RESUMEN
Primary cilia on granule cell neuron progenitors in the developing cerebellum detect sonic hedgehog to facilitate proliferation. Following differentiation, cerebellar granule cells become the most abundant neuronal cell type in the brain. While granule cell cilia are essential during early developmental stages, they become infrequent upon maturation. Here, we provide nanoscopic resolution of cilia in situ using large-scale electron microscopy volumes and immunostaining of mouse cerebella. In many granule cells, we found intracellular cilia, concealed from the external environment. Cilia were disassembled in differentiating granule cell neurons-in a process we call cilia deconstruction-distinct from premitotic cilia resorption in proliferating progenitors. In differentiating granule cells, cilia deconstruction involved unique disassembly intermediates, and, as maturation progressed, mother centriolar docking at the plasma membrane. Unlike ciliated neurons in other brain regions, our results show the deconstruction of concealed cilia in differentiating granule cells, which might prevent mitogenic hedgehog responsiveness. Ciliary deconstruction could be paradigmatic of cilia removal during differentiation in other tissues.
Asunto(s)
Diferenciación Celular , Cerebelo , Cilios , Proteínas Hedgehog , Neuronas , Cilios/metabolismo , Cilios/ultraestructura , Animales , Neuronas/metabolismo , Neuronas/citología , Neuronas/ultraestructura , Ratones , Cerebelo/metabolismo , Cerebelo/citología , Proteínas Hedgehog/metabolismo , Proteínas Hedgehog/genética , Neurogénesis , Centriolos/metabolismo , Centriolos/ultraestructura , Ratones Endogámicos C57BLRESUMEN
As daughter centrioles assemble during G2, they recruit conserved Ana3/RTTN followed by its partner Rcd4/PPP1R35. Together, this contributes to the subsequent recruitment of Ana1/CEP295, required for the centriole's conversion to a centrosome. Here, we show that Rcd4/PPP1R35 is also required to maintain 9-fold centriole symmetry in the Drosophila male germline; its absence causes microtubule triplets to disperse into a reduced number of doublet or singlet microtubules. rcd4-null mutant spermatocytes display skinny centrioles that elongate normally and localize centriolar components correctly. Mutant spermatocytes also have centrioles of normal girth that splay at their proximal ends when induced to elongate by Ana1 overexpression. Skinny and splayed spermatid centrioles can still recruit a proximal centriole-like (PCL) structure marking a capability to initiate features of centriole duplication in developing sperm. Thus, stable 9-fold symmetry of microtubule triplets is not essential for centriole growth, correct longitudinal association of centriole components, and aspects of centriole duplication.
Asunto(s)
Centriolos , Proteínas de Drosophila , Microtúbulos , Espermatocitos , Centriolos/metabolismo , Centriolos/ultraestructura , Centriolos/genética , Animales , Masculino , Proteínas de Drosophila/metabolismo , Proteínas de Drosophila/genética , Espermatocitos/metabolismo , Microtúbulos/metabolismo , Drosophila melanogaster/genética , Drosophila melanogaster/metabolismo , Espermátides/metabolismo , Espermátides/citología , Proteínas de Ciclo Celular/metabolismo , Proteínas de Ciclo Celular/genética , Proteínas Asociadas a Microtúbulos/metabolismo , Proteínas Asociadas a Microtúbulos/genética , Mutación , DrosophilaRESUMEN
The Commander complex, a 16-protein assembly, plays multiple roles in cell homeostasis, cell cycle and immune response. It consists of copper-metabolism Murr1 domain proteins (COMMD1-10), coiled-coil domain-containing proteins (CCDC22 and CCDC93), DENND10 and the Retriever subcomplex (VPS26C, VPS29 and VPS35L), all expressed ubiquitously in the body and linked to various diseases. Here, we report the structure and key interactions of the endogenous human Commander complex by cryogenic-electron microscopy and mass spectrometry-based proteomics. The complex consists of a stable core of COMMD1-10 and an effector containing DENND10 and Retriever, scaffolded together by CCDC22 and CCDC93. We establish the composition of Commander and reveal major interaction interfaces. These findings clarify its roles in intracellular transport, and uncover a strong association with cilium assembly, and centrosome and centriole functions.
