The molecular structure of mammalian primary cilia revealed by cryo-electron tomography
Jazyk angličtina Země Spojené státy americké Médium print-electronic
Typ dokumentu časopisecké články, práce podpořená grantem
Grantová podpora
819826
European Research Council - International
PubMed
32989303
PubMed Central
PMC7610599
DOI
10.1038/s41594-020-0507-4
PII: 10.1038/s41594-020-0507-4
Knihovny.cz E-zdroje
- MeSH
- buněčné kultury MeSH
- buňky MDCK MeSH
- Chlamydomonas metabolismus ultrastruktura MeSH
- cilie metabolismus ultrastruktura MeSH
- elektronová kryomikroskopie MeSH
- exprese genu MeSH
- lidé MeSH
- mikrofilamenta metabolismus ultrastruktura MeSH
- mikrotubuly metabolismus ultrastruktura MeSH
- proteiny asociované s mikrotubuly genetika metabolismus MeSH
- psi MeSH
- tomografie elektronová MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- psi MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Názvy látek
- proteiny asociované s mikrotubuly MeSH
Primary cilia are microtubule-based organelles that are important for signaling and sensing in eukaryotic cells. Unlike the thoroughly studied motile cilia, the three-dimensional architecture and molecular composition of primary cilia are largely unexplored. Yet, studying these aspects is necessary to understand how primary cilia function in health and disease. We developed an enabling method for investigating the structure of primary cilia isolated from MDCK-II cells at molecular resolution by cryo-electron tomography. We show that the textbook '9 + 0' arrangement of microtubule doublets is only present at the primary cilium base. A few microns out, the architecture changes into an unstructured bundle of EB1-decorated microtubules and actin filaments, putting an end to a long debate on the presence or absence of actin filaments in primary cilia. Our work provides a plethora of insights into the molecular structure of primary cilia and offers a methodological framework to study these important organelles.
Institute of Molecular Genetics of the Czech Academy of Sciences Prague Czech Republic
Max Planck Institute of Molecular Cell Biology and Genetics Dresden Germany
Zobrazit více v PubMed
Badano JL, Mitsuma N, Beales PL, Katsanis N. The ciliopathies: an emerging class of human genetic disorders. Annu Rev Genomics Hum Genet. 2006;7:125–148. PubMed
Fliegauf M, Benzing T, Omran H. When cilia go bad: cilia defects and ciliopathies. Nat Rev Mol Cell Biol. 2007;8:880–893. PubMed
Waters AM, Beales PL. Ciliopathies: an expanding disease spectrum. Pediatr Nephrol. 2011;26:1039–1056. PubMed PMC
Afzelius BA. A human syndrome caused by immotile cilia. Science. 1976;193:317–319. PubMed
Mitchison HM, Valente EM. Motile and non-motile cilia in human pathology: from function to phenotypes. J Pathol. 2017;241:294–309. PubMed
Satir P, Heuser T, Sale WS. A structural basis for how motile cilia beat. Bioscience. 2014;64:1073–1083. PubMed PMC
Harris EH, Stern DB, Witman GB. The Chlamydomonas Sourcebook. 2nd. Academic Press; 2009.
Pigino G, et al. Comparative structural analysis of eukaryotic flagella and cilia from Chlamydomonas, Tetrahymena, and sea urchins. J Struct Biol. 2012;178:199–206. PubMed
Bui KH, Sakakibara H, Movassagh T, Oiwa K, Ishikawa T. Asymmetry of inner dynein arms and inter-doublet links in Chlamydomonas flagella. J Cell Biol. 2009;186:437–446. PubMed PMC
Jordan MA, Diener DR, Stepanek L, Pigino G. The cryo-EM structure of intraflagellar transport trains reveals how dynein is inactivated to ensure unidirectional anterograde movement in cilia. Nat Cell Biol. 2018;20:1250–1255. PubMed
Nicastro D, et al. The molecular architecture of axonemes revealed by cryoelectron tomography. Science. 2006;313:944–948. PubMed
Witman GB. In: Methods in Enzymology. Vallee RB, editor. Vol. 134. Academic Press; 1986. pp. 280–290. PubMed
Ma M, et al. Structure of the decorated ciliary doublet microtubule. Cell. 2019;179:909–922.:e12. PubMed PMC
Lin J, Nicastro D. Asymmetric distribution and spatial switching of dynein activity generates ciliary motility. Science. 2018;360:eaar1968. PubMed PMC
