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  • 1
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    Primary cilia are not calcium-responsive mechanosensors (2016)
    Springer Nature
    In: Nature
    Details
    Publication Date: 2016-03-31
    Description: Primary cilia are not calcium-responsive mechanosensors Nature 531, 7596 (2016). doi:10.1038/nature17426 Authors: M. Delling, A. A. Indzhykulian, X. Liu, Y. Li, T. Xie, D. P. Corey & D. E. Clapham Primary cilia are solitary, generally non-motile, hair-like protrusions that extend from the surface of cells between cell divisions. Their antenna-like structure leads naturally to the assumption that they sense the surrounding environment, the most common hypothesis being sensation of mechanical force through calcium-permeable ion channels within the cilium. This Ca2+-responsive mechanosensor hypothesis for primary cilia has been invoked to explain a large range of biological responses, from control of left–right axis determination in embryonic development to adult progression of polycystic kidney disease and some cancers. Here we report the complete lack of mechanically induced calcium increases in primary cilia, in tissues upon which this hypothesis has been based. We developed a transgenic mouse, Arl13b–mCherry–GECO1.2, expressing a ratiometric genetically encoded calcium indicator in all primary cilia. We then measured responses to flow in primary cilia of cultured kidney epithelial cells, kidney thick ascending tubules, crown cells of the embryonic node, kinocilia of inner ear hair cells, and several cell lines. Cilia-specific Ca2+ influxes were not observed in physiological or even highly supraphysiological levels of fluid flow. We conclude that mechanosensation, if it originates in primary cilia, is not via calcium signalling.
    Print ISSN: 0028-0836
    Electronic ISSN: 1476-4687
    Topics: Biology , Chemistry and Pharmacology , Medicine , Natural Sciences in General , Physics
    Published by Springer Nature
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  • 2
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    Proliferation of functional hair cells in vivo in the absence of the retinoblastoma protein (2005)
    American Association for the Advancement of Science (AAAS)
    Details
    Publication Date: 2005-01-18
    Description: In mammals, hair cell loss causes irreversible hearing and balance impairment because hair cells are terminally differentiated and do not regenerate spontaneously. By profiling gene expression in developing mouse vestibular organs, we identified the retinoblastoma protein (pRb) as a candidate regulator of cell cycle exit in hair cells. Differentiated and functional mouse hair cells with a targeted deletion of Rb1 undergo mitosis, divide, and cycle, yet continue to become highly differentiated and functional. Moreover, acute loss of Rb1 in postnatal hair cells caused cell cycle reentry. Manipulation of the pRb pathway may ultimately lead to mammalian hair cell regeneration.〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Sage, Cyrille -- Huang, Mingqian -- Karimi, Kambiz -- Gutierrez, Gabriel -- Vollrath, Melissa A -- Zhang, Duan-Sun -- Garcia-Anoveros, Jaime -- Hinds, Philip W -- Corwin, Jeffrey T -- Corey, David P -- Chen, Zheng-Yi -- DC-00200/DC/NIDCD NIH HHS/ -- DC-04546/DC/NIDCD NIH HHS/ -- DC-AG20208/DC/NIDCD NIH HHS/ -- New York, N.Y. -- Science. 2005 Feb 18;307(5712):1114-8. Epub 2005 Jan 13.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉Neurology Service, MGH-HMS Center for Nervous System Repair, Massachusetts General Hospital and Harvard Medical School, Boston, MA 02114, USA.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/15653467" target="_blank"〉PubMed〈/a〉