Asunto(s)
Microscopía por Crioelectrón , Humanos , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Proteínas Adaptadoras Transductoras de Señales/química , Modelos Moleculares , Proteínas de Transporte Vesicular/metabolismo , Proteínas de Transporte Vesicular/química , Complejos Multiproteicos/metabolismo , Complejos Multiproteicos/química , Células HEK293 , Unión Proteica , Cilios/metabolismo , Cilios/ultraestructura , Centriolos/metabolismo , Centriolos/ultraestructuraRESUMEN
BACKGROUND: The centrosome is the main center of the organization of microtubules (MT) in the cell, the origin for the formation of flagella and cilia, as well as the site of many regulatory intracellular processes. In diploid cells, the centrosome includes two centrioles connected to some additional structures and surrounded by pericentriolar material. METHODS: The ultrastructure of the cells was studied using transmission electron microscopy on serial ultrathin sections. RESULTS: Here, using transmission electron microscopy on a complete series of ultrathin sections of the centrosome region, we studied the relation between the number of centrioles and ploidy in diploid cells of female wasps and haploid cells of male in the parasitoid wasp Anisopteromalus calandrae (Hymenoptera). It showed that the haploid cells of the male insect have the same number of centrioles as the diploid cells of the female. CONCLUSIONS: It can be concluded that there is no strict correlation between the number of chromosome sets (ploidy) and the number of centrioles in haplodiploid insects.
Asunto(s)
Centriolos , Avispas , Animales , Masculino , Femenino , Centriolos/genética , Centriolos/ultraestructura , Avispas/genética , Haploidia , Diploidia , CentrosomaRESUMEN
Centrosome formation during early development in mice and rats occurs due to the appearance of centrioles de novo. In contrast, in humans and other non-rodent mammals, centrioles are thought to be derived from spermatozoa. Ultrastructural study of zygotes and early embryos of cattle at full series of ultrathin sections show that the proximal centriole of the spermatozoon disappears by the end of the first cleavage division. Centrioles appear in two to four cell embryos in fertilized oocytes and in parthenogenetic embryos. Centriole formation includes the appearance of atypical centrioles with randomly arranged triplets and centrioles with microtubule triplets of various lengths. After the third cleavage, four centriolar cylinders appear for the first time in the blastomeres while each embryo still has two atypical centrioles. Our results showed that the mechanisms of centriole formation in different groups of mammals are universal, differing only in the stage of development in which they occur.
Asunto(s)
Centrosoma , Oocitos , Humanos , Masculino , Bovinos , Animales , Ratones , Ratas , Oocitos/ultraestructura , Centrosoma/ultraestructura , Centriolos/ultraestructura , Espermatozoides/ultraestructura , MamíferosRESUMEN
The sperm competition theory, as proposed by Geoff Parker, predicts that sperm evolve through a cascade of changes. As an example, internal fertilization is followed by sperm morphology diversification. However, little is known about the evolution of internal sperm structures. The centriole has an ancient and evolutionarily conserved canonical structure with signature 9-fold, radially symmetric microtubules that form the cell's centrosomes, cilia, and flagella. Most animal spermatozoa have two centrioles, one of which forms the spermatozoan flagellum. Both are delivered to the egg and constitute the embryo's first two centrosomes. The spermatozoa of mammals and insects only have one recognizable centriole with a canonical structure. A second sperm centriole with an atypical structure was recently reported in both animal groups and which, prior to this, eluded discovery by standard techniques and criteria. Because the ancestors of both mammals and insects reproduced by internal fertilization, we hypothesized that the transition from two centrioles with canonical composition in ancestral sperm to an atypical centriolar composition characterized by only one canonical centriole evolved preferentially after internal fertilization. We examined fish because of the diversity of species available to test this hypothesis−as some species reproduce via internal and others via external fertilization−and because their spermatozoan ultrastructure has been extensively studied. Our literature search reports on 277 fish species. Species reported with atypical centriolar composition are specifically enriched among internal fertilizers compared to external fertilizers (7/34, 20.6% versus 2/243, 0.80%; p < 0.00001, odds ratio = 32.4) and represent phylogenetically unrelated fish. Atypical centrioles are present in the internal fertilizers of the subfamily Poeciliinae. Therefore, internally fertilizing fish preferentially and independently evolved spermatozoa with atypical centriolar composition multiple times, agreeing with Parker's cascade theory.
