Dynein is required for receptor sorting and the morphogenesis of early endosomes

Driskell, Owen J; Mironov, Aleksandr; Allan, Viki; Woodman, Philip

doi:10.1038/ncb1525

Cited by 166 publications

(177 citation statements)

References 35 publications

Supporting

Mentioning

158

Contrasting

Order By: Relevance

“…For the control versus treatment condition, we estimated the normalizedT 2D ¼ 4.160 andT 3D ¼ 6.670 giving rise to P 2D -value of 1.589 Ã 10 −5 and P 3D -value ¼ 1.280 Ã 10 −11 , indicating that there is strong evidence that the drug treatment significantly affects the distribution of MVB. These results agree with previous studies demonstrating microtubule-dependent movement of MVB (35). To further evaluate our approach, we performed additional analysis of diverse subcellular structures (SI Text, Fig.…”

Section: Detection Of Morphological Changes In Micropatterned Cells Asupporting

confidence: 79%

“…2A), we analyzed the morphology changes of multivesicular bodies (MVB) induced by a treatment with the drug nocodazole (NZ) that depolymerizes microtubules, a major component of the cellular cytoskeleton. MVB are endosomes involved in several important cellular functions, including processing of nutrients, ligands and receptors during endocytosis, exosome secretion, and autophagy (34) that are transported along microtubules (35). Intracellular MVB were visualized by indirect immunofluorescence against CD63, a transmembrane protein enriched in MVB (Fig.…”

Section: Detection Of Morphological Changes In Micropatterned Cells Amentioning

confidence: 99%

See 1 more Smart Citation

Closed-form density-based framework for automatic detection of cellular morphology changes

Duong

Goud

Schauer

2012

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

A primary method for studying cellular function is to examine cell morphology after a given manipulation. Fluorescent markers attached to proteins/intracellular structures of interest in conjunction with 3D fluorescent microscopy are frequently exploited for functional analysis. Despite the central role of morphology comparisons in cell biological approaches, few statistical tools are available that allow biological scientists without a high level of statistical training to quantify the similarity or difference of fluorescent images containing multifactorial information. We transform intracellular structures into kernels and develop a multivariate two-sample test that is nonparametric and asymptotically normal to directly and quantitatively compare cellular morphologies. The asymptotic normality bypasses the computationally intensive calculations used by the usual resampling techniques to compute the P-value. Because all parameters required for the statistical test are estimated directly from the data, it does not require any subjective decisions. Thus, we provide a black-box method for unbiased, automated comparison of cell morphology. We validate the performance of our test statistic for finite synthetic samples and experimental data. Employing our test for the comparison of the morphology of intracellular multivesicular bodies, we detect changes in their distribution after disruption of the cellular microtubule cytoskeleton with high statistical significance in fixed samples and live cell analysis. These results demonstrate that density-based comparison of multivariate image information is a powerful tool for automated detection of cell morphology changes. Moreover, the underlying mathematics of our test statistic is a general technique, which can be applied in situations where two data samples are compared.hypothesis test | integrated density functional | optimal bandwidth selection | quantitative cell comparison F luorescent markers attached to proteins of interest in conjunction with modern fluorescent microscopy technologies are a useful proxy for studying subcellular compartments and their behavior after a given manipulation. Treatment with chemical compounds or specific gene silencing by RNA interference are commonly used at the scale of individual experiments to highthroughput studies. Visual inspection by expert biologists has been performed for several decades, ranging from early studies by microscopists like Ramon y Cajal to contemporary large scale, high-throughput screens (1-4). Although human observation may be very accurate, the three major drawbacks are that (i) it lacks quantitative measures, (ii) it may be biased, and (iii) it is time consuming.The structural features of cells and the topological relationships between the numerous intracellular compartments give rise to multivariate data whose unbiased, automatic comparison is a major challenge. Importantly, alterations in cellular morphology also occur in many diseases, including cancer, requiring quantitative tools for their detection. Give...

show abstract

Section: Detection Of Morphological Changes In Micropatterned Cells Asupporting

confidence: 79%

Section: Detection Of Morphological Changes In Micropatterned Cells Amentioning

confidence: 99%

Closed-form density-based framework for automatic detection of cellular morphology changes

Duong

Goud

Schauer

2012

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

show abstract

“…Rather, as was found in studies of early endocytic vesicles from the rat, all motility in both wild-type and Kifc2 knockout mouse vesicles was inhibited by 1 mM AMP-PNP, an inhibitor of kinesin based motility (Vale et al, 1992;Bananis et al, 2000;Murray et al, 2000). A recent study in which HeLa cells were loaded with fluorescent epidermal growth factor for 2-3 min and followed for as long as 30 min was interpreted as showing a role for dynein in processing of early endocytic vesicles (Driskell et al, 2007), in contrast to other studies (Nielsen et al, 1999;Bananis et al, 2003Bananis et al, . 2004.…”

Section: Discussionmentioning

confidence: 99%

Kif5B and Kifc1 Interact and Are Required for Motility and Fission of Early Endocytic Vesicles in Mouse Liver