    Keywords: Animals ; Apoptosis ; Cell Count ; Cell Cycle ; Cell Differentiation ; *Cell Proliferation ; Cell Shape ; Cochlea/cytology/embryology ; Female ; Gene Deletion ; Gene Expression Profiling ; Genes, Retinoblastoma ; Hair Cells, Auditory, Inner/*cytology/*physiology ; Mice ; Mice, Knockout ; Mitosis ; Oligonucleotide Array Sequence Analysis ; Pregnancy ; Pyridinium Compounds/metabolism ; Quaternary Ammonium Compounds/metabolism ; Regeneration ; Retinoblastoma Protein/genetics/*physiology ; Saccule and Utricle/embryology/metabolism ; Stem Cells/cytology/physiology
    Print ISSN: 0036-8075
    Electronic ISSN: 1095-9203
    Topics: Biology , Chemistry and Pharmacology , Computer Science , Medicine , Natural Sciences in General , Physics
    Published by American Association for the Advancement of Science (AAAS)
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  • 3
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    Multi-isotope imaging mass spectrometry reveals slow protein turnover in hair-cell stereocilia (2012)
    Nature Publishing Group (NPG)
    Details
    Publication Date: 2012-01-17
    Description: Hair cells of the inner ear are not normally replaced during an animal's life, and must continually renew components of their various organelles. Among these are the stereocilia, each with a core of several hundred actin filaments that arise from their apical surfaces and that bear the mechanotransduction apparatus at their tips. Actin turnover in stereocilia has previously been studied by transfecting neonatal rat hair cells in culture with a beta-actin-GFP fusion, and evidence was found that actin is replaced, from the top down, in 2-3 days. Overexpression of the actin-binding protein espin causes elongation of stereocilia within 12-24 hours, also suggesting rapid regulation of stereocilia lengths. Similarly, the mechanosensory 'tip links' are replaced in 5-10 hours after cleavage in chicken and mammalian hair cells. In contrast, turnover in chick stereocilia in vivo is much slower. It might be that only certain components of stereocilia turn over quickly, that rapid turnover occurs only in neonatal animals, only in culture, or only in response to a challenge like breakage or actin overexpression. Here we quantify protein turnover by feeding animals with a (15)N-labelled precursor amino acid and using multi-isotope imaging mass spectrometry to measure appearance of new protein. Surprisingly, in adult frogs and mice and in neonatal mice, in vivo and in vitro, the stereocilia were remarkably stable, incorporating newly synthesized protein at 〈10% per day. Only stereocilia tips had rapid turnover and no treadmilling was observed. Other methods confirmed this: in hair cells expressing beta-actin-GFP we bleached fiducial lines across hair bundles, but they did not move in 6 days. When we stopped expression of beta- or gamma-actin with tamoxifen-inducible recombination, neither actin isoform left the stereocilia, except at the tips. Thus, rapid turnover in stereocilia occurs only at the tips and not by a treadmilling process.〈br /〉〈br /〉〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3267870/" target="_blank"〉〈img src="https://static.pubmed.gov/portal/portal3rc.fcgi/4089621/img/3977009" border="0"〉〈/a〉   〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3267870/" target="_blank"〉This paper as free author manuscript - peer-reviewed and accepted for publication〈/a〉〈br /〉〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Zhang, Duan-Sun -- Piazza, Valeria -- Perrin, Benjamin J -- Rzadzinska, Agnieszka K -- Poczatek, J Collin -- Wang, Mei -- Prosser, Haydn M -- Ervasti, James M -- Corey, David P -- Lechene, Claude P -- 2P41RR0112553-12/RR/NCRR NIH HHS/ -- F32DC009539/DC/NIDCD NIH HHS/ -- P41EB001974/EB/NIBIB NIH HHS/ -- P41RR14579/RR/NCRR NIH HHS/ -- R01 AR042423/AR/NIAMS NIH HHS/ -- R01 AR042423-08/AR/NIAMS NIH HHS/ -- R01 AR049899/AR/NIAMS NIH HHS/ -- R01 DC000033/DC/NIDCD NIH HHS/ -- R01 DC002281/DC/NIDCD NIH HHS/ -- R01AR049899/AR/NIAMS NIH HHS/ -- R01D K58762/PHS HHS/ -- R01DC00033/DC/NIDCD NIH HHS/ -- R01DC02281/DC/NIDCD NIH HHS/ -- R01DC03463/DC/NIDCD NIH HHS/ -- R01DC04179/DC/NIDCD NIH HHS/ -- R01EY12963/EY/NEI NIH HHS/ -- R01GM47214/GM/NIGMS NIH HHS/ -- R37DK39773/DK/NIDDK NIH HHS/ -- WT079643/Wellcome Trust/United Kingdom -- Howard Hughes Medical Institute/ -- England -- Nature. 2012 Jan 15;481(7382):520-4. doi: 10.1038/nature10745.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉Department of Neurobiology, Harvard Medical School and Howard Hughes Medical Institute, Boston, Massachusetts 02115, USA.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/22246323" target="_blank"〉PubMed〈/a〉