Asunto(s)
Centriolos , Fertilizantes , Animales , Centriolos/ultraestructura , Centrosoma/ultraestructura , Fertilización , Masculino , Mamíferos , Espermatozoides/ultraestructuraRESUMEN
Centriole biogenesis and maintenance are crucial for cells to generate cilia and assemble centrosomes that function as microtubule organizing centers (MTOCs). Centriole biogenesis and MTOC function both require the microtubule nucleator γ-tubulin ring complex (γTuRC). It is widely accepted that γTuRC nucleates microtubules from the pericentriolar material that is associated with the proximal part of centrioles. However, γTuRC also localizes more distally and in the centriole lumen, but the significance of these findings is unclear. Here we identify spatially and functionally distinct subpopulations of centrosomal γTuRC. Luminal localization is mediated by augmin, which is linked to the centriole inner scaffold through POC5. Disruption of luminal localization impairs centriole integrity and interferes with cilium assembly. Defective ciliogenesis is also observed in γTuRC mutant fibroblasts from a patient suffering from microcephaly with chorioretinopathy. These results identify a non-canonical role of augmin-γTuRC in the centriole lumen that is linked to human disease.
Asunto(s)
Proteínas de Ciclo Celular/aislamiento & purificación , Proteínas de Ciclo Celular/metabolismo , Centriolos/metabolismo , Proteínas Asociadas a Microtúbulos/aislamiento & purificación , Proteínas Asociadas a Microtúbulos/metabolismo , Centro Organizador de los Microtúbulos/metabolismo , Animales , Proteínas Portadoras/metabolismo , Proteínas de Ciclo Celular/ultraestructura , Línea Celular , Centriolos/ultraestructura , Centrosoma/metabolismo , Centrosoma/ultraestructura , Cilios , Femenino , Humanos , Masculino , Ratones , Proteínas Asociadas a Microtúbulos/ultraestructura , Centro Organizador de los Microtúbulos/ultraestructura , Microtúbulos/metabolismo , NeuronasRESUMEN
Among the morphological processes that characterize the early stages of Drosophila oogenesis, the dynamic of the centrioles deserves particular attention. We re-examined the architecture and the distribution of the centrioles within the germarium and early stages of the vitellarium. We found that most of the germ cell centrioles diverge from the canonical model and display notable variations in size. Moreover, duplication events were frequently observed within the germarium in the absence of DNA replication. Finally, we report the presence of an unusually long centriole that is first detected in the cystoblast and is always associated with the developing oocyte. This centriole is directly inherited after the asymmetric division of the germline stem cells and persists during the process of oocyte selection, thus already representing a marker for oocyte identification at the beginning of its formation and during the ensuing developmental stages.
Asunto(s)
Centriolos/fisiología , Drosophila melanogaster/fisiología , Oocitos/fisiología , Oogénesis , Animales , Centriolos/genética , Centriolos/ultraestructura , Drosophila melanogaster/genética , Drosophila melanogaster/ultraestructura , Femenino , Microscopía Electrónica de Transmisión , Oocitos/ultraestructura , Factores de TiempoRESUMEN
Cells inherit two centrioles, the older of which is uniquely capable of generating a cilium. Using proteomics and superresolved imaging, we identify a module that we term DISCO (distal centriole complex). The DISCO components CEP90, MNR, and OFD1 underlie human ciliopathies. This complex localizes to both distal centrioles and centriolar satellites, proteinaceous granules surrounding centrioles. Cells and mice lacking CEP90 or MNR do not generate cilia, fail to assemble distal appendages, and do not transduce Hedgehog signals. Disrupting the satellite pools does not affect distal appendage assembly, indicating that it is the centriolar populations of MNR and CEP90 that are critical for ciliogenesis. CEP90 recruits the most proximal known distal appendage component, CEP83, to root distal appendage formation, an early step in ciliogenesis. In addition, MNR, but not CEP90, restricts centriolar length by recruiting OFD1. We conclude that DISCO acts at the distal centriole to support ciliogenesis by restraining centriole length and assembling distal appendages, defects in which cause human ciliopathies.
