Brain dynein crossbridges microtubules into bundles

1989 ◽  
Vol 93 (1) ◽  
pp. 19-28 ◽  
Author(s):  
L.A. Amos
Keyword(s):  
Cytoplasmic Dynein ◽  
Molecular Forms ◽  
Mammalian Brain ◽  
Structural Homology ◽  
Greek Letter ◽  
Major Band ◽  
Negative Stain ◽  
Globular Particle ◽  
Axonemal Dynein ◽  
Individual Particles

Cytoplasmic dynein was purified from pig brain, using a modified version of published procedures, in order to study its interaction with microtubules. Since the preparation produces ATP-dependent sliding of taxol-stabilized purified microtubules over glass and runs on SDS-containing gels as a major band exceeding 300,000 Mr plus a medium chain band at about 75,000 Mr, it is assumed to be identical to the mammalian brain dynein (MAP 1C) purified by Vallee and colleagues. When viewed by electron microscopy in negative stain, individual particles show two distinct configurations. Some are clearly similar to the two-headed bouquet structure already shown for MAP 1C. A larger number of molecules in the present preparation appear to have two heads fused together, forming a dimeric globular particle with two separate tails. They are referred to as phiparticles, because of their resemblance to the greek letter phi. A model for the structural relationship between the two molecular forms is presented. The stems of two associated dynein subunits may separate beyond the base, to form a bouquet, or they may remain fused to form the larger tail of a phi-particle. The smaller tail probably represents a combined pair of features equivalent to the ‘stalks’ shown to emanate from axonemal dynein heads by Goodenough and colleagues. Both tails of a phi-particle can bind to microtubules, even in the presence of ATP, and cause microtubule bundling. These results suggest a complete structural homology between axonemal and cytoplasmic dynein.

scholarly journals PURIFICATION OF INTACT MICROTUBULES FROM BRAIN

1970 ◽  
Vol 47 (2) ◽  
pp. 384-394 ◽  
Author(s):  
Joel B. Kirkpatrick ◽  
Lyon Hyams ◽  
Virginia L. Thomas ◽  
Peter M. Howley
Keyword(s):  
Low Temperature ◽  
Gel Electrophoresis ◽  
Combined Effects ◽  
Acidic Ph ◽  
Electron Microscopic ◽  
Major Band ◽  
Negative Stain ◽  
Hexylene Glycol ◽  
Electrophoresis System ◽  
Microtubule Structure

Hundred-fold purification of intact microtubules from homogenates of rat brain is reported. The success of purification depends on stabilizing the microtubule structure by the combined effects of hexylene glycol, acidic Ph, and low temperature. A practical, negative stain, electron microscopic assay is used to study purity and stability of microtubule fractions. The purified fractions show a major band which migrates like purified tubulin in the SDS gel electrophoresis system.

A novel cytoplasmic dynein heavy chain: expression of DHC1b in mammalian ciliated epithelial cells

1996 ◽  
Vol 109 (7) ◽  
pp. 1891-1898 ◽  
Author(s):  
P.S. Criswell ◽  
L.E. Ostrowski ◽  
D.J. Asai
Keyword(s):  
Heavy Chain ◽  
Minor Component ◽  
Cytoplasmic Dynein ◽  
Heart Cells ◽  
Rat Tissues ◽  
Dynein Heavy Chain ◽  
Undifferentiated Cells ◽  
Tracheal Epithelial ◽  
Axonemal Dynein ◽  
Ciliated Epithelial Cells