Pigino G, Ishikawa T. Axonemal radial spokes: 3D structure, function and assembly. Bioarchitecture. 2012;2:50–58. PubMed PMC
Oda T, Yanagisawa H, Kamiya R, Kikkawa M. A molecular ruler determines the repeat length in eukaryotic cilia and flagella. Science. 2014;346:857–860. PubMed
Ishikawa H, Thompson J, Yates JR, III, Marshall WF. Proteomic analysis of mammalian primary cilia. Curr Biol. 2012;22:414–419. PubMed PMC
Mick DU, et al. Proteomics of primary cilia by proximity labeling. Dev Cell. 2015;35:497–512. PubMed PMC
Ott C, Lippincott-Schwartz J. Visualization of live primary cilia dynamics using fluorescence microscopy. Curr Protoc Cell Biol. 2012;57 4.26.1-4.26.22. PubMed PMC
Sun S, Fisher RL, Bowser SS, Pentecost BT, Sui H. Three-dimensional architecture of epithelial primary cilia. Proc Natl Acad Sci USA. 2019;116:9370–9379. PubMed PMC
Gluenz E, et al. Beyond 9+0: noncanonical axoneme structures characterize sensory cilia from protists to humans. FASEB J. 2010;24:3117–3121. PubMed PMC
Doroquez DB, Berciu C, Anderson JR, Sengupta P, Nicastro D. A high-resolution morphological and ultrastructural map of anterior sensory cilia and glia in Caenorhabditis elegans. Elife. 2014;3:e01948. PubMed PMC
Ishikawa H, Marshall WF. In: Methods in Enzymology. Marshall WF, editor. Vol. 525. Academic Press; 2013. pp. 311–325. PubMed
Huang B, Masyuk T, LaRusso N. In: Methods in Cell Biology. Sloboda RD, editor. Vol. 94. Academic Press; 2009. pp. 103–115. PubMed
Bujakowska KM, Liu Q, Pierce EA. Photoreceptor cilia and retinal ciliopathies. Cold Spring Harb Perspect Biol. 2017;9:a028274. PubMed PMC
Yuan S, Zhao L, Brueckner M, Sun Z. Intraciliary calcium oscillations initiate vertebrate left-right asymmetry. Curr Biol. 2015;25:556–567. PubMed PMC
Essner JJ, Amack JD, Nyholm MK, Harris EB, Yost HJ. Kupffer’s vesicle is a ciliated organ of asymmetry in the zebrafish embryo that initiates left-right development of the brain, heart and gut. Development. 2005;132:1247–1260. PubMed
Wheway G, Nazlamova L, Hancock JT. Signaling through the primary cilium. Front Cell Dev Biol. 2018 doi: 10.3389/fcell.2018.00008. PubMed DOI PMC
Anvarian Z, Mykytyn K, Mukhopadhyay S, Pedersen LB, Christensen ST. Cellular signalling by primary cilia in development, organ function and disease. Nat Rev Nephrol. 2019;15:199–219. PubMed PMC
Goetz JG, et al. Endothelial cilia mediate low flow sensing during zebrafish vascular development. Cell Rep. 2014;6:799–808. PubMed
Praetorius HA. The primary cilium as sensor of fluid flow: new building blocks to the model. A review in the theme: cell signaling: proteins, pathways and mechanisms. Am J Physiol Cell Physiol. 2015;308:C198–C208. PubMed
Yoder BK. Role of primary cilia in the pathogenesis of polycystic kidney disease. J Am Soc Nephrol. 2007;18:1381–1388. PubMed
Kathem SH, Mohieldin AM, Nauli SM. The roles of primary cilia in polycystic kidney disease. AIMS Mol Sci. 2014;1:27–46. PubMed PMC
Stepanek L, Pigino G. Microtubule doublets are double-track railways for intraflagellar transport trains. Science. 2016;352:721–724. PubMed
Kozminsky KG, Johnson KA, Forscher P, Rosenbaum JL. A motility in the eukaryotic flagellum unrelated to flagellar beating. Proc Natl Acad Sci USA. 1993;90:5519–5523. PubMed PMC
Ishikawa H, Marshall WF. Efficient live fluorescence imaging of intraflagellar transport in mammalian primary cilia. Methods Cell Biol. 2015;127:189–201. PubMed PMC
Sui H, Downing KH. Molecular architecture of axonemal microtubule doublets revealed by cryo-electron tomography. Nature. 2006;442:475–478. PubMed
Song K, et al. In situ structure determination at nanometer resolution using TYGRESS. Nat Methods. 2020;17:201–208. PubMed PMC
von Loeffelholz O, et al. Nucleotide- and Mal3-dependent changes in fission yeast microtubules suggest a structural plasticity view of dynamics. Nat Commun. 2017;8:2110. PubMed PMC
Fisch C, Dupuis-Williams P. Ultrastructure of cilia and flagella—back to the future! Biol Cell. 2011;103:249–270. PubMed