Asunto(s)
Centriolos/metabolismo , Cilios/metabolismo , Ciliopatías/genética , Proteínas Asociadas a Microtúbulos/genética , Proteínas/genética , Animales , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Línea Celular , Centriolos/patología , Centriolos/ultraestructura , Cilios/patología , Cilios/ultraestructura , Ciliopatías/metabolismo , Ciliopatías/patología , Embrión de Mamíferos , Células Epiteliales/citología , Células Epiteliales/metabolismo , Regulación de la Expresión Génica , Genes Reporteros , Proteínas Fluorescentes Verdes/genética , Proteínas Fluorescentes Verdes/metabolismo , Humanos , Proteínas Luminiscentes/genética , Proteínas Luminiscentes/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Proteínas Asociadas a Microtúbulos/deficiencia , Proteínas Asociadas a Microtúbulos/metabolismo , Proteínas/metabolismo , Epitelio Pigmentado de la Retina/citología , Epitelio Pigmentado de la Retina/metabolismo , Transducción de SeñalRESUMEN
Centrioles are evolutionarily conserved multi-protein organelles essential for forming cilia and centrosomes. Centriole biogenesis begins with self-assembly of SAS-6 proteins into 9-fold symmetrical ring polymers, which then stack into a cartwheel that scaffolds organelle formation. The importance of this architecture has been difficult to decipher notably because of the lack of precise tools to modulate the underlying assembly reaction. Here, we developed monobodies against Chlamydomonas reinhardtii SAS-6, characterizing three in detail with X-ray crystallography, atomic force microscopy and cryo-electron microscopy. This revealed distinct monobody-target interaction modes, as well as specific consequences on ring assembly and stacking. Of particular interest, monobody MBCRS6-15 induces a conformational change in CrSAS-6, resulting in the formation of a helix instead of a ring. Furthermore, we show that this alteration impairs centriole biogenesis in human cells. Overall, our findings identify monobodies as powerful molecular levers to alter the architecture of multi-protein complexes and tune centriole assembly.
Asunto(s)
Proteínas Portadoras/metabolismo , Proteínas de Ciclo Celular/metabolismo , Centriolos/metabolismo , Proteínas Algáceas/química , Proteínas Algáceas/metabolismo , Proteínas Portadoras/química , Proteínas de Ciclo Celular/antagonistas & inhibidores , Proteínas de Ciclo Celular/química , Centriolos/ultraestructura , Chlamydomonas reinhardtii/metabolismo , Chlamydomonas reinhardtii/ultraestructura , Microscopía por Crioelectrón , Cristalografía por Rayos X , Microscopía de Fuerza Atómica , Modelos Moleculares , Unión Proteica , Multimerización de Proteína , Estructura Terciaria de ProteínaRESUMEN
Reproductive success depends on efficient sperm movement driven by axonemal dynein-mediated microtubule sliding. Models predict sliding at the base of the tail - the centriole - but such sliding has never been observed. Centrioles are ancient organelles with a conserved architecture; their rigidity is thought to restrict microtubule sliding. Here, we show that, in mammalian sperm, the atypical distal centriole (DC) and its surrounding atypical pericentriolar matrix form a dynamic basal complex (DBC) that facilitates a cascade of internal sliding deformations, coupling tail beating with asymmetric head kinking. During asymmetric tail beating, the DC's right side and its surroundings slide ~300 nm rostrally relative to the left side. The deformation throughout the DBC is transmitted to the head-tail junction; thus, the head tilts to the left, generating a kinking motion. These findings suggest that the DBC evolved as a dynamic linker coupling sperm head and tail into a single self-coordinated system.
Asunto(s)
Motilidad Espermática/fisiología , Animales , Centriolos/fisiología , Centriolos/ultraestructura , Humanos , Masculino , Mamíferos , Microtúbulos/fisiología , Microtúbulos/ultraestructura , Cabeza del Espermatozoide/fisiología , Cola del Espermatozoide/fisiología , Cola del Espermatozoide/ultraestructuraRESUMEN
Expansion microscopy is an imaging method based on isotropic physical expansion of biological samples, which improves optical resolution and allows imaging of subresolutional cellular components by conventional microscopes. Centrioles are small microtubule-based cylindrical structures that build centrosomes and cilia, two organelles essential for vertebrates. Due to a centriole's small size, electron microscopy has traditionally been used to study centriole length and ultrastructural features. Recently, expansion microscopy has been successfully used as an affordable and accessible alternative to electron microscopy in the analysis of centriole and cilia length and structural features. Here, we describe an expansion microscopy approach for the analysis of centrioles and cilia in large populations of mammalian adherent and nonadherent cells and multiciliated cultures.
