Nature Genet. Park, T. Dishevelled controls apical docking and planar polarization of basal bodies in ciliated epithelial cells. Supports a direct connection between the orientation of the basal body and planar polarity.
Ciliogenesis defects in embryos lacking inturned or fuzzy function are associated with failure of planar cell polarity and Hedgehog signaling.
Watnick, T. From cilia to cyst. Boisvieux-Ulrich, E. Determination of ciliary polarity precedes differentiation in the epithelial cells of quail oviduct. Cell 72 , 3—14 Lecuit, T.
Cell surface mechanics and the control of cell shape, tissue patterns and morphogenesis. Bahmanyar, S. Liu, W. Cellular and multicellular form and function. Drug Deliv. Nie, Z. Patterning surfaces with functional polymers. Nature Mater.
The extracellular matrix guides the orientation of the cell division axis. Anisotropy of cell adhesive microenvironment governs cell internal organization and orientation of polarity.
Shows that the orientation of cell polarity is governed by the spatial distribution of cell adhesions. Wang, N. Micropatterning tractional forces in living cells.
Cytoskeleton 52 , 97— Parker, K. Directional control of lamellipodia extension by constraining cell shape and orienting cell tractional forces. Brock, A. Geometric determinants of directional cell motility revealed using microcontact printing. Langmuir 19 , — Kodama, A. ACF7: an essential integrator of microtubule dynamics. McBeath, R. Cell shape, cytoskeletal tension, and RhoA regulate stem cell lineage commitment. Cell 6 , — Huang, S. Control of cyclin D1, p27 Kip1 , and cell cycle progression in human capillary endothelial cells by cell shape and cytoskeletal tension.
James, J. Subcellular curvature at the perimeter of micropatterned cells influences lamellipodial distribution and cell polarity. Cytoskeleton 1 Aug doi Goffin, J. Cell distribution of stress fibres in response to the geometry of the adhesive environment. Cytoskeleton 63 , — Csucs, G. Locomotion of fish epidermal keratocytes on spatially selective adhesion patterns. Cytoskeleton 64 , — Jiang, X.
Directing cell migration with asymmetric micropatterns. Pouthas, F. In migrating cells, the Golgi complex and the position of the centrosome depend on geometrical constraints of the substratum. Symmetry-breaking in mammalian cell cohort migration during tissue pattern formation: role of random-walk persistence. Cytoskeleton 61 , — Gogendeau, D. Functional diversification of centrins and cell morphological complexity.
Geimer, S. Centrin scaffold in Chlamydomonas reinhardtii revealed by immunoelectron microscopy. Cell 4 , — Bastiaens, P. Gradients in the self-organization of the mitotic spindle. Trends Cell Biol. Fuller, B. Midzone activation of Aurora B in anaphase produces an intracellular phosphorylation gradient. Neumann, F. Nuclear size control in fission yeast. Cytoplasmic inheritance of the organization of the cell cortex in Paramecium aurelia.
USA 53 , — The first demonstration of an epigenetic process by which structural memory can be observed during cell reproduction. Meyer, E. Epigenetics: paramecium as a model system.
Paris 21 , — in French. Preformed cell structure and cell heredity. Prion 2 , 1—8 Basal body-associated nucleation center for the centrin-based cortical cytoskeletal network in Paramecium. Protist , — Chen, T. Multigenerational cortical inheritance of the Rax2 protein in orienting polarity and division in yeast. Experimental and theoretical study of mitotic spindle orientation. Grill, S. The distribution of active force generators controls mitotic spindle position.
Kwon, M. Mechanisms to suppress multipolar divisions in cancer cells with extra centrosomes. Paintrand, M. Centrosome organization and centriole architecture: their sensitivity to divalent cations. Klotz, C. A protein of , daltons associated with striated rootlets in ciliated epithelia, as revealed by a monoclonal antibody. Cytoskeleton 6 , 56—67 Lemullois, M. Relationships between cytokeratin filaments and centriolar derivatives during ciliogenesis in the quail oviduct.
Cell 61 , 39—49 Yang, J. Rootletin, a novel coiled-coil protein, is a structural component of the ciliary rootlet. Microtubule minus-end anchorage at centrosomal and non-centrosomal sites: the role of ninein.
Lechler, T. Desmoplakin: an unexpected regulator of microtubule organization in the epidermis. Shows how adhesion remodelling during cell differentiation induces centrosomal protein translocation to cell—cell contacts and microtubule reorganization. Cell adhesion guides cell polarity. Paris 23 , — in French. Download references. The author would like to thank M. Piel and M. They have produced original research with creative approaches over the past years, which inspired the shaping of this review.
