scholarly journals Interspecies conservation of outer arm dynein intermediate chain sequences defines two intermediate chain subclasses.

1995 ◽  
Vol 6 (6) ◽  
pp. 685-696 ◽  
Author(s):  
K Ogawa ◽  
R Kamiya ◽  
C G Wilkerson ◽  
G B Witman
Keyword(s):  
Sea Urchin ◽  
Related Species ◽  
Cytoplasmic Dynein ◽  
Cross Reactivity ◽  
The Other ◽  
Immunological Analysis ◽  
Dynein Intermediate Chain ◽  
Structural Differences ◽  
Intermediate Chains ◽  
Intermediate Chain

Immunological analysis showed that antibodies against the intermediate chains (ICs) IC2 and IC3 of sea urchin outer arm dynein specifically cross-reacted with intermediate chains IC78 and IC69, respectively, of Chlamydomonas outer arm dynein. In contrast, no specific cross-reactivity with any Chlamydomonas outer arm polypeptide was observed using antibody against IC1 of sea urchin outer arm dynein. To learn more about the relationships between the different ICs, overlapping cDNAs encoding all of IC2 and IC3 of sea urchin were isolated and sequenced. Comparison of these sequences with those previously obtained for the Chlamydomonas ICs revealed that, although all four chains are homologous, sea urchin IC2 is much more closely related to Chlamydomonas IC78 (45.8% identity), and sea urchin IC3 is much more closely related to Chlamydomonas IC69 (48.5% identity), than either sea urchin chain is related to the other (23.5% identity). For homologous pairs, the similarities extend throughout the full lengths of the chains. Regions of similarity between all four ICs and the IC (IC74) of cytoplasmic dynein, located in the C-terminal halves of the chains, are due primarily to conservation of the WD repeats present in all of these ICs. This is the first demonstration that structural differences between individual ICs within an outer arm dynein have been highly conserved in the dyneins of distantly related species. The results provide a basis for the subclassification of these chains.

scholarly journals Is outer arm dynein intermediate chain 1 multifunctional?

1996 ◽  
Vol 7 (12) ◽  
pp. 1895-1907 ◽  
Author(s):  
K Ogawa ◽  
H Takai ◽  
A Ogiwara ◽  
E Yokota ◽  
T Shimizu ◽  
...  
Keyword(s):  
Sea Urchin ◽  
Related Species ◽  
Repetitive Sequences ◽  
Middle Part ◽  
Affinity Column ◽  
Terminal Part ◽  
Related Part ◽  
Dynein Intermediate Chain ◽  
Intermediate Chains ◽  
Intermediate Chain

The outer arm dynein of sea urchin sperm axoneme contains three intermediate chains (IC1, IC2, and IC3; M(r) 128,000, 98,000, and 74,000, respectively). IC2 and IC3 are members of the WD family; the WD motif is responsible for a protein-protein interaction. We describe here the molecular cloning of IC1. IC1 has a unique primary structure, the N-terminal part is homologous to the sequence of thioredoxin, the middle part consists of three repetitive sequences homologous to the sequence of nucleoside diphosphate kinase, and the C-terminal part contains a high proportion of negatively charged glutamic acid residues. Thus, IC1 is a novel dynein intermediate chain distinct from IC2 and IC3 and may be a multifunctional protein. The thioredoxin-related part of IC1 is more closely related to those of two redox-active Chlamydomonas light chains than thioredoxin. Antibodies were prepared against the N-terminal and middle domains of IC1 expressed as His-tagged proteins in bacteria. These antibodies cross-reacted with some dynein polypeptides (potential homologues of IC1) from distantly related species. We propose here that the three intermediate chains are the basic core units of sperm outer arm dynein because of their ubiquitous existence. The recombinant thioredoxin-related part of IC1 and outer arm dyneins from sea urchin and distantly related species were specifically bound to and eluted from a phenylarsine oxide affinity column with 2-mercaptoethanol, indicating that they contain vicinal dithiols competent to undergo reversible oxidation/reduction.

scholarly journals Structure of Tctex-1 and Its Interaction with Cytoplasmic Dynein Intermediate Chain

2001 ◽  
Vol 276 (17) ◽  
pp. 14067-14074 ◽  
Author(s):  
Yu-Keung Mok ◽  
Kevin W.-H. Lo ◽  
Mingjie Zhang
Keyword(s):  
Cytoplasmic Dynein ◽  
Dynein Intermediate Chain ◽  
Intermediate Chain

scholarly journals PLAC-24 Is a Cytoplasmic Dynein-Binding Protein That Is Recruited to Sites of Cell-Cell Contact

