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Richard B Vallee

Publications and source records attributed to Richard B Vallee.

At least 19 recordsLinked to original sources
PubMed2007

Expression patterns of LIS1, dynein and their interaction partners dynactin, NudE, NudEL and NudC in human gliomas suggest roles in invasion and proliferation.

Diffusely infiltrating gliomas are the most common type of primary intracranial neoplasm in humans. One of the major obstacles to the effective treatment of these tumors is their highly infiltrative growth. However, mechanisms controlling their migration and proliferation are poorly understood. Glioma cells resemble neural progenitors, and we hypothesize that gliomas recapitulate the capacity of migration and proliferation of progenitors that takes place during brain development. Based on recent evidence implicating cytoplasmic dynein and its regulatory proteins in neural progenitor migration and division, we conducted immunohistochemical evaluation of surgically resected human glioma samples for the presence and distribution of these proteins. We examined expression of LIS1, the gene responsible for type I lissencephaly, cytoplasmic dynein and the dynein- and LIS1-interacting factors dynactin, NudE/NudEL and NudC, which play significant roles in neural progenitor cell behavior. We found that each of these proteins is expressed in all histological types and grades of human neuroectodermal tumors examined. Immunohistochemical analysis revealed that the levels of expression varied from cell to cell within each tumor, ranging from very high to undetectable. This stands in contrast to the low levels of diffuse staining seen in non-neoplastic brain tissue. Of particular interest, we noted tumor cells infiltrating the white matter and tumor cells undergoing cell division amongst the cells with notably high expression levels. These findings are compatible with the idea that LIS1 and its interacting proteins play a role in glioma migration and proliferation analogous to their role during brain development.

1-Alkyl-2-acetylglycerophosphocholine Esterase↗
PubMed2006

ZW10 function in mitotic checkpoint control, dynein targeting and membrane trafficking: is dynein the unifying theme?

ZW10 was initially identified as a mitotic checkpoint protein involved in chromosome segregation. It was subsequently implicated in targeting cytoplasmic dynein and dynactin to mitotic kinetochores, though the relationship between these functions remains incompletely understood. Recent studies have revealed that ZW10 performs important functions in nondividing cells as well. These include cytoplasmic dynein targeting to Golgi and other membranes, but also SNARE-mediated ER-Golgi trafficking. Identifying a unifying function for ZW10 in these diverse contexts has been elusive, but likely involves cytoplasmic dynein, as discussed here.

Animals↗
PubMed2006

Autoinhibitory and other autoregulatory elements within the dynein motor domain.

The dyneins are a family of microtubule motor proteins. The motor domain, which represents the C-terminal 2/3 of the dynein heavy chain, exhibits homology to the AAA family of ATPases. It consists of a ring of six related but divergent AAA+ units, with two substantial sized protruding projections, the stem, or tail, which anchors the protein to diverse subcellular sites, and the stalk, which binds microtubules. This article reviews recent efforts to probe the mechanism by which the dyneins produce force, and work from the authors' lab regarding long-range conformational regulation of dynein enzymatic activity.

Amino Acid Motifs↗
PubMed2006

Synthesis and biological evaluation of purealin and analogues as cytoplasmic dynein heavy chain inhibitors.

Cytoplasmic dynein plays important roles in membrane transport, mitosis, and other cellular processes. A few small-molecule inhibitors of cytoplasmic dynein have been identified. We report here the first synthesis of purealin, a natural product isolated from the sea sponge Psammaplysilla purea, which is known to inhibit axonemal dynein. Also described are the first syntheses, by modular amide coupling reactions, of the natural product purealidin A (a component of purealin) and a small library of analogues. The library was examined for inhibition of cytoplasmic dynein heavy chain and cell growth. The compounds showed effective antiproliferative activity against a mouse leukemia cell line but selective activities against human carcinoma cell lines. Purealin and some of the analogues inhibited the microtubule-stimulated ATPase activity of recombinant cytoplasmic dynein heavy chain motor domain. The inhibitory effect of purealin was concentration dependent and uncompetitive, supporting the hypothesis that it does not compete with the binding of ATP.

Adenosine Triphosphatases↗
PubMed2006

Role of the kinetochore/cell cycle checkpoint protein ZW10 in interphase cytoplasmic dynein function.

Zeste white 10 (ZW10) is a mitotic checkpoint protein and the anchor for cytoplasmic dynein at mitotic kinetochores, though it is expressed throughout the cell cycle. We find that ZW10 localizes to pericentriolar membranous structures during interphase and cosediments with Golgi membranes. Dominant-negative ZW10, anti-ZW10 antibody, and ZW10 RNA interference (RNAi) caused Golgi dispersal. ZW10 RNAi also dispersed endosomes and lysosomes. Live imaging of Golgi, endosomal, and lysosomal markers after reduced ZW10 expression showed a specific decrease in the frequency of minus end-directed movements. Golgi membrane-associated dynein was markedly decreased, suggesting a role for ZW10 in dynein cargo binding during interphase. We also find ZW10 enriched at the leading edge of migrating fibroblasts, suggesting that ZW10 serves as a general regulator of dynein function throughout the cell cycle.