They also contributed significantly and generously towards the figures of this manuscript. This manuscript also benefited from discussions with P.
Bastin and J. You can also search for this author in PubMed Google Scholar. One of an intracellular dynamic array of membrane-bound organelles that have distinct components and functions.
A constant flow of membranes and proteins occurs through these organelles. A single-copy structure that is generally localized at the cell centre because of its microtubule-nucleating and -anchoring activity. The centrosome is physically associated with the nucleus and duplicates once during the cell cycle.
A centrosome can be isolated, whereas a microtubule-organizing centre MTOC is not specifically an isolatable structure. Whereas a centrosome is necessarily an MTOC, the reverse is not true. A structure that is similar to the basal body organelle. A pair of centrioles is required to form the centrosome in animal cells.
The oldest centriole in the pair can convert to the basal body of a primary cilium in many types of differentiated cells. This ancient structure is present at the apparition of the early eukaryotic cells. A cytoskeletal filament with a 6-nm diameter that consists of polymerized actin.
Microfilaments form the main component of the cellular contractile machinery. The formation of blebs. Blebs are spherical cellular protrusions that occur in many physiological situations and depend on membrane—cortex adhesion. One of a set of proteins that are associated with centrosomal structures in most eukaryotes.
Centrins belong to two ancient subfamilies of the calcium-binding, EF-hand superfamily of proteins that are defined by calmodulin. The separation of a cell into two, marked by ingression of the cleavage furrow between the two nuclei. A thin, flat extension at the cell periphery that is filled with a branching meshwork of actin filaments.
A cellular structure that links the extracellular matrix on the outside of the cell to the actin cytoskeleton inside the cell through integrin receptors. A complex of myosin and actin filaments that is responsible for a range of cellular movements in eukaryotic cells.
Myosins can translocate vesicles or other cargo on actin filaments. Reprints and Permissions. Organelle positioning and cell polarity. Nat Rev Mol Cell Biol 9, — Download citation. Issue Date : November Anyone you share the following link with will be able to read this content:. Sorry, a shareable link is not currently available for this article. Provided by the Springer Nature SharedIt content-sharing initiative.
Cell and Tissue Research Chemical Research in Chinese Universities Nature Communications Nature Advanced search. Skip to main content Thank you for visiting nature. Key Points Cell polarity can be defined as a structurally and functionally asymmetric organization, in which the non-random positioning of each organelle, the function of which contributes to cell asymmetry, is preserved and transmitted through cell division.
Abstract In spite of conspicuous differences in their polarized architecture, swimming unicellular eukaryotes and migrating cells from metazoa display a conserved hierarchical interlocking of the main cellular compartments, in which the microtubule network has a dominant role. Access through your institution. Buy or subscribe. Rent or Buy article Get time limited or full article access on ReadCube. Figure 1: Intrinsic polarizing properties of the actin and tubulin cytoskeleton.
Figure 2: The centriole or basal body organelle. Figure 3: The two main polar cell architectures in eukaryotes. Figure 4: The transmission of cell polarities. Figure 5: Adhesive control of organelle positioning and cell polarity. References 1 Lee, M. Google Scholar 66 Salisbury, J. Google Scholar Tassin, A. Google Scholar Kodama, A. Google Scholar Beisson, J. Google Scholar Download references.
Acknowledgements The author would like to thank M. Glossary Endomembrane One of an intracellular dynamic array of membrane-bound organelles that have distinct components and functions.
Centrosome A single-copy structure that is generally localized at the cell centre because of its microtubule-nucleating and -anchoring activity. Centriole A structure that is similar to the basal body organelle. Microfilament A cytoskeletal filament with a 6-nm diameter that consists of polymerized actin.
Cytotaxis An epigenetic process that confers a structural memory in cell reproduction. Blebbing The formation of blebs.
These modified molecules are transported through vesicles, which pinch off from the Golgi, towards the lysosome, cell membrane or to the exterior of the cells.
Function The Golgi complexes primarily modifies the products generated from the ER and also synthesize polysaccharides on its own. The number of Golgi bodies and the products generated depend upon the cell type. The exported products are generally secretions of proteins or glycoproteins that are part of the cell's function in the organism. The additional products are returned to the endoplasmic reticulum or undergo maturation to become lysosomes. Golgi bodies synthesize glycolipids and sphingomyelin as well as serve as a site for the synthesis of complex polysaccharides for the cell wall synthesis in plants.