    Keywords: Actins/metabolism ; Animals ; Animals, Newborn ; Bleaching Agents ; Chickens ; Epithelium/drug effects/metabolism ; Fiducial Markers ; Hair Cells, Auditory, Inner/*cytology ; Homologous Recombination/drug effects ; Mass Spectrometry/*methods ; Mice ; Mice, Inbred C57BL ; Proteins/*metabolism ; Rana catesbeiana ; Stereocilia/*metabolism ; Tamoxifen/pharmacology
    Print ISSN: 0028-0836
    Electronic ISSN: 1476-4687
    Topics: Biology , Chemistry and Pharmacology , Medicine , Natural Sciences in General , Physics
    Published by Nature Publishing Group (NPG)
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  • 4
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    Structure of a force-conveying cadherin bond essential for inner-ear mechanotransduction (2012)
    Nature Publishing Group (NPG)
    Details
    Publication Date: 2012-11-09
    Description: Hearing and balance use hair cells in the inner ear to transform mechanical stimuli into electrical signals. Mechanical force from sound waves or head movements is conveyed to hair-cell transduction channels by tip links, fine filaments formed by two atypical cadherins known as protocadherin 15 and cadherin 23 (refs 4, 5). These two proteins are involved in inherited deafness and feature long extracellular domains that interact tip-to-tip in a Ca(2+)-dependent manner. However, the molecular architecture of this complex is unknown. Here we combine crystallography, molecular dynamics simulations and binding experiments to characterize the protocadherin 15-cadherin 23 bond. We find a unique cadherin interaction mechanism, in which the two most amino-terminal cadherin repeats (extracellular cadherin repeats 1 and 2) of each protein interact to form an overlapped, antiparallel heterodimer. Simulations predict that this tip-link bond is mechanically strong enough to resist forces in hair cells. In addition, the complex is shown to become unstable in response to Ca(2+) removal owing to increased flexure of Ca(2+)-free cadherin repeats. Finally, we use structures and biochemical measurements to study the molecular mechanisms by which deafness mutations disrupt tip-link function. Overall, our results shed light on the molecular mechanics of hair-cell sensory transduction and on new interaction mechanisms for cadherins, a large protein family implicated in tissue and organ morphogenesis, neural connectivity and cancer.〈br /〉〈br /〉〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3518760/" target="_blank"〉〈img src="https://static.pubmed.gov/portal/portal3rc.fcgi/4089621/img/3977009" border="0"〉〈/a〉   〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3518760/" target="_blank"〉This paper as free author manuscript - peer-reviewed and accepted for publication〈/a〉〈br /〉〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Sotomayor, Marcos -- Weihofen, Wilhelm A -- Gaudet, Rachelle -- Corey, David P -- R01 DC002281/DC/NIDCD NIH HHS/ -- R01 DC02281/DC/NIDCD NIH HHS/ -- RC2GM093307/GM/NIGMS NIH HHS/ -- RR-15301/RR/NCRR NIH HHS/ -- Howard Hughes Medical Institute/ -- England -- Nature. 2012 Dec 6;492(7427):128-32. doi: 10.1038/nature11590. Epub 2012 Nov 7.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉Howard Hughes Medical Institute and Department of Neurobiology, Harvard Medical School, Boston, Massachusetts 02115, USA.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/23135401" target="_blank"〉PubMed〈/a〉