Asunto(s)
Centriolos/ultraestructura , Cilios/ultraestructura , Adhesión Celular , Técnicas de Cultivo de Célula/métodos , Línea Celular , Humanos , MicroscopíaRESUMEN
How cells control the numbers of subcellular components is a fundamental question in biology. Given that biosynthetic processes are fundamentally stochastic it is utterly puzzling that some structures display no copy number variation within a cell population. Centriole biogenesis, with each centriole being duplicated once and only once per cell cycle, stands out due to its remarkable fidelity. This is a highly controlled process, which depends on low-abundance rate-limiting factors. How can exactly one centriole copy be produced given the variation in the concentration of these key factors? Hitherto, tentative explanations of this control evoked lateral inhibition- or phase separation-like mechanisms emerging from the dynamics of these rate-limiting factors but how strict centriole number is regulated remains unsolved. Here, a novel solution to centriole copy number control is proposed based on the assembly of a centriolar scaffold, the cartwheel. We assume that cartwheel building blocks accumulate around the mother centriole at supercritical concentrations, sufficient to assemble one or more cartwheels. Our key postulate is that once the first cartwheel is formed it continues to elongate by stacking the intermediate building blocks that would otherwise form supernumerary cartwheels. Using stochastic models and simulations, we show that this mechanism may ensure formation of one and only one cartwheel robustly over a wide range of parameter values. By comparison to alternative models, we conclude that the distinctive signatures of this novel mechanism are an increasing assembly time with cartwheel numbers and the translation of stochasticity in building block concentrations into variation in cartwheel numbers or length.
Asunto(s)
Centriolos/metabolismo , Centriolos/ultraestructura , Modelos Biológicos , Ciclo Celular/fisiología , Proteínas de Ciclo Celular/química , Proteínas de Ciclo Celular/metabolismo , Centriolos/química , Biología Computacional , Simulación por Computador , Humanos , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Estructura Cuaternaria de Proteína , Procesos EstocásticosRESUMEN
Motile cilia are molecular machines used by a myriad of eukaryotic cells to swim through fluid environments. However, available molecular structures represent only a handful of cell types, limiting our understanding of how cilia are modified to support motility in diverse media. Here, we use cryo-focused ion beam milling-enabled cryo-electron tomography to image sperm flagella from three mammalian species. We resolve in-cell structures of centrioles, axonemal doublets, central pair apparatus, and endpiece singlets, revealing novel protofilament-bridging microtubule inner proteins throughout the flagellum. We present native structures of the flagellar base, which is crucial for shaping the flagellar beat. We show that outer dense fibers are directly coupled to microtubule doublets in the principal piece but not in the midpiece. Thus, mammalian sperm flagella are ornamented across scales, from protofilament-bracing structures reinforcing microtubules at the nano-scale to accessory structures that impose micron-scale asymmetries on the entire assembly. Our structures provide vital foundations for linking molecular structure to ciliary motility and evolution.
Asunto(s)
Cola del Espermatozoide/ultraestructura , Animales , Axonema/ultraestructura , Movimiento Celular , Centriolos/ultraestructura , Cilios/fisiología , Microscopía por Crioelectrón , Tomografía con Microscopio Electrónico , Caballos , Masculino , Ratones , Ratones Endogámicos C57BL , Cola del Espermatozoide/fisiología , PorcinosRESUMEN
The centrosome is the main microtubule-organizing center in animal cells. It comprises of two centrioles and the surrounding pericentriolar material. Protein organization at the outer layer of the centriole and outward has been studied extensively; however, an overall picture of the protein architecture at the centriole core has been missing. Here we report a direct view of Drosophila centriolar proteins at â¼50-nm resolution. This reveals a Sas6 ring at the C-terminus, where it overlaps with the C-terminus of Cep135. The ninefold symmetrical pattern of Cep135 is further conveyed through Ana1-Asterless axes that extend past the microtubule wall from between the blades. Ana3 and Rcd4, whose termini are close to Cep135, are arranged in ninefold symmetry that does not match the above axes. During centriole biogenesis, Ana3 and Rcd4 are sequentially loaded on the newly formed centriole and are required for centriole-to-centrosome conversion through recruiting the Cep135-Ana1-Asterless complex. Together, our results provide a spatiotemporal map of the centriole core and implications of how the structure might be built.