An active sorting role for the Golgi apparatus has also been established for the asymmetric transport of lipid raft-associated GPI-anchored proteins and proteins modified with N- and O-glycans via microtubule motor protein interactions [80]. On the other hand, it has been proposed that the initial events of cell polarization originate at the plasma membrane, based on cell-cell and cell-substrate contacts, inducing cytoskeletal reorganization and, consequently, Golgi polarization, which functions in a stabilizing and reinforcing capacity [73].
Early plasma membrane polarization, and specifically the generation of plasma membrane potential via polarization of ion channels and pumps, has also been shown to directly modulate the actin cytoskeleton and drive other polarization events [81].
Other Golgi-independent mechanisms for achieving plasma membrane polarity have also been revealed, including self assembly and clustering [48] , actin-mediated trapping [49] , microtubule-based active transport [43] , or asymmetric membrane insertion mediated by non-classical pathways [52] , [82] — [84]. Many protein and lipid species associated with lipid rafts have been shown to follow non-classical, BFA-insensitive secretion, including cholesterol [52] , [85] , sphingomyelin [86] — [88] , and the ganglioside GD3 [89].
Interestingly, all of these lipids follow similar kinetics to those demonstrated here for GM1, taking approximately 10 min to reach the plasma membrane. The intermediated compartment IC , formed by a stable tubular structure at the interface between the ER and Golgi apparatus, is thought to play a central role in BFA-insensitive pathways by providing either a direct route to the plasma membrane or by bypassing the Golgi compartment and fusing with post-Golgi endocytic recycling compartments trafficked via microtubules to the plasma membrane.
The precise mechanisms, specific coat proteins, and membrane fusion factors remain, however, unknown [82]. The observed BFA-insensitive polarization behavior of GM1 in the plasma membrane could suggest that the biosynthetic pathway of GM1 parallels that of cholesterol and sphingomyelin. However, a puzzling factor is that the total amount of GM1 in the plasma membrane was greatly reduced upon BFA treatment, implying at least partial reliance on classical, BFA-sensitive Golgi synthesis pathways.
Another explanation may lie in recent studies describing a novel system of endocytic membrane recycling pathways termed CLICs clathrin-independent carriers [83] , [84]. CLICs are thought to represent a mechanism for bulk plasma membrane recycling that contributes to rapid changes in plasma membrane morphology. It might be the case that GM1 in the plasma membrane is sequestered into intracellular CLIC compartments until stimulation with polarization-inducing growth factors causes leading edge CLIC compartments to fuse with the plasma membrane.
This scenario would explain our observation that wortmannin treated cells had significantly reduced GM1 content in the plasma membrane Figure 5 , while providing an attractive mechanism for achieving fast polarization of GM1. The question remains as to what function the polarized Golgi apparatus serves, if membrane asymmetry can be achieved independently.
It could be that Golgi-derived vesicles carry specific proteins or lipid species essential for the regulation or modulation of plasma membrane polarity. The lack of these essential Golgi-derived proteins or lipid species might explain the aberrant clustering pattern of GM1 that we observed after BFA treatment inhibited Golgi exit. Cdc42 localization is BFA-sensitive and thought to regulate Golgi secretion pathways, but the exact role in this context is unknown [23] , [93].
Activating Cdc42 through the expression of the oncogene Dbl causes a translocation of Cdc42 from the Golgi apparatus to newly formed lamellipodia in the plasma membrane [92]. Although to our knowledge it has never been directly shown, it may be possible that the Golgi apparatus is involved in directly trafficking Cdc42, and possibly other Rho proteins, to the plasma membrane during cell polarization and that the polarized localization assists in this function.
It is also possible that the polarized Golgi apparatus contributes to the later stages of polarization maintenance and cell migration. Blocking traffic from the Golgi apparatus has been known for some time to inhibit cell migration [33] — [35].
That the Golgi apparatus plays its essential role in the later stages of cell migration rather than in the initial stages of establishing polarity is consistent with the idea that Golgi polarity acts in a stabilizing or reinforcing capacity [94] , and, in any case, does not exclude the other possible roles described above.
Finally, the Golgi apparatus has also been shown to play an important role in cell signaling, and its polarized position within the cell could contribute to this role. Two kinases involved in cell polarization and migration, YSK1 and MST4, were shown to localize to the Golgi apparatus through interactions with Golgi matrix protein GM, which directly activated the kinases [95].
Ras shows diverse activation kinetics and second messenger binding partners based on its localization to the plasma membrane or Golgi apparatus, and can even be activated independently in one location or the other [96]. Based on its role in cell signaling, it is possible that the polarized location of the Golgi apparatus contributes to the maintenance of intracellular gradients by localizing activated kinases like YSK1 and Ras towards the front of polarized cells.