    Keywords: Animals ; Cadherins/*chemistry/genetics/*metabolism ; Calcium/metabolism/pharmacology ; Chromatography, Gel ; Crystallography, X-Ray ; Deafness/genetics ; Ear, Inner/cytology/*physiology ; Mechanotransduction, Cellular/*physiology ; Mice ; Models, Molecular ; Molecular Dynamics Simulation ; Mutagenesis, Site-Directed ; Mutation/genetics ; Protein Binding/drug effects ; Protein Multimerization/drug effects ; Protein Precursors/*chemistry/genetics/*metabolism ; Repetitive Sequences, Amino Acid
    Print ISSN: 0028-0836
    Electronic ISSN: 1476-4687
    Topics: Biology , Chemistry and Pharmacology , Medicine , Natural Sciences in General , Physics
    Published by Nature Publishing Group (NPG)
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  • 5
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    Mechanosensation and the DEG/ENaC ion channels (1996)
    American Association for the Advancement of Science (AAAS)
    Details
    Publication Date: 1996-07-19
    Description: 〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Corey, D P -- Garcia-Anoveros, J -- New York, N.Y. -- Science. 1996 Jul 19;273(5273):323-4.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉Howard Hughes Medical Institute, Boston, MA 02114, USA.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/8685718" target="_blank"〉PubMed〈/a〉
    Keywords: Amino Acid Sequence ; Animals ; Caenorhabditis elegans/genetics/*physiology ; Genes, Helminth ; Helminth Proteins/chemistry/genetics/*physiology ; Molecular Sequence Data ; Muscle Contraction ; Mutation ; Phenotype ; Sensation/genetics/*physiology ; Sodium Channels/chemistry/genetics/*physiology ; Touch/genetics/physiology
    Print ISSN: 0036-8075
    Electronic ISSN: 1095-9203
    Topics: Biology , Chemistry and Pharmacology , Computer Science , Medicine , Natural Sciences in General , Physics
    Published by American Association for the Advancement of Science (AAAS)
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  • 6
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    Primary cilia are not calcium-responsive mechanosensors (2016)
    Nature Publishing Group (NPG)
    Details
    Publication Date: 2016-03-24
    Description: Primary cilia are solitary, generally non-motile, hair-like protrusions that extend from the surface of cells between cell divisions. Their antenna-like structure leads naturally to the assumption that they sense the surrounding environment, the most common hypothesis being sensation of mechanical force through calcium-permeable ion channels within the cilium. This Ca(2+)-responsive mechanosensor hypothesis for primary cilia has been invoked to explain a large range of biological responses, from control of left-right axis determination in embryonic development to adult progression of polycystic kidney disease and some cancers. Here we report the complete lack of mechanically induced calcium increases in primary cilia, in tissues upon which this hypothesis has been based. We developed a transgenic mouse, Arl13b-mCherry-GECO1.2, expressing a ratiometric genetically encoded calcium indicator in all primary cilia. We then measured responses to flow in primary cilia of cultured kidney epithelial cells, kidney thick ascending tubules, crown cells of the embryonic node, kinocilia of inner ear hair cells, and several cell lines. Cilia-specific Ca(2+) influxes were not observed in physiological or even highly supraphysiological levels of fluid flow. We conclude that mechanosensation, if it originates in primary cilia, is not via calcium signalling.