Organisms that have cilia or flagella express over a dozen dynein heavy chain genes. Of these heavy chain genes, most appear to encode axonemal dyneins, one encodes conventional cytoplasmic dynein (MAP1C or DHC1a), and one, here referred to as DHC1b, encodes an unclassified heavy chain. Previous analysis of sea urchin DHC1b (Gibbons et al. (1994) Mol. Biol. Cell 5, 57–70) indicated that this isoform is either an axonemal dynein with an unusual protein sequence or a cytoplasmic dynein whose expression increases during ciliogenesis. In the present study, we examined the expression of DHC1b in rat tissues. The DHC1b gene is expressed in all tissues examined, including unciliated liver and heart cells. In contrast, rat axonemal dyneins are only expressed in tissues that produce cilia or flagella. In cultured rat tracheal epithelial (RTE) cells, DHC1b is expressed in undifferentiated cells and increases in expression during ciliogenesis. In contrast, the expression of conventional cytoplasmic dynein, DHC1a, does not change during RTE differentiation and axonemal dynein is not expressed until after differentiation commences. In order to examine the expression of DHC1b protein, we produced an isoform-specific antibody to a synthetic peptide derived from the rat DHC1b sequence. The antibody demonstrated that DHC1b is a relatively minor component of partially purified cytoplasmic dynein. Indirect immunofluorescence microscopy revealed that DHC1b is not detected in cilia and remains in the cytoplasm of ciliated RTE cells, often accumulating at the apical ends of the cells. These results suggest that DHC1b is a cytoplasmic dynein that may participate in intracellular trafficking in polarized cells.

scholarly journals Opioid receptors in GtoPdb v.2021.3

2021 ◽  
Vol 2021 (3) ◽  
Author(s):  
Anna Borsodi ◽  
Michael Bruchas ◽  
Girolamo Caló ◽  
Charles Chavkin ◽  
MacDonald J. Christie ◽  
...  
Keyword(s):  
Opioid Receptors ◽  
Opioid Analgesics ◽  
Structural Homology ◽  
Orphanin Fq ◽  
Dynorphin A ◽  
Greek Letter ◽  
Endogenous Peptides ◽  
High Degree ◽  
Μ Receptor

Opioid and opioid-like receptors are activated by a variety of endogenous peptides including [Met]enkephalin (met), [Leu]enkephalin (leu), β-endorphin (β-end), α-neodynorphin, dynorphin A (dynA), dynorphin B (dynB), big dynorphin (Big dyn), nociceptin/orphanin FQ (N/OFQ); endomorphin-1 and endomorphin-2 are also potential endogenous peptides. The Greek letter nomenclature for the opioid receptors, μ, δ and κ, is well established, and NC-IUPHAR considers this nomenclature appropriate, along with the symbols spelled out (mu, delta, and kappa), and the acronyms, MOP, DOP, and KOP. [121, 100, 91]. The human N/OFQ receptor, NOP, is considered 'opioid-related' rather than opioid because, while it exhibits a high degree of structural homology with the conventional opioid receptors [294], it displays a distinct pharmacology. Currently there are numerous clinically used drugs, such as morphine and many other opioid analgesics, as well as antagonists such as naloxone, however only for the μ receptor.

scholarly journals Opioid receptors (version 2019.4) in the IUPHAR/BPS Guide to Pharmacology Database

2019 ◽  
Vol 2019 (4) ◽  
Author(s):  
Anna Borsodi ◽  
Michael Bruchas ◽  
Girolamo Caló ◽  
Charles Chavkin ◽  
MacDonald J. Christie ◽  
...  
Keyword(s):  
Opioid Receptors ◽  
Opioid Analgesics ◽  
Structural Homology ◽  
Orphanin Fq ◽  
Dynorphin A ◽  
Greek Letter ◽  
Endogenous Peptides ◽  
High Degree ◽  
Μ Receptor

Opioid and opioid-like receptors are activated by a variety of endogenous peptides including [Met]enkephalin (met), [Leu]enkephalin (leu), β-endorphin (β-end), α-neodynorphin, dynorphin A (dynA), dynorphin B (dynB), big dynorphin (Big dyn), nociceptin/orphanin FQ (N/OFQ); endomorphin-1 and endomorphin-2 are also potential endogenous peptides. The Greek letter nomenclature for the opioid receptors, μ, δ and κ, is well established, and NC-IUPHAR considers this nomenclature appropriate, along with the symbols spelled out (mu, delta, and kappa), and the acronyms, MOP, DOP, and KOP. [116, 96, 88]. The human N/OFQ receptor, NOP, is considered 'opioid-related' rather than opioid because, while it exhibits a high degree of structural homology with the conventional opioid receptors [282], it displays a distinct pharmacology. Currently there are numerous clinically used drugs, such as morphine and many other opioid analgesics, as well as antagonists such as naloxone, however only for the μ receptor.