2002 ◽  
Vol 13 (5) ◽  
pp. 1722-1734 ◽  
Author(s):  
Sher Karki ◽  
Lee A. Ligon ◽  
Jamison DeSantis ◽  
Mariko Tokito ◽  
Erika L. F. Holzbaur
Keyword(s):  
Epithelial Cells ◽  
Recent Observation ◽  
Polyclonal Antibodies ◽  
Adherens Junction ◽  
Cytoplasmic Dynein ◽  
Cell Contact ◽  
Dynein Intermediate Chain ◽  
Cellular Cytoskeleton ◽  
Cell Cell ◽  
Intermediate Chain

We screened for polypeptides that interact specifically with dynein and identified a novel 24-kDa protein (PLAC-24) that binds directly to dynein intermediate chain (DIC). PLAC-24 is not a dynactin subunit, and the binding of PLAC-24 to the dynein intermediate chain is independent of the association between dynein and dynactin. Immunocytochemistry using PLAC-24–specific polyclonal antibodies revealed a punctate perinuclear distribution of the polypeptide in fibroblasts and isolated epithelial cells. However, as epithelial cells in culture make contact with adjacent cells, PLAC-24 is specifically recruited to the cortex at sites of contact, where the protein colocalizes with components of the adherens junction. Disruption of the cellular cytoskeleton with latrunculin or nocodazole indicates that the localization of PLAC-24 to the cortex is dependent on intact actin filaments but not on microtubules. Overexpression of β-catenin also leads to a loss of PLAC-24 from sites of cell-cell contact. On the basis of these data and the recent observation that cytoplasmic dynein is also localized to sites of cell-cell contact in epithelial cells, we propose that PLAC-24 is part of a multiprotein complex localized to sites of intercellular contact that may function to tether microtubule plus ends to the actin-rich cellular cortex.

scholarly journals Somatic CRISPR–Cas9-induced mutations reveal roles of embryonically essential dynein chains in Caenorhabditis elegans cilia

2015 ◽  
Vol 208 (6) ◽  
pp. 683-692 ◽  
Author(s):  
Wenjing Li ◽  
Peishan Yi ◽  
Guangshuo Ou
Keyword(s):  
Caenorhabditis Elegans ◽  
Intraflagellar Transport ◽  
Cytoplasmic Dynein ◽  
Regulatory Mechanisms ◽  
Light Chains ◽  
Induced Mutations ◽  
Functional Studies ◽  
Cilium Formation ◽  
Dynein Intermediate Chain ◽  
Intermediate Chain

Cilium formation and maintenance require intraflagellar transport (IFT). Although much is known about kinesin-2–driven anterograde IFT, the composition and regulation of retrograde IFT-specific dynein remain elusive. Components of cytoplasmic dynein may participate in IFT; however, their essential roles in cell division preclude functional studies in postmitotic cilia. Here, we report that inducible expression of the clustered regularly interspaced short palindromic repeats (CRISPR)–Cas9 system in Caenorhabditis elegans generated conditional mutations in IFT motors and particles, recapitulating ciliary defects in their null mutants. Using this method to bypass the embryonic requirement, we show the following: the dynein intermediate chain, light chain LC8, and lissencephaly-1 regulate retrograde IFT; the dynein light intermediate chain functions in dendrites and indirectly contributes to ciliogenesis; and the Tctex and Roadblock light chains are dispensable for cilium assembly. Furthermore, we demonstrate that these components undergo biphasic IFT with distinct transport frequencies and turnaround behaviors. Together, our results suggest that IFT–dynein and cytoplasmic dynein have unique compositions but also share components and regulatory mechanisms.

scholarly journals The APC-associated protein EB1 associates with components of the dynactin complex and cytoplasmic dynein intermediate chain

1999 ◽  
Vol 9 (8) ◽  
pp. 425-428 ◽  
Author(s):  
Lisbeth Berrueta ◽  
Jennifer S. Tirnauer ◽  
Scott C. Schuyler ◽  
David Pellman ◽  
Barbara E. Bierer
Keyword(s):  
Cytoplasmic Dynein ◽  
Dynein Intermediate Chain ◽  
Dynactin Complex ◽  
Intermediate Chain

scholarly journals The M r 140,000 Intermediate Chain ofChlamydomonas Flagellar Inner Arm Dynein Is a WD-Repeat Protein Implicated in Dynein Arm Anchoring

1998 ◽  
Vol 9 (12) ◽  
pp. 3335-3349 ◽  
Author(s):  
Pinfen Yang ◽  
Winfield S. Sale
Keyword(s):  
Fusion Protein ◽  
Protein A ◽  
Dynein Intermediate Chain ◽  
In Vitro Reconstitution ◽  
Unique Site ◽  
Intermediate Chains ◽  
Β Sheet ◽  
Chemical Cross Linking ◽  
Intermediate Chain