Asunto(s)
Centriolos/metabolismo , Centriolos/ultraestructura , Animales , Línea Celular , Centriolos/genética , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Drosophila melanogaster , Dominios ProteicosRESUMEN
Centrosomes are composed of two orthogonally arranged centrioles surrounded by an electron-dense matrix called the pericentriolar material (PCM). Centrioles are cylinders with diameters of ~250 nm, are several hundred nanometres in length and consist of 9-fold symmetrically arranged microtubules (MT). In dividing animal cells, centrosomes act as the principal MT-organising centres and they also organise actin, which tunes cytoplasmic MT nucleation. In some specialised cells, the centrosome acquires additional critical structures and converts into the base of a cilium with diverse functions including signalling and motility. These structures are found in most eukaryotes and are essential for development and homoeostasis at both cellular and organism levels. The ultrastructure of centrosomes and their derived organelles have been known for more than half a century. However, recent advances in a number of techniques have revealed the high-resolution structures (at Å-to-nm scale resolution) of centrioles and have begun to uncover the molecular principles underlying their properties, including: protein components; structural elements; and biogenesis in various model organisms. This review covers advances in our understanding of the features and processes that are critical for the biogenesis of the evolutionarily conserved structures of the centrosomes. Furthermore, it discusses how variations of these aspects can generate diversity in centrosome structure and function among different species and even between cell types within a multicellular organism.
Asunto(s)
Centriolos/ultraestructura , Cilios/ultraestructura , Proteínas Asociadas a Microtúbulos/genética , Microtúbulos/ultraestructura , Biogénesis de Organelos , Actinas/genética , Actinas/metabolismo , Animales , Biodiversidad , Evolución Biológica , Ciclo Celular/genética , Centriolos/metabolismo , Chlorophyta/genética , Chlorophyta/metabolismo , Chlorophyta/ultraestructura , Cilios/metabolismo , Células Eucariotas/metabolismo , Células Eucariotas/ultraestructura , Regulación de la Expresión Génica , Humanos , Proteínas Asociadas a Microtúbulos/clasificación , Proteínas Asociadas a Microtúbulos/metabolismo , Microtúbulos/metabolismo , Especificidad de la Especie , Tubulina (Proteína)/genética , Tubulina (Proteína)/metabolismoRESUMEN
The primary cilium is a ubiquitous microtubule-based organelle that senses external environment and modulates diverse signaling pathways in different cell types and tissues. The cilium originates from the mother centriole through a complex set of cellular events requiring hundreds of distinct components. Aberrant ciliogenesis or ciliary transport leads to a broad spectrum of clinical entities with overlapping yet highly variable phenotypes, collectively called ciliopathies, which include sensory defects and syndromic disorders with multi-organ pathologies. For efficient light detection, photoreceptors in the retina elaborate a modified cilium known as the outer segment, which is packed with membranous discs enriched for components of the phototransduction machinery. Retinopathy phenotype involves dysfunction and/or degeneration of the light sensing photoreceptors and is highly penetrant in ciliopathies. This review will discuss primary cilia biogenesis and ciliopathies, with a focus on the retina, and the role of CP110-CEP290-CC2D2A network. We will also explore how recent technologies can advance our understanding of cilia biology and discuss new paradigms for developing potential therapies of retinal ciliopathies.
Asunto(s)
Antígenos de Neoplasias/genética , Proteínas de Ciclo Celular/genética , Cilios/metabolismo , Ciliopatías/genética , Proteínas del Citoesqueleto/genética , Proteínas Asociadas a Microtúbulos/genética , Fosfoproteínas/genética , Degeneración Retiniana/genética , Retinitis Pigmentosa/genética , Animales , Antígenos de Neoplasias/metabolismo , Transporte Biológico , Proteínas de Ciclo Celular/metabolismo , Centriolos/metabolismo , Centriolos/ultraestructura , Cilios/ultraestructura , Ciliopatías/metabolismo , Ciliopatías/patología , Proteínas del Citoesqueleto/metabolismo , Modelos Animales de Enfermedad , Regulación de la Expresión Génica , Humanos , Fototransducción , Ratones , Proteínas Asociadas a Microtúbulos/metabolismo , Microtúbulos/metabolismo , Microtúbulos/ultraestructura , Fosfoproteínas/metabolismo , Células Fotorreceptoras de Vertebrados/metabolismo , Células Fotorreceptoras de Vertebrados/patología , Degeneración Retiniana/metabolismo , Degeneración Retiniana/patología , Retinitis Pigmentosa/metabolismo , Retinitis Pigmentosa/patologíaRESUMEN
Multiciliated cells (MCC) project dozens to hundreds of motile cilia from the cell surface to generate fluid flow across epithelial surfaces or turbulence to promote the transport of gametes. The MCC differentiation program is initiated by GEMC1 and MCIDAS, members of the geminin family, that activate key transcription factors, including p73 and FOXJ1, to control the multiciliogenesis program. To support the generation of multiple motile cilia, MCCs must undergo massive centriole amplification to generate a sufficient number of basal bodies (modified centrioles). This transcriptional program involves the generation of deuterosomes, unique structures that act as platforms to regulate centriole amplification, the reactivation of cell cycle programs to control centriole amplification and release, and extensive remodeling of the cytoskeleton. This review will focus on providing an overview of the transcriptional regulation of MCCs and its connection to key processes, in addition to highlighting exciting recent developments and open questions in the field.