Developmental regulation of two microtubule-associated proteins (MAP2 and MAP5) in the embryonic avian retina

Development ◽  
1987 ◽  
Vol 101 (3) ◽  
pp. 535-546
Author(s):  
R.P. Tucker ◽  
A.I. Matus
Keyword(s):  
Ganglion Cell ◽  
Developmental Regulation ◽  
Neonatal Rat ◽  
Molecular Forms ◽  
Mammalian Brain ◽  
Cellular Distribution ◽  
Inner Plexiform Layer ◽  
Western Blots ◽  
Microtubule Associated Proteins ◽  
Associated Proteins

Previous studies with the mammalian brain have shown that the expression of a number of neuronal microtubule-associated proteins (MAPs) is developmentally regulated. For example, the low-molecular-weight form of MAP2 (MAP2c) is abundant in neonatal rat brains and is less abundant in adults. Similarly, MAP5 levels decrease during postnatal development. Using monoclonal antibodies, we have followed the time of first appearance, cellular distribution, and molecular form of MAP2 and MAP5 during the morphogenesis of the quail retina. MAP2 first appears in ganglion cell bodies and in the axons of the optic fibre layer (OFL) at embryonic day 4 (E4). Anti-MAP2 staining remains restricted to these sites until E10, when staining appears in the inner plexiform layer (IPL). At E14, one day before hatching, anti-MAP2 staining is found in three broad laminae in the IPL, as well as in photosensitive cells. MAP5 is present in ganglion cell axons from the onset of neurite elongation at E3 and is limited to the OFL until E10. The intensity of anti-MAP5 staining in the OFL and optic nerve decreases after E7, which corresponds with a decrease in the number of actively growing ganglion cell axons. By E14, anti-MAP5 stains five layers in the IPL that correspond with layers of amacrine cell process arborizations. Western blots of E10 brain microtubule proteins show that MAP2 is represented by both a 260 × 10(3) Mr protein and a 60–65 × 10(3) Mr protein; the latter is much more abundant. Anti-MAP5 recognizes a 320 × 10(3) Mr brain microtubule protein in both the quail and the rat. We conclude that the cellular distribution, developmental regulation and molecular forms of MAP2 and MAP5 are similar in the rat and quail, suggesting that these molecules have conserved and hence fundamental roles in the growth and differentiation of neuronal processes.

scholarly journals Studies on the Structure and Subunit Composition of Human Antihaemophilic Factor

1977 ◽  
Author(s):  
J. J. Gorman
Keyword(s):  
Molecular Weight ◽  
Factor Viii ◽  
Peptide Mapping ◽  
Sodium Dodecyl ◽  
Gel Filtration ◽  
Protein S ◽  
Molecular Forms ◽  
Major Band ◽  
Human Factor Viii ◽  
Von Willebrand

