse.This article has an associated First Person interview with the first author of the paper.

scholarly journals Higher throughput drug screening for rare respiratory diseases: Readthrough therapy in primary ciliary dyskinesia

2021 ◽  
pp. 2000455
Author(s):  
Dani Do Hyang Lee ◽  
Daniela Cardinale ◽  
Ersilia Nigro ◽  
Colin R. Butler ◽  
Andrew Rutman ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
High Throughput Screening ◽  
Primary Ciliary Dyskinesia ◽  
Respiratory Diseases ◽  
Beat Frequency ◽  
Screening Method ◽  
Respiratory Epithelium ◽  
Basal Cells ◽  
Translational Readthrough ◽  
Ciliary Dyskinesia

Development of therapeutic approaches for rare respiratory diseases is hampered by the lack of systems that allow medium-to-high-throughput screening of fully differentiated respiratory epithelium from affected patients. This is a particular problem for primary ciliary dyskinesia (PCD), a rare genetic disease caused by mutations in genes that adversely affect ciliary movement and consequently mucociliary transport. Primary cell culture of basal epithelial cells from nasal brush biopsies, followed by ciliated differentiation at air-liquid interface (ALI) has proven to be a useful tool in PCD diagnostics but the technique's broader utility, including in pre-clinical PCD research, has been restricted by the limited number of basal cells that it is possible to expand from such biopsies. Here, we describe an immunofluorescence screening method, enabled by extensive expansion of PCD patient basal cells and their culture into differentiated respiratory epithelium in miniaturised 96-well transwell format ALI cultures. Analyses of ciliary ultrastructure, beat pattern and beat frequency indicate that a range of different PCD defects can be retained in these cultures. As proof-of-principle, we performed a personalised investigation in a patient with a rare and severe form of PCD (reduced generation of motile cilia, RGMC), in this case caused by a homozygous nonsense mutation in the MCIDAS gene. The screening system allowed drugs that induce translational readthrough to be evaluated alone or in combination with nonsense-mediated decay inhibitors. Restoration of basal body formation in the patient's nasal epithelial cells was seen in vitro, suggesting a novel avenue for drug evaluation and development in PCD.

scholarly journals Structural insights into the cause of human RSPH4A primary ciliary dyskinesia

2021 ◽  
pp. mbc.E20-12-0806
Author(s):  
Yanhe Zhao ◽  
Justine Pinskey ◽  
Jianfeng Lin ◽  
Weining Yin ◽  
Patrick R. Sears ◽  
...  
Keyword(s):  
Primary Ciliary Dyskinesia ◽  
Electron Tomography ◽  
Three Dimensional ◽  
Dimensional Structure ◽  
Structural Basis ◽  
Radial Spoke ◽  
Radial Spokes ◽  
Cilia And Flagella ◽  
Tract Infections ◽  
Ciliary Dyskinesia

Cilia and flagella are evolutionarily conserved eukaryotic organelles involved in cell motility and signaling. In humans, mutations in Radial Spoke Head Protein 4 homolog A ( RSPH4A) can lead to primary ciliary dyskinesia (PCD), a life-shortening disease characterized by chronic respiratory tract infections, abnormal organ positioning, and infertility. Despite its importance for human health, the location of RSPH4A in human cilia has not been resolved, and the structural basis of RSPH4A-/- PCD remains elusive. Here, we present the native, three-dimensional structure of RSPH4A-/- human respiratory cilia using samples collected non-invasively from a PCD patient. Using cryo-electron tomography and subtomogram averaging, we compared the structures of control and RSPH4A-/- cilia, revealing primary defects in two of the three radial spokes (RSs) within the axonemal repeat and secondary (heterogeneous) defects in the central pair complex. Similar to RSPH1-/- cilia, the radial spoke heads of RS1 and RS2, but not RS3, were missing in RSPH4A-/- cilia. However, RSPH4A-/- cilia also exhibited defects within the arch domains adjacent to the RS1 and RS2 heads, which were not observed with RSPH1 loss. Our results provide insight into the underlying structural basis for RSPH4A-/- PCD and highlight the benefits of applying cryo-ET directly to patient samples for molecular structure determination. [Media: see text]

scholarly journals CiliarMove: new software for evaluating ciliary beat frequency helps find novel mutations by a Portuguese multidisciplinary team on primary ciliary dyskinesia