Previous structural and biochemical studies have revealed that the inner arm dynein I1 is targeted and anchored to a unique site located proximal to the first radial spoke in each 96-nm axoneme repeat on flagellar doublet microtubules. To determine whether intermediate chains mediate the positioning and docking of dynein complexes, we cloned and characterized the 140-kDa intermediate chain (IC140) of the I1 complex. Sequence and secondary structural analysis, with particular emphasis on β-sheet organization, predicted that IC140 contains seven WD repeats. Reexamination of other members of the dynein intermediate chain family of WD proteins indicated that these polypeptides also bear seven WD/β-sheet repeats arranged in the same pattern along each intermediate chain protein. A polyclonal antibody was raised against a 53-kDa fusion protein derived from the C-terminal third of IC140. The antibody is highly specific for IC140 and does not bind to other dynein intermediate chains or proteins in Chlamydomonasflagella. Immunofluorescent microscopy of Chlamydomonascells confirmed that IC140 is distributed along the length of both flagellar axonemes. In vitro reconstitution experiments demonstrated that the 53-kDa C-terminal fusion protein binds specifically to axonemes lacking the I1 complex. Chemical cross-linking indicated that IC140 is closely associated with a second intermediate chain in the I1 complex. These data suggest that IC140 contains domains responsible for the assembly and docking of the I1 complex to the doublet microtubule cargo.

scholarly journals The Gene for the Intermediate Chain Subunit of Cytoplasmic Dynein Is Essential in Drosophila

Genetics ◽  
2002 ◽  
Vol 162 (3) ◽  
pp. 1211-1220 ◽  
Author(s):  
Kristin L M Boylan ◽  
Thomas S Hays
Keyword(s):  
Intracellular Transport ◽  
De Novo ◽  
Cytoplasmic Dynein ◽  
Chain Gene ◽  
Temperature Sensitive ◽  
Wing Development ◽  
Dynein Motor ◽  
Dynein Intermediate Chain ◽  
Microtubule Motor ◽  
Intermediate Chain

Abstract The microtubule motor cytoplasmic dynein powers a variety of intracellular transport events that are essential for cellular and developmental processes. A current hypothesis is that the accessory subunits of the dynein complex are important for the specialization of cytoplasmic dynein function. In a genetic approach to understanding the range of dynein functions and the contribution of the different subunits to dynein motor function and regulation, we have identified mutations in the gene for the cytoplasmic dynein intermediate chain, Dic19C. We used a functional Dic transgene in a genetic screen to recover X-linked lethal mutations that require this transgene for viability. Three Dic mutations were identified and characterized. All three Dic alleles result in larval lethality, demonstrating that the intermediate chain serves an essential function in Drosophila. Like a deficiency that removes Dic19C, the Dic mutations dominantly enhance the rough eye phenotype of Glued1, a dominant mutation in the gene for the p150 subunit of the dynactin complex, a dynein activator. Additionally, we used complementation analysis to identify an existing mutation, shortwing (sw), as an allele of the dynein intermediate chain gene. Unlike the Dic alleles isolated de novo, shortwing is homozygous viable and exhibits recessive and temperature-sensitive defects in eye and wing development. These phenotypes are rescued by the wild-type Dic transgene, indicating that shortwing is a viable allele of the dynein intermediate chain gene and revealing a novel role for dynein function during wing development.

Activation of Human Platelets Causes Post-translational Modifications to Cytoplasmic Dynein

1997 ◽  
Vol 78 (02) ◽  
pp. 910-918 ◽  
Author(s):  
Stephen W Rothwell ◽  
Valerie S Calvert
Keyword(s):  
Molecular Motors ◽  
Soluble Fraction ◽  
Cytoplasmic Dynein ◽  
Human Platelets ◽  
Particulate Fraction ◽  
Cortical Region ◽  
Transient Time ◽  
Post Translational Modifications ◽  
Dynein Intermediate Chain ◽  
Intermediate Chain

SummaryIn our studies of human platelets we have detected the presence of the molecular motors kinesin and dynein. Dynein is present at a concentration (0.8 μg/g tissue) that is approximately 1/3 the concentration reported for neuronal tissue. Immunofluorescence microscopy of resting platelets shows that, while platelet microtubules are arranged in coiled hoops forming the marginal band in the cortical region of the platelet, dynein is distributed in a pattern of punctate staining throughout the cytoplasm of the platelets. Fractionation of unactivated platelets shows that dynein partitions to the soluble fraction. Stimulation of platelets with thrombin, ADP or epinephrine causes a partial translocation of dynein from the soluble fraction to the particulate fraction with thrombin being the most efficient agent at promoting this shift. Dynein intermediate chain recovered in the soluble fraction of disrupted platelets following activation displays a transient, time-dependent phosphorylation. In contrast, dynein intermediate chain recovered in the particulate fraction shows decreased phosphorylation. These results indicate that human platelets contain a complex microtubule-based system of motor proteins that is an integral part of the physiological changes occurring during platelet activation.

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