The experiments indicate that the polarization events are controlled by separate biochemical pathways, yet migration is more efficient when both pathways are active. This suggests that it may be advantageous for the cell to separately control the activation of individual pathways, and by extension, individual cell polarization events.
We can speculate that the two divergent pathways controlling Golgi apparatus and GM1 polarization may function as a synergistic regulator of cell migration. Only when both pathways are activated and both systems engaged can migration proceed efficiently. The need for two separately engaged pathways may serve as a necessary regulation that constrains migration except for where strictly required.
Consequently, the interplay between these separately regulated pathways has been a prime target for cancer therapeutics. Cancerous cells have been shown to take advantage of the uncoupled nature of the pathways; where one pathway is inhibited by chemotherapeutic drugs, the other seems to compensate to drive cell proliferation and migration [99] , []. A clinical trial combining inhibition of MEK and AKT showed more effective tumor reduction and increased positive outcomes than inhibition of a single pathway alone [].
Further research into the mechanisms involved in cell polarization and migration, and the interplay between cellular pathways and systems will contribute to the development of alternative strategies to effectively treat an array of diseases, including metastases in cancer. Paola Defilippi []. Chiara Lanzuolo. For scratch experiments, ECV cells were plated on 18 mm No.
A wound edge was created with a sterilized razor blade by removing the cells from one half of the cover slip. Stimulation and drug treatments were performed on unlabeled live ECV cells to avoid aggregation effects of cholera toxin subunit B, and to avoid any possibility of affecting plasma membrane diffusion rates of labeled molecules. We implemented a protocol of two separate rounds of fixation and staining that was used for all experiments to prevent non-specific binding of QDots.
First, cover slips were fixed with 1. Washing was performed between each step in PBS. Excitation light was provided by a mercury lamp HgW. Fluorescence signal was detected by a Hamamatsu Photonics K. The Argus Image Processor was set to average 8 frames of the standard 30 frames per second camera output in order to improve signal-to-noise and reduce the visible blinking effects of QDots.
Digital images were captured using a custom-made Virtual Instrument produced with Labview 7. Images were collected along the scratch surface. Prior to capturing digital images of the wound edge cells, we carefully aligned the coverslip so that the wound was parallel to the x -axis of both the microscope stage and the camera's ccd by scanning back and forth and adjusting the slide manually.
In this way, our images always had the wound in the upper quadrant, and the positive y -axis facing the wound was used as a reference to calculate polarization. Three QD images representing different focal layers of the plasma membrane were transformed into a maximum intensity projection that was combined with the Golgi apparatus and Hoechst images into an RGB composite. Image processing was performed using the program ImageJ64 v1.
Background fluorescence was subtracted using the Brightness and Contrast function in ImageJ by increasing the minimum threshold until the background read as 0. Using center of mass measurements requires that the background of an image or ROI be reduced to 0 because even low levels of background noise do not allow an accurate calculation of the center of mass. Care was taken not to over aggressively reduce the background by assuring that signals from single QDs remained in the final image.
Representative images were cropped and levels adjusted uniformly in Photoshop CS4 during the compilation of figures. GM1 polarization was determined by comparing the weighted center of mass from QD fluorescence distribution in the plasma membrane with the center of mass calculated for the area of the cell.
All pixels within the ROI were then set to 1, and the geometric center of mass of the cell C m cell was measured. The Golgi angle was calculated from the vector coordinates as follows:. The second factor in the equation enables measurement of the defined angle on all four quadrants of the coordinate system we adapted, breaking the symmetry of the arctan function.
Background noise was subtracted as described above prior to center of mass measurements. Background was subtracted using a rolling radius of 25 pixels, and images were thresholded below a value of 29, which was determined empirically to retain a signal for single QDots in the majority of images. Particle analysis was run on previously determined single cell ROIs with no constraints on particle size or circularity, and the number, average size, area fraction, and total area of the cell were returned as raw data.
Labeling density is equivalent to the area fraction, or the percent of the total cell area covered by fluorescent signal after thresholding. For two-dimensional cell migration assays, ECV cells were grown as described above were plated in 6 well plates on glass cover slips.
Images were collected of the 0 h time point using a 10 X objective and inverted microscope. After 24 h and again after 48 h, cells were removed from the incubator and imaged using the same 10 X objective and inverted microscope. Care was taken to align the scratch along the y axis of the camera to aid subsequent image quantification. Matrigel invasion assays were performed in 24 well plates using 0. Finally, cells were dispersed with TE solution 0.
