The Roadblock Light Chain Binds a Novel Region of the Cytoplasmic Dynein Intermediate Chain

Susalka, Stephen J.; Nikulina, Karina; Salata, Mark W.; Vaughan, Patricia S.; King, Stephen M.; Vaughan, Kevin T.; Pfister, Karin

doi:10.1074/jbc.m205510200

Cited by 72 publications

(73 citation statements)

References 65 publications

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“…Yeast Two-hybrid Studies-Pairwise yeast two-hybrid analysis was performed as described previously with slight modification (16,34). Briefly, AH109 yeast strains were co-transformed with the constructs as indicated in the figures according to the manufacturer's instructions (Clontech).…”

Section: Methodsmentioning

confidence: 99%

Interaction of the DYNLT (TCTEX1/RP3) Light Chains and the Intermediate Chains Reveals Novel Intersubunit Regulation during Assembly of the Dynein Complex

Kogoy

Rasoul

et al. 2007

Journal of Biological Chemistry

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Cytoplasmic dynein 1 is a microtubule-based molecular motor that functions to generate force for cargo transport to microtubule minus-ends (1-3). It is involved in numerous eukaryotic cell processes including the trafficking of membranous vesicles, viruses, and other intracellular particles. Cytoplasmic dynein 1 is a large multisubunit complex (ϳ1.5 MDa) containing two copies of six subunits, the heavy chain (DYNC1H), the intermediate chain (DYNC1I), the light intermediate chain (DYNC1LI), and three distinct light chains, DYNLT (previously called Tctex1), DYNLRB (previously called roadblock), and DYNLL (previously called LC8) (2, 4 -6). The motor domains of cytoplasmic dynein 1 are located in the C-terminal globular heads of the two identical heavy chains Whereas there is only a single heavy chain isoform, there are multiple isoforms of the five subunits that make up the cargo binding domain (2,3,17). In mammals, at least six intermediate chain isoforms are produced by the alternative splicing of two genes, and there are at least two genes for each of the other four subunits (2, 3, 5, 18). Assembly of individual subunit isoforms into the dynein complex creates different populations of the motor protein that are thought to be involved in specific cargo binding and regulation (2, 19 -22). For example, pericentrin is transported to the centrosome exclusively by the dynein complexes that contain the light intermediate chain isoform DYNC1LI-1 (23). The DYNLT and DYNLL light chains have been shown to interact with numerous functionally unrelated proteins (3, 14, 15, 24 -27).The two members of the DYNLT family, DYNLT1 (previously called Tctex1) 2 and DYNLT3 (previously called rp3) are found in all cultured cells and adult and fetal tissues so far examined (19,28,29). Unlike the DYNLT isoforms, the expression of the six intermediate chains isoforms are tissue and cell type-specific (18, 30 -32). One intermediate chain isoform, DYNC1I-2C (IC-2C), 3 is found in all cells and it is often the only isoform found in cultured cells (18). Most tissues express only * This work was supported by National Institutes of Health NINDS Grants NS29996 (to K. K. P.) and GM 51293 (to S. M. K.). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact. 1 To whom correspondence should be addressed: P. O. Box 800732, University of Virginia, Charlottesville, VA 22908-0732. Tel.: 434-924-1912; Fax: 434-982-3912; E-mail: kkp9w@virginia.edu.2 Cytoplasmic dynein 1 light chain and intermediate chain subunit nomenclature. There are three functionally distinct light chain families in the cytoplasmic dynein complex and each family has at least two members (or isoforms). The names of the light chains all begin with DYN for dynein, followed by L for light chain, then additional letters that designate the families. The family names are based on the old common name of the first identifie...

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Section: Methodsmentioning

confidence: 99%

Interaction of the DYNLT (TCTEX1/RP3) Light Chains and the Intermediate Chains Reveals Novel Intersubunit Regulation during Assembly of the Dynein Complex

Kogoy

Rasoul

et al. 2007

Journal of Biological Chemistry

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“…The precise order of association sites on IC1 and within the oda6-r88-defective region of IC2 is speculative. Analysis of other dyneins has suggested that members of the LC7/Roadblock family interact with IC regions either N-terminal to the first WD-repeat (Susalka et al, 2002) or C-terminal of the last repeat (Hendrickson et al, 2004); due to the interlocking design of the WD-repeat ␤-propeller, these regions are spatially close together. Maximal distances (in angstroms) between components are based on DMP and EDC cross-linking (King et al, 1991;DiBella et al, 2001DiBella et al, , 2004a.…”

Section: Heavy Chain-independent Assembly Of the Motor Domain-associamentioning

confidence: 99%

“…Analysis of cytoplasmic dynein has suggested that an IC region immediately N-terminal of the first WD-repeat (or possibly including the repeat) is necessary (Susalka et al, 2002), whereas study of the bop5 mutant version of IC138 within Chlamydomonas inner arm I1 has implicated the seventh WD-repeat and/or the C-terminal region in the interaction of LC7b with that dynein (Hendrickson et al, 2004). Due to the interlocking assembly of the WD-repeat ␤-propeller (the first ␤ strand of repeat #1 hydrogen bonds to the last strand of repeat #7; see, e.g., Sondek et al, 1996), these regions are likely in proximity at the tertiary structure level and may both contribute to LC7/ roadblock LC binding (Hendrickson et al, 2004).…”