Asunto(s)
Proteínas de Ciclo Celular/genética , Centriolos/metabolismo , Cilios/metabolismo , Ciliopatías/genética , Factores de Transcripción/genética , Transcripción Genética , Animales , Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Diferenciación Celular , Centriolos/ultraestructura , Cilios/ultraestructura , Ciliopatías/metabolismo , Ciliopatías/patología , Citoesqueleto/metabolismo , Citoesqueleto/ultraestructura , Factores de Transcripción Forkhead/genética , Factores de Transcripción Forkhead/metabolismo , Regulación de la Expresión Génica , Humanos , Transducción de Señal , Factores de Transcripción/metabolismo , Proteína Tumoral p73/genética , Proteína Tumoral p73/metabolismoRESUMEN
Multiciliogenesis is a cascading process for generating hundreds of motile cilia in single cells. In vertebrates, this process has been investigated in the ependyma of brain ventricles and the ciliated epithelia of the airway and oviduct. Although the early steps to amplify centrioles have been characterized in molecular detail, subsequent steps to establish multicilia have been relatively overlooked. Here, we focused on unusual cilia-related structures previously observed in wild-type mouse ependyma using transmission electron microscopy and analyzed their ultrastructural features and the frequency of their occurrence. In the ependyma, $\sim$5% of cilia existed as bundles; while the majority of the bundles were paired, bundles of more than three cilia were also found. Furthermore, apical protrusions harboring multiple sets of axonemes were occasionally observed (0-2 per section), suggesting an unusual mode of ciliogenesis. In trachea and oviduct epithelia, ciliary bundles were absent, but protrusions containing multiple axonemes were observed. At the base of such protrusions, certain axonemes were completely enwrapped by membranes, whereas others remained incompletely enwrapped. These data suggested that the late steps of multiciliogenesis might include a unique process underlying the development of cilia, which is distinct from the ciliogenesis of primary cilia.
Asunto(s)
Centriolos/ultraestructura , Cilios/ultraestructura , Epéndimo/citología , Células Epiteliales/citología , Oviductos/citología , Tráquea/citología , Animales , Femenino , Ratones , Ratones Endogámicos C57BL , Microscopía Electrónica de TransmisiónRESUMEN
Centrioles are polarized microtubule-based organelles that seed the formation of cilia, and which assemble from a cartwheel containing stacked ring oligomers of SAS-6 proteins. A cryo-tomography map of centrioles from the termite flagellate Trichonympha spp. was obtained previously, but higher resolution analysis is likely to reveal novel features. Using sub-tomogram averaging (STA) in T. spp. and Trichonympha agilis, we delineate the architecture of centriolar microtubules, pinhead, and A-C linker. Moreover, we report ~25 Å resolution maps of the central cartwheel, revealing notably polarized cartwheel inner densities (CID). Furthermore, STA of centrioles from the distant flagellate Teranympha mirabilis uncovers similar cartwheel architecture and a distinct filamentous CID. Fitting the CrSAS-6 crystal structure into the flagellate maps and analyzing cartwheels generated in vitro indicate that SAS-6 rings can directly stack onto one another in two alternating configurations: with a slight rotational offset and in register. Overall, improved STA maps in three flagellates enabled us to unravel novel architectural features, including of centriole polarity and cartwheel stacking, thus setting the stage for an accelerated elucidation of underlying assembly mechanisms.