Human antihaemophilic factor has been purified by hydroxylapatite chromatography following precipitation from plasma and gel filtration on Sepharose 6B.Application to hydroxyiapatite was in 0.02 M tris HCl (pH 7.35) – 0.14 M NaCl and after washing with 5mM phosphate (pH 6.8) – 0.1 M NaCI the antihaemophilic factor was eluted with 0. 1M phosphate (pH 6.8) – 0.1M NaCl. Factor VIII coagulant activity, factor VIII related antigen and von Willebrand factor activity eluted simultaneously.The protein(s) had a molecular weight in excess of 500,000 and multiple subunits as shown by electrophoresis in 5% acrylamide gels containing sodium dodecyl sulphate;without reduction the protein failed to enter these gels but following reduction multiple bands were observed, the major band had a molecular weight around 200,000.Thin layer peptide mapping demonstrated structural inter-relationship between the 200,000 dalton protein and three of the smaller species, however, two other unrelated smaller species were evident.It is apparent from these findings that human factor VIII may exist as multiple molecular forms due to heterogeneity of one subunit (MW around 200,000) and the molecular structure may include other smaller non-identical subunits. The structure-function relationships of these subunits remains to be elucidated.

scholarly journals Ciliary Dyneins and Dynein Related Ciliopathies

Cells ◽  
2021 ◽  
Vol 10 (8) ◽  
pp. 1885
Author(s):  
Dinu Antony ◽  
Han G. Brunner ◽  
Miriam Schmidts
Keyword(s):  
Intraflagellar Transport ◽  
Cytoplasmic Dynein ◽  
Vertebrate Development ◽  
Kartagener Syndrome ◽  
Dynein Motor ◽  
Axonemal Dynein ◽  
Ciliary Dyskinesia ◽  
Laterality Defects ◽  
Renal Phenotype ◽  
Ciliary Localization

Although ubiquitously present, the relevance of cilia for vertebrate development and health has long been underrated. However, the aberration or dysfunction of ciliary structures or components results in a large heterogeneous group of disorders in mammals, termed ciliopathies. The majority of human ciliopathy cases are caused by malfunction of the ciliary dynein motor activity, powering retrograde intraflagellar transport (enabled by the cytoplasmic dynein-2 complex) or axonemal movement (axonemal dynein complexes). Despite a partially shared evolutionary developmental path and shared ciliary localization, the cytoplasmic dynein-2 and axonemal dynein functions are markedly different: while cytoplasmic dynein-2 complex dysfunction results in an ultra-rare syndromal skeleto-renal phenotype with a high lethality, axonemal dynein dysfunction is associated with a motile cilia dysfunction disorder, primary ciliary dyskinesia (PCD) or Kartagener syndrome, causing recurrent airway infection, degenerative lung disease, laterality defects, and infertility. In this review, we provide an overview of ciliary dynein complex compositions, their functions, clinical disease hallmarks of ciliary dynein disorders, presumed underlying pathomechanisms, and novel developments in the field.

scholarly journals Cryo-EM of dynein microtubule-binding domains shows how an axonemal dynein distorts the microtubule

eLife ◽  
2019 ◽  
Vol 8 ◽  
Author(s):  
Samuel E Lacey ◽  
Shaoda He ◽  
Sjors HW Scheres ◽  
Andrew P Carter
Keyword(s):  
Conformational Changes ◽  
Coiled Coil ◽  
Cytoplasmic Dynein ◽  
Cross Sectional ◽  
Microtubule Binding ◽  
High Affinity ◽  
Binding Domains ◽  
Microtubule Binding Domain ◽  
Axonemal Dynein ◽  
Cilia And Flagella

Dyneins are motor proteins responsible for transport in the cytoplasm and the beating of axonemes in cilia and flagella. They bind and release microtubules via a compact microtubule-binding domain (MTBD) at the end of a coiled-coil stalk. We address how cytoplasmic and axonemal dynein MTBDs bind microtubules at near atomic resolution. We decorated microtubules with MTBDs of cytoplasmic dynein-1 and axonemal dynein DNAH7 and determined their cryo-EM structures using helical Relion. The majority of the MTBD is rigid upon binding, with the transition to the high-affinity state controlled by the movement of a single helix at the MTBD interface. DNAH7 contains an 18-residue insertion, found in many axonemal dyneins, that contacts the adjacent protofilament. Unexpectedly, we observe that DNAH7, but not dynein-1, induces large distortions in the microtubule cross-sectional curvature. This raises the possibility that dynein coordination in axonemes is mediated via conformational changes in the microtubule.