Section: Implications For Intermediate Chain/light Chain Complex Archmentioning

confidence: 99%

Differential Light Chain Assembly Influences Outer Arm Dynein Motor Function

DiBella

Gorbatyuk

Sakato

et al. 2005

MBoC

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Tctex1 and Tctex2 were originally described as potential distorters/sterility factors in the non-Mendelian transmission of t-haplotypes in mice. These proteins have since been identified as subunits of cytoplasmic and/or axonemal dyneins. Within the Chlamydomonas flagellum, Tctex1 is a subunit of inner arm I1. We have now identified a second Tctex1-related protein (here termed LC9) in Chlamydomonas. LC9 copurifies with outer arm dynein in sucrose density gradients and is missing only in those strains completely lacking this motor. Zero-length cross-linking of purified outer arm dynein indicates that LC9 interacts directly with both the IC1 and IC2 intermediate chains. Immunoblot analysis revealed that LC2, LC6, and LC9 are missing in an IC2 mutant strain (oda6-r88) that can assemble outer arms but exhibits significantly reduced flagellar beat frequency. This defect is unlikely to be due to lack of LC6, because an LC6 null mutant (oda13) exhibits only a minor swimming abnormality. Using an LC2 null mutant (oda12-1), we find that although some outer arm dynein components assemble in the absence of LC2, they are nonfunctional. In contrast, dyneins from oda6-r88, which also lack LC2, retain some activity. Furthermore, we observed a synthetic assembly defect in an oda6-r88 oda12-1 double mutant. These data suggest that LC2, LC6, and LC9 have different roles in outer arm assembly and are required for wild-type motor function in the Chlamydomonas flagellum.

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“…Varma and R. Vallee, unpublished results) were suitable for the chick DRG neurons, which remained responsive to acute addition of laminin for only ϳ18 h. Instead, we injected the cells with dynein and LIS1 function-blocking antibodies. The antidynein antibody reacts with the N-terminal 63 aa of the IC polypeptide (Susalka et al, 2002; K. Vaughan and R. Vallee, unpublished observations) and, like other antibodies that react within this region, is thought to block the interaction of the IC with the p150…”

Section: Inhibition Of Dynein and Lis1mentioning

confidence: 99%

Cytoplasmic Dynein and LIS1 Are Required for Microtubule Advance during Growth Cone Remodeling and Fast Axonal Outgrowth

Grabham¹,

Seale²,

Bennecib³

et al. 2007

J. Neurosci.

151

138

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Recent evidence has implicated dynein and its regulatory factors dynactin and LIS1 in neuronal and non-neuronal cell migration. In the current study we sought to test whether effects on neuronal cell motility might reflect, in part, a role for these proteins in the growth cone. In chick sensory neurons subjected to acute laminin treatment dynein, dynactin, and LIS1 were mobilized strikingly and rapidly to the leading edge of the growth cone, where they were seen to be associated with microtubules converging into the laminin-induced axonal outgrowths. To interfere acutely with LIS1 and dynein function and to minimize secondary phenotypic effects, we injected antibodies to these proteins just before axon initiation. Antibody to both proteins produced an almost complete block of laminin-induced growth cone remodeling and the underlying reorganization of microtubules. Penetration of microtubules into the peripheral zone of differentiating axonal growth cones was decreased dramatically by antibody injection, as judged by live analysis of enhanced green fluorescent proteintubulin and the microtubule tip-associated EB3 (end-binding protein 3). Dynein and LIS1 inhibition had no detectable effect on microtubule assembly but reduced the ability of microtubules to resist retrograde actin flow. In hippocampal neurons dynein, dynactin, and LIS1 were enriched in axonal growth cones at stage 3, and both growth cone organization and axon elongation were altered by LIS1 RNA interference. Together, our data indicate that dynein and LIS1 play a surprisingly prominent role in microtubule advance during growth cone remodeling associated with axonogenesis. These data may explain, in part, the role of these proteins in brain developmental disease and support an important role in diverse aspects of neuronal differentiation and nervous system development.

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The Roadblock Light Chain Binds a Novel Region of the Cytoplasmic Dynein Intermediate Chain

Cited by 72 publications

References 65 publications

Interaction of the DYNLT (TCTEX1/RP3) Light Chains and the Intermediate Chains Reveals Novel Intersubunit Regulation during Assembly of the Dynein Complex

Interaction of the DYNLT (TCTEX1/RP3) Light Chains and the Intermediate Chains Reveals Novel Intersubunit Regulation during Assembly of the Dynein Complex

Differential Light Chain Assembly Influences Outer Arm Dynein Motor Function

Cytoplasmic Dynein and LIS1 Are Required for Microtubule Advance during Growth Cone Remodeling and Fast Axonal Outgrowth

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