Animals↗
PubMed2005

Cytoplasmic dynein nomenclature.

A variety of names has been used in the literature for the subunits of cytoplasmic dynein complexes. Thus, there is a strong need for a more definitive consensus statement on nomenclature. This is especially important for mammalian cytoplasmic dyneins, many subunits of which are encoded by multiple genes. We propose names for the mammalian cytoplasmic dynein subunit genes and proteins that reflect the phylogenetic relationships of the genes and the published studies clarifying the functions of the polypeptides. This nomenclature recognizes the two distinct cytoplasmic dynein complexes and has the flexibility to accommodate the discovery of new subunits and isoforms.

Animals↗
PubMed2005

LIS1 RNA interference blocks neural stem cell division, morphogenesis, and motility at multiple stages.

Mutations in the human LIS1 gene cause the smooth brain disease classical lissencephaly. To understand the underlying mechanisms, we conducted in situ live cell imaging analysis of LIS1 function throughout the entire radial migration pathway. In utero electroporation of LIS1 small interference RNA and short hairpin dominant negative LIS1 and dynactin cDNAs caused a dramatic accumulation of multipolar progenitor cells within the subventricular zone of embryonic rat brains. This effect resulted from a complete failure in progression from the multipolar to the migratory bipolar state, as revealed by time-lapse analysis of brain slices. Surprisingly, interkinetic nuclear oscillations in the radial glial progenitors were also abolished, as were cell divisions at the ventricular surface. Those few bipolar cells that reached the intermediate zone also exhibited a complete block in somal translocation, although, remarkably, process extension persisted. Finally, axonal growth also ceased. These results identify multiple distinct and novel roles for LIS1 in nucleokinesis and process dynamics and suggest that nuclear position controls neural progenitor cell division.

Animals↗
PubMed2005

Crystal structure of the GTPase domain of rat dynamin 1.

Here, we present the 1.9-A crystal structure of the nucleotide-free GTPase domain of dynamin 1 from Rattus norvegicus. The structure corresponds to an extended form of the canonical GTPase fold observed in Ras proteins. Both nucleotide-binding switch motifs are well resolved, adopting conformations that closely resemble a GTP-bound state not previously observed for nucleotide-free GTPases. Two highly conserved arginines, Arg-66 and Arg-67, greatly restrict the mobility of switch I and are ideally positioned to relay information about the nucleotide state to other parts of the protein. Our results support a model in which switch I residue Arg-59 gates GTP binding in an assembly-dependent manner and the GTPase effector domain functions as an assembly-dependent GTPase activating protein in the fashion of RGS-type GAPs.

Animals↗
PubMed2005

Long range allosteric control of cytoplasmic dynein ATPase activity by the stalk and C-terminal domains.

The dynein motor domain consists of a ring of six AAA domains with a protruding microtubule-binding stalk and a C-terminal domain of unknown function. To understand how conformational information is communicated within this complex structure, we produced a series of recombinant and proteolytic rat motor domain fragments, which we analyzed enzymatically. A recombinant 210-kDa half-motor domain fragment surprisingly exhibited a 6-fold higher steady state ATPase activity than a 380-kDa complete motor domain fragment. The increased ATPase activity was associated with a complete loss of sensitivity to inhibition by vanadate and an approximately 100-fold increase in the rate of ADP release. The time course of product release was discovered to be biphasic, and each phase was stimulated approximately 1000-fold by microtubule binding to the 380-kDa motor domain. Both the half-motor and full motor domain fragments were remarkably resistant to tryptic proteolysis, exhibiting either two or three major cleavage sites. Cleavage near the C terminus of the 380-kDa motor domain released a 32-kDa fragment and abolished sensitivity to vanadate. Cleavage at this site was insensitive to ATP or 5'-adenylyl-beta,gamma-imidodiphosphate but was blocked by ADP-AlF3 or ADP-vanadate. Based on these data, we proposed a model for long range allosteric control of product release at AAA1 and AAA3 through the microtubule-binding stalk and the C-terminal domain, the latter of which may interact with AAA1 to close the motor domain ring in a cross-bridge cycle-dependent manner.

Adenosine Diphosphate↗
PubMed2005

How dynein helps the cell find its center: a servomechanical model.

Cytoplasmic dynein is the major minus-end-directed microtubule motor protein in interphase cells. In addition to its well-established roles in vesicular transport and chromosome dynamics, cytoplasmic dynein also associates with the cell cortex. From this site, it appears to pull on the cytoplasmic microtubule network, influencing mitotic spindle orientation, nuclear position and other aspects of cell polarity and organization. Recent evidence indicates that the cell has the remarkable ability to calculate is geometric center, and, with the help of dynein, to position the centrosome at this central site. Here, we outline models to account for this behavior.

Animals↗
PubMed2004

Dynein: An ancient motor protein involved in multiple modes of transport.