A Tetragonal Lattice in Mammalian Lens Junctions

1981 ◽  
Vol 39 ◽  
pp. 632-633
Author(s):  
J.D. Robertson ◽  
G. Zampighi ◽  
S. Simon ◽  
T.J. McIntosh ◽  
M.J. Costello
Keyword(s):  
Freeze Fracture ◽  
X Ray Diffraction ◽  
Major Band ◽  
Direction Parallel ◽  
X Ray ◽  
Negative Stain ◽  
Tetragonal Lattice ◽  
Section Electron ◽  
A Minor ◽  
Ray Patterns

A pure fraction of junctional membranes has been isolated in milligram quantities from bovine lens without detergents. SDS-PAGE analysis shows a major band at 27,000 d and a minor one at 21,000 d. Thin section electron microscopy (EM) of this fraction revealed junctions of two closely apposed membranes with an overall thickness of 135-140 Å with a wavy contour and lengths greater than 10 μm. Negative stain and freeze-fracture (FF) EM (only unetched specimens so far) and X-ray diffraction studies of unfixed and unglycerinated specimens were in agreement regarding the structure of the junctions. They consisted primarily of extensive domains of sub-units arranged in a 66 Å tetragonal lattice. Specimens were spun down centrifugally in a Beem capsule and partially dried for the x-ray diffraction studies. The X-ray patterns contained several reflections which indexed on a tetragonal lattice of 66 Å in the equatorial direction parallel to the plane of the membranes and several bands of a continuous transform in the meridional direction consistent with membrane pairs. No diffraction was seen at 85 Å in the equatorial direction.

Distribution of cytoplasmic and axonemal dyneins in rat tissues

1992 ◽  
Vol 101 (3) ◽  
pp. 579-587
Author(s):  
T. Yoshida ◽  
H. Takanari ◽  
K. Izutsu
Keyword(s):  
Cytoplasmic Dynein ◽  
Cross Reactivity ◽  
Bovine Brain ◽  
Secretory Cells ◽  
Rat Tissues ◽  
Antibody Staining ◽  
Type Ii Pneumocytes ◽  
Neuronal Tissues ◽  
Axonemal Dynein ◽  
Cilia And Flagella

Microtubule-associated protein 1C (MAP 1C) is now defined as brain cytoplasmic dynein. Recent studies have suggested that cytoplasmic dynein is a motor protein responsible for the intracellular microtubule-based motility in neuronal and non-neuronal cells. We have prepared an antibody against bovine brain MAP 1C and have examined the localizations of cytoplasmic dynein in rat tissues. Immunoblots of extracts from the tissues showed that the dynein was present in brain, testis, liver, kidney and lung. Immunohistochemical experiments have demonstrated that dynein is localized in Purkinje cells of cerebellum and axons of central and peripheral nervous systems. In non-neuronal tissues, the antibody staining was intense in many types of cells, such as hepatocytes, epithelia of renal convoluted tubules, secretory cells of adrenal medulla and spermatids. Glomeruli of kidney, bronchial epithelia and type II pneumocytes of lung, pancreatic islets and acini, adrenal cortex and Sertoli cells were moderately positive upon exposure to the cytoplasmic dynein antibody. On the other hand, the localization of axonemal dynein was examined using antibodies against flagellar dynein of sea urchin spermatozoa. Anti-axonemal dynein labeled cilia and flagella in rat tissues whereas anti-MAP 1C did not stain axonemes. We also tested for immunological cross-reactivity between cytoplasmic and axonemal dyneins to probe for molecular similarities. Anti-axonemal dynein reacted with MAP 1C weakly. These results have confirmed that cytoplasmic dyneins are distributed widely among rat organs, not only in neuronal but also in non-neuronal tissues. There is no similarity in the localization of cytoplasmic and axonemal dyneins but there is some similarity in molecular antigenicity.

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