Immediately after adding the cells, inhibitory drugs were added to the top chamber as well. Finally, a cotton swab was used to remove cells from the surface of the 0. After a brief drying period, the whole insert was placed on a cover slide and imaged with a 10x objective on an inverted microscope.
Experiments were repeated a minimum of three times. N values reported in the figure legends represent the total number of cells analyzed over three or more experiments. For the wound healing assay, the area of the scratch was measured at various points along the scratch during at least three independent experiments.
The results were pooled for each experimental condition, and two-way ANOVA and Bonferroni post-tests comparing all columns were applied to assess significance. Similarly, for matrigel invasion assays, migrated cells were counted for several frames per experiment, and the results for at least three independent experiments were pooled and subjected to analysis by one way ANOVA with Tukey post-tests to compare all experimental conditions. N values represent the total number of measurements, either scratch area or number of cells per frame.
Comparison of classical and semi-automated methods for measuring Golgi apparatus polarization. B 30 min stimulation with either serum, EGF, or LPA, leads to significant Golgi polarization levels compared to control, and no significant differences were present between the two evaluation methods.
We thank Drs. We thank Dr. Performed the experiments: BB. Wrote the paper: BB. Browse Subject Areas? Click through the PLOS taxonomy to find articles in your field. Abstract Cell polarization is a process of coordinated cellular rearrangements that prepare the cell for migration. Introduction Cell polarization and cell migration are interrelated, highly coordinated processes that allow complex, stratified tissue morphology and guided navigation in response to chemical cues [1] — [4].
Results Quantifying Golgi apparatus and GM1 polarization An accurate quantification of polarization is essential to investigate the relationship between the Golgi apparatus and GM1 in the plasma membrane, and allows us to determine the individual contributions of each structure, define the effects of various drugs, and assess correlation between the two events. Download: PPT. Figure 1. Quantification of Golgi apparatus and GM1 polarization.
Figure 2. Golgi apparatus and GM1 polarization depend on separate intracellular pathways To confirm the assertion that Golgi polarization is not required for GM1 polarization, we decided to utilize a drug known to block Golgi apparatus polarization.
Figure 3. Drug treatments reveal uncoupled pathways to Golgi apparatus and GM1 polarization. Simultaneous measurements of Golgi apparatus and GM1 polarization in single cells Next, we tested whether any correlation could be determined between Golgi and GM1 polarization at the level of the individual cell.
Figure 4. Analysis of GM1 distribution among experimental populations While assessing membrane polarization, we observed that the pattern of GM1 staining was highly variable not only between different experimental conditions, but also between cells of the same population Figure 5.
Figure 5. Analysis of GM1 distribution in response to drug treatments. Figure 6. Cell migration wound closure assay after treatment with drugs. Discussion Correlation of polarization events in single cells We have described new methodologies for quantifying Golgi apparatus and GM1 distributions in polarizing cells.
Fast, Golgi-independent membrane polarization We have shown that polarization of GM1 in the plasma membrane occurs before Golgi apparatus polarization and is not affected by either the position or functional state of the Golgi. Role of the polarized Golgi apparatus The question remains as to what function the polarized Golgi apparatus serves, if membrane asymmetry can be achieved independently.
Biochemical and mechanistic separation in polarization response We have demonstrated that the uncoupled polarization responses of the Golgi apparatus and GM1 in the plasma membrane depend on MEK and PI3K activation, respectively. Fixation and labeling We implemented a protocol of two separate rounds of fixation and staining that was used for all experiments to prevent non-specific binding of QDots.
Quantification of GM1 and Golgi apparatus polarization GM1 polarization was determined by comparing the weighted center of mass from QD fluorescence distribution in the plasma membrane with the center of mass calculated for the area of the cell.
Cell migration and invasion assays For two-dimensional cell migration assays, ECV cells were grown as described above were plated in 6 well plates on glass cover slips.
Statistical analysis Experiments were repeated a minimum of three times. Supporting Information. Figure S1. Figure S2. File S1. Acknowledgments We thank Drs. References 1. Nabi I The polarization of the motile cell. J Cell Sci Pt 12 : — View Article Google Scholar 2. Condliffe A, Hawkins P Cell biology.
Moving in mysterious ways. Nature , Nature — View Article Google Scholar 4. Science — View Article Google Scholar 5. Annu Rev Immunol — View Article Google Scholar 6. Immunity 4: — View Article Google Scholar 7. Semin Cell Dev Biol —
0コメント