2021 ◽  
Vol 7 (1) ◽  
pp. 00792-2020
Author(s):  
Pedro Sampaio ◽  
Mónica Ferro da Silva ◽  
Inês Vale ◽  
Mónica Roxo-Rosa ◽  
Andreia Pinto ◽  
...  
Keyword(s):  
Healthy Volunteers ◽  
Open Source ◽  
Primary Ciliary Dyskinesia ◽  
High Speed ◽  
Beat Frequency ◽  
Ciliary Beat Frequency ◽  
Control Group ◽  
Observation Method ◽  
Ciliary Dyskinesia ◽  
Ciliary Beat

Evaluation of ciliary beat frequency (CBF) performed by high-speed videomicroscopy analysis (HVMA) is one of the techniques required for the correct diagnosis of primary ciliary dyskinesia (PCD). Currently, due to lack of open-source software, this technique is widely performed by visually counting the ciliary beatings per a given time-window. Our aim was to generate open-source, fast and intuitive software for evaluating CBF, validated in Portuguese PCD patients and healthy volunteers.Nasal brushings collected from 17 adult healthy volunteers and 34 PCD-referred subjects were recorded using HVMA. Evaluation of CBF was compared by two different methodologies: the new semi-automated computer software CiliarMove and the manual observation method using slow-motion movies. Clinical history, nasal nitric oxide and transmission electron microscopy were performed for diagnosis of PCD in the patient group. Genetic analysis was performed in a subset (n=8) of suspected PCD patients.The correlation coefficient between the two methods was R2=0.9895. The interval of CBF values obtained from the healthy control group (n=17) was 6.18–9.17 Hz at 25°C. In the PCD-excluded group (n=16), CBF ranged from 6.84 to 10.93 Hz and in the PCD group (n=18), CBF ranged from 0 to 14.30 Hz.We offer an automated open-source programme named CiliarMove, validated by the manual observation method in a healthy volunteer control group, a PCD-excluded group and a PCD-confirmed group. In our hands, comparisons between CBF intervals alone could discern between healthy and PCD groups in 78% of the cases.

scholarly journals Wampa is a dynein subunit required for axonemal assembly and male fertility in Drosophila

2019 ◽  
Author(s):  
Elisabeth Bauerly ◽  
Kexi Yi ◽  
Matthew C. Gibson
Keyword(s):  
Male Fertility ◽  
Primary Ciliary Dyskinesia ◽  
Functional Role ◽  
Essential Role ◽  
Respiratory Infections ◽  
Pathological Features ◽  
Dynein Arms ◽  
Cilia And Flagella ◽  
Ciliary Dyskinesia

AbstractAxonemal dyneins are motor proteins that form the inner and outer arms of the axoneme in cilia and flagella. Defects in dynein arms are the leading cause of primary ciliary dyskinesia (PCD), which is characterized by chronic respiratory infections, situs inversus, and sterility. Despite current understanding of pathological features associated with PCD, many of their causative genes still remain elusive. Here we analyze genetic requirements for wampa (wam), a previously uncharacterized component of the outer dynein arm that is essential for male fertility. In addition to a role in outer dynein arm formation, we uncovered additional requirements during spermatogenesis, including regulation of remodeling events for the mitochondria and the nucleus. Due to the conserved nature of axonemal dyneins and their essential role in both PCD and fertility, this study advances our understanding of the pathology of PCD, as well as the functional role of dyneins in axonemal formation and spermatogenesis.

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