〈br /〉〈br /〉〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4851444/" target="_blank"〉〈img src="https://static.pubmed.gov/portal/portal3rc.fcgi/4089621/img/3977009" border="0"〉〈/a〉   〈a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4851444/" target="_blank"〉This paper as free author manuscript - peer-reviewed and accepted for publication〈/a〉〈br /〉〈br /〉〈span class="detail_caption"〉Notes: 〈/span〉Delling, M -- Indzhykulian, A A -- Liu, X -- Li, Y -- Xie, T -- Corey, D P -- Clapham, D E -- 5R01 DC000304/DC/NIDCD NIH HHS/ -- P30-HD 18655/HD/NICHD NIH HHS/ -- R01 DC000304/DC/NIDCD NIH HHS/ -- Howard Hughes Medical Institute/ -- England -- Nature. 2016 Mar 31;531(7596):656-60. doi: 10.1038/nature17426. Epub 2016 Mar 23.〈br /〉〈span class="detail_caption"〉Author address: 〈/span〉Department of Cardiology, Howard Hughes Medical Institute, Boston Children's Hospital, Boston, Massachusetts 02115, USA. ; Department of Neurobiology, Howard Hughes Medical Institute, Harvard Medical School, Boston, Massachusetts 02115, USA. ; Image and Data Analysis Core (IDAC), Harvard Medical School, Boston, Massachusetts 02115, USA.〈br /〉〈span class="detail_caption"〉Record origin:〈/span〉 〈a href="http://www.ncbi.nlm.nih.gov/pubmed/27007841" target="_blank"〉PubMed〈/a〉
    Keywords: Animals ; Calcium/analysis/*metabolism ; Calcium Signaling ; Cilia/*metabolism ; Embryo, Mammalian/cytology ; Epithelial Cells/cytology ; Female ; Hair Cells, Auditory, Inner/cytology ; Kidney/cytology ; Male ; *Mechanotransduction, Cellular ; Mice ; Mice, Transgenic ; Models, Biological
    Print ISSN: 0028-0836
    Electronic ISSN: 1476-4687
    Topics: Biology , Chemistry and Pharmacology , Medicine , Natural Sciences in General , Physics
    Published by Nature Publishing Group (NPG)
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  • 7
    Electronic Resource
    Electronic Resource
    Ion Channels in Vertebrate Glia (1990)
    Palo Alto, Calif. : Annual Reviews
    Annual Review of Neuroscience 13 (1990), S. 441-474 
    Details
    ISSN: 0147-006X
    Source: Annual Reviews Electronic Back Volume Collection 1932-2001ff
    Topics: Biology , Medicine
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1146/annurev.ne.13.030190.002301
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  • 8
    Electronic Resource
    Electronic Resource
    A reinterpretation of mammalian sodium channel gating based on single channel recording (1983)
    [s.l.] : Nature Publishing Group
    Nature 306 (1983), S. 436-441 
    Details
    ISSN: 1476-4687
    Source: Nature Archives 1869 - 2009
    Topics: Biology , Chemistry and Pharmacology , Medicine , Natural Sciences in General , Physics
    Notes: [Auszug] Some of the traditionally held views about how sodium channels work are shown to be incorrect and a new approach to physical theories of sodium channel operation is ...
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1038/306436a0
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  • 9
    Electronic Resource
    Electronic Resource
    Ionic basis of the receptor potential in a vertebrate hair cell (1979)
    [s.l.] : Nature Publishing Group
    Nature 281 (1979), S. 675-677 
    Details
    ISSN: 1476-4687
    Source: Nature Archives 1869 - 2009
    Topics: Biology , Chemistry and Pharmacology , Medicine , Natural Sciences in General , Physics
    Notes: [Auszug] Hair cells were prepared for intracellular recording as described previously510. Sacculi were removed from adult bullfrogs and their otolithic membranes peeled away after loosening by mild proteolysis (incubation for 60 min with 0.03mg ml'1 subtilopeptidase A, EC 3.4.4.16, at 22 C). The tissue was ...
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1038/281675a0
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  • 10
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    Actin cores of hair-cell stereocilia support myosin motility. (1990)
    National Academy of Sciences
    Details
    Publication Date: 1990-11-01
    Print ISSN: 0027-8424
    Electronic ISSN: 1091-6490
    Topics: Biology , Medicine , Natural Sciences in General
    Published by National Academy of Sciences
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