Nath

Bananis

Sarkar

et al. 2007

MBoC

View full text Add to dashboard Cite

Early endocytic vesicles loaded with Texas Red asialoorosomucoid were prepared from mouse liver. These vesicles bound to microtubules in vitro, and upon ATP addition, they moved bidirectionally, frequently undergoing fission into two daughter vesicles. There was no effect of vanadate (inhibitor of dynein) on motility, whereas 5 -adenylylimido-diphosphate (kinesin inhibitor) was highly inhibitory. Studies with specific antibodies confirmed that dynein was not associated with these vesicles and that Kif5B and the minus-end kinesin Kifc1 mediated their plus-and minus-end motility, respectively. More than 90% of vesicles associated with Kifc1 also contained Kif5B, and inhibition of Kifc1 with antibody resulted in enhancement of plus-end-directed motility. There was reduced vesicle fission when either Kifc1 or Kif5B activity was inhibited by antibody, indicating that the opposing forces resulting from activity of both motors are required for fission to occur. Immunoprecipitation of native Kif5B by FLAG antibody after expression of FLAG-Kifc1 in 293T cells indicates that these two motors can interact with each other. Whether they interact directly or through a complex of potential regulatory proteins will need to be clarified in future studies. However, the present study shows that coordinated activity of these kinesins is essential for motility and processing of early endocytic vesicles. INTRODUCTIONReceptor-mediated endocytosis is a process in which ligands bind to specific cell surface receptors and internalize via clathrin-coated pits. After internalization the clathrin coat is released and uncoated vesicles mature into early endosomes (Mellman, 1996;Mukherjee et al., 1997;Marsh and McMahon, 1999;Higgins and McMahon, 2002;Perrais and Merrifield, 2005). After acidification of early endosomes, ligands such as asialoorosomucoid (ASOR) that are destined for lysosomes dissociate from their receptors (Harford et al., 1983a,b), and a series of fission events results in their segregation into separate daughter vesicles (Wolkoff et al., 1984;Mellman, 1996;Mukherjee et al., 1997;Murray and Wolkoff, 2003). The resulting ligand-enriched late endosomes traffic to lysosomes for degradation, whereas the receptor-containing daughter endosomes traffic back to the cell surface where receptor is reused (Harford et al., 1983a;Wolkoff et al., 1984;Mukherjee et al., 1997). Previous studies indicated that this segregation event requires an intact microtubule cytoskeleton (Goltz et al., 1992;Novikoff et al., 1996;Murray et al., 2000;Bananis et al., 2003Bananis et al., , 2004. In more recent studies, microtubule-based endosome motility and segregation were reconstituted in vitro using fluorescent early endosomes prepared from rat liver 5 min after portal venous injection of Texas Red-labeled ASOR, a substrate for the hepatocytespecific asialoglycoprotein receptor (ASGPR) (Bananis et al., , 2003(Bananis et al., , 2004Murray et al., 2000;Murray and Wolkoff, 2003). On addition of ATP to the in vitro assay, these vesicles moved bidirectionall...

show abstract

“…In experiments on dynein alone, is seen to move progressively, however it is a simplification of what is seen in experiments on the dynein moving cargo. Experiments do show cargos within a cell being moved in the direction of the nucleus over time, however there are also pauses in movement and the cargo can travel backwards or move in a direction which is not solely radial [21,12,42,46]. Currently, the available experimental data does not show how the dynein itself is behaving at these times, as we only see the position of the endosome.…”

Section: Variation Of Parametersmentioning

confidence: 78%

“…Microtubules are polarised, with the plus-end usually pointing towards the cell membrane, away from the nucleus [40]. There are numerous articles providing supporting evidence as to which motor proteins are involved in each stage of the process, although they do not always agree [3,5,12,23,33]. These differences may be due to the differences in results from the studies of different types of cells, experimental techniques and differences between in vitro and in vivo studies.…”

Section: Introductionmentioning

confidence: 99%

From the Cell Membrane to the Nucleus: Unearthing Transport Mechanisms for Dynein

et al. 2012

View full text Add to dashboard Cite

Mutations in the motor protein cytoplasmic dynein have been found to cause Charcot-MarieTooth disease in humans, spinal muscular atrophy and severe intellectual disabilities in humans.In mouse models neurodegeneration is observed. We sought to develop a novel model which could incorporate the effect of mutations on distance travelled and velocity. A mechanical model for the dynein mediated transport of endosomes is derived from first principles and solved numerically. The effects of variations in model parameter values are analysed to find those that have a significant impact on velocity and distance travelled. The model successfully describes the processivity of dynein and matches qualitatively the velocity profiles observed in experiments.

show abstract

Dynein is required for receptor sorting and the morphogenesis of early endosomes

Cited by 166 publications

References 35 publications

Closed-form density-based framework for automatic detection of cellular morphology changes

Closed-form density-based framework for automatic detection of cellular morphology changes

Kif5B and Kifc1 Interact and Are Required for Motility and Fission of Early Endocytic Vesicles in Mouse Liver

From the Cell Membrane to the Nucleus: Unearthing Transport Mechanisms for Dynein

Contact Info

Product

Resources

About