Cytoplasmic dynein has long been thought to be responsible for retrograde axonal transport. As the number of cellular roles for this multifunctional protein has expanded, the complexity of its contribution to axonal transport has increased. In this article the increasing evidence for a role for cytoplasmic dynein in anterograde as well as retrograde transport is discussed. The current status of the complex dynein cargo-binding mechanism is evaluated. Finally, recent genetic evidence supporting a role in axonal transport and revealing a role in neurodegenerative conditions is reviewed.

Animals↗
PubMed2003

A role for cytoplasmic dynein and LIS1 in directed cell movement.

Cytoplasmic dynein has been implicated in numerous aspects of intracellular movement. We recently found dynein inhibitors to interfere with the reorientation of the microtubule cytoskeleton during healing of wounded NIH3T3 cell monolayers. We now find that dynein and its regulators dynactin and LIS1 localize to the leading cell cortex during this process. In the presence of serum, bright diffuse staining was observed in regions of active ruffling. This pattern was abolished by cytochalasin D, and was not observed in cells treated with lysophosphatidic acid, conditions which allow microtubule reorientation but not forward cell movement. Under the same conditions, using total internal reflection fluorescence microscopy, clear punctate dynein/dynactin containing structures were observed along the sides and at the tips of microtubules at the leading edge. Overexpression of dominant negative dynactin and LIS1 cDNAs or injection of antidynein antibody interfered with the rate of cell migration. Together, these results implicate a leading edge cortical pool of dynein in both early and persistent steps in directed cell movement.

1-Alkyl-2-acetylglycerophosphocholine Esterase↗
PubMed2003

Differential regulation of dynein-driven melanosome movement.

Cytoplasmic dyneins are multisubunit minus-end-directed microtubule motors. Different isoforms of dynein are thought to provide a means for independent movement of different organelles. We investigated the differential regulation of dynein-driven transport of pigment organelles (melanosomes) in Xenopus melanophores. Aggregation of melanosomes to the cell center does not change the localization of mitochondria, nor does dispersion of melanosomes cause a change in the perinuclear localization of the Golgi complex, indicating that melanosomes bear a dedicated form of dynein. We examined the subcellular fractionation behavior of dynein light intermediate chains (LIC) and identified at least three forms immunologically, only one of which fractionated with melanosomes. Melanosome aggregation was specifically blocked after injection of an antibody recognizing this LIC. Our data indicate that melanosome-associated dynein is regulated independently of bulk cytoplasmic dynein and involves a subfraction of dynein with a distinct subunit composition.

Animals↗
PubMed2002

Molecular structure of cytoplasmic dynein 2 and its distribution in neuronal and ciliated cells.

Cytoplasmic dynein is involved in a wide variety of cellular functions. In addition to the initially characterized form (MAP 1C/dynein 1), a second form of cytoplasmic dynein (dynein 2) has been identified and implicated in intraflagellar transport (IFT) in lower eukaryotes and in Golgi organization in vertebrates. In the current study, the primary structure of the full-length dynein 2 heavy chain (HC) was determined from cDNA sequence. The dynein 1 and dynein 2 sequences were similar within the motor region, and around the light intermediate chain (LIC)-binding site within the N-terminal stem region. The dynein 2 HC co-immunoprecipitated with LIC3, a homologue of dynein 1 LICs. Dynein 2 mRNA was abundant in the ependymal layer of the neural tube and in the olfactory epithelium. Antibodies to dynein 2 HC, LIC3 and a component of IFT particles strongly stained the ependymal layer lining the lateral ventricles. Both dynein 2 HC and LIC3 staining was also observed associated with connecting cilia in the retina and within primary cilia of non-neuronal cultured cells. These data support a specific role for dynein 2 in the generation and maintenance of cilia.

Animals↗
PubMed2002

A requirement for cytoplasmic dynein and dynactin in intermediate filament network assembly and organization.

We present evidence that vimentin intermediate filament (IF) motility in vivo is associated with cytoplasmic dynein. Immunofluorescence reveals that subunits of dynein and dynactin are associated with all structural forms of vimentin in baby hamster kidney-21 cells. This relationship is also supported by the presence of numerous components of dynein and dynactin in IF-enriched cytoskeletal preparations. Overexpression of dynamitin biases IF motility toward the cell surface, leading to a perinuclear clearance of IFs and their redistribution to the cell surface. IF-enriched cytoskeletal preparations from dynamitin-overexpressing cells contain decreased amounts of dynein, actin-related protein-1, and p150Glued relative to controls. In contrast, the amount of dynamitin is unaltered in these preparations, indicating that it is involved in linking vimentin cargo to dynactin. The results demonstrate that dynein and dynactin are required for the normal organization of vimentin IF networks in vivo. These results together with those of previous studies also suggest that a balance among the microtubule (MT) minus and plus end-directed motors, cytoplasmic dynein, and kinesin are required for the assembly and maintenance of type III IF networks in interphase cells. Furthermore, these motors are to a large extent responsible for the long recognized relationships between vimentin IFs and MTs.

Animals↗