Hess RA. Small tubules, surprising discoveries: from efferent ductules in the turkey to the discovery that estrogen receptor alpha is essential for fertility in the male. Anim Reprod. 2015;1:7–23. PubMed PMC
Nguyen AM, Young Y-N, Jacobs CR. The primary cilium is a self-adaptable, integrating nexus for mechanical stimuli and cellular signaling. Biol Open. 2015;4:1733–1738. PubMed PMC
Owa M, et al. Inner lumen proteins stabilize doublet microtubules in cilia and flagella. Nat Commun. 2019;10:1143. PubMed PMC
Garvalov BK, et al. Luminal particles within cellular microtubules. J Cell Biol. 2006;174:759–765. PubMed PMC
Coombes C, et al. Mechanism of microtubule lumen entry for the α-tubulin acetyltransferase enzyme αTAT1. Proc Natl Acad Sci USA. 2016;113:E7176-E7184. PubMed PMC
Boehlke C, et al. Differential role of Rab proteins in ciliary trafficking: Rab23 regulates Smoothened levels. J Cell Sci. 2010;123:1460–1467. PubMed
Schrøder JM, et al. EB1 and EB3 promote cilia biogenesis by several centrosome-related mechanisms. J Cell Sci. 2011;124:2539–2551. PubMed PMC
Pedersen LB, Geimer S, Sloboda RD, Rosenbaum JL. The microtubule plus end-tracking protein EB1 is localized to the flagellar tip and basal bodies in Chlamydomonas reinhardtii. Curr Biol. 2003;13:1969–1974. PubMed
Roth D, Fitton BP, Chmel NP, Wasiluk N, Straube A. Spatial positioning of EB family proteins at microtubule tips involves distinct nucleotide-dependent binding properties. J Cell Sci. 2018;132:jcs219550. PubMed PMC
Leterrier C, et al. End-binding proteins EB3 and EB1 link microtubules to ankyrin G in the axon initial segment. Proc Natl Acad Sci USA. 2011;108:8826–8831. PubMed
Schrøder JM, Schneider L, Christensen ST, Pedersen LB. EB1 is required for primary cilia assembly in fibroblasts. Curr Biol. 2007;17:1134–1139. PubMed
Lopez BJ, Valentine MT. Mechanical effects of EB1 on microtubules depend on GTP hydrolysis state and presence of paclitaxel. Cytoskeleton (Hoboken) 2014;71:530–541. PubMed
Zhang R, LaFrance B, Nogales E. Separating the effects of nucleotide and EB binding on microtubule structure. Proc Natl Acad Sci USA. 2018;115:E6191-E6200. PubMed PMC
Stroud MJ, et al. GAS2-like proteins mediate communication between microtubules and actin through interactions with end-binding proteins. J Cell Sci. 2014;127:2672–2682. PubMed PMC
Nazgiewicz A, Atherton P, Ballestrem C. GAS2-like 1 coordinates cell division through its association with end-binding proteins. Sci Rep. 2019;9:5805. PubMed PMC
Slep KC, et al. Structural determinants for EB1-mediated recruitment of APC and spectraplakins to the microtubule plus end. J Cell Biol. 2005;168:587–598. PubMed PMC
Lee S, Tan HY, Geneva II, Kruglov A, Calvert PD. Actin filaments partition primary cilia membranes into distinct fluid corrals. J Cell Biol. 2018;217:2831–2849. PubMed PMC
Copeland SJ, McRae A, Guarguaglini G, Trinkle-Mulcahy L, Copeland JW. Actin-dependent regulation of cilia length by the inverted formin FHDC1. Mol Biol Cell. 2018;29:1611–1627. PubMed PMC
Kohli P, et al. The ciliary membrane-associated proteome reveals actin-binding proteins as key components of cilia. EMBO Rep. 2017;18:1521–1535. PubMed PMC
Phua SC, et al. Dynamic remodeling of membrane composition drives cell cycle through primary cilia excision. Cell. 2017;168:264–279.:e15. PubMed PMC
Zuo X, et al. Primary cilia and the exocyst are linked to urinary extracellular vesicle production and content. J Biol Chem. 2019;294:19099–19110. PubMed PMC
Nager AR, et al. An actin network dispatches ciliary GPCRs into extracellular vesicles to modulate signaling. Cell. 2017;168:252–263.:e14. PubMed PMC
Mirvis M, Stearns T, James Nelson W. Cilium structure, assembly, and disassembly regulated by the cytoskeleton. Biochem J. 2018;475:2329–2353. PubMed PMC
BBSome-deficient cells activate intraciliary CDC42 to trigger actin-dependent ciliary ectocytosis
MAP9/MAPH-9 supports axonemal microtubule doublets and modulates motor movement
A protocol for generation and live-cell imaging analysis of primary cilia reporter cell lines