Cell-to-cell heterogeneity in cortical tension specifies curvature of contact surfaces in Caenorhabditis elegans embryos

In the two-cell stage embryos of Caenorhabditis elegans, the contact surface of the two blastomeres forms a curve that bulges from the AB blastomere to the P₁ blastomere. This curve is a consequence of the high intracellular hydrostatic pressure of AB compared with that of P₁. However, the higher pressure in AB is intriguing because AB has a larger volume than P₁. In soap bubbles, which are a widely used model of cell shape, a larger bubble has lower pressure than a smaller bubble. Here, we reveal that the higher pressure in AB is mediated by its higher cortical tension. The cell fusion experiments confirmed that the curvature of the contact surface is related to the pressure difference between the cells. Chemical and genetic interferences showed that the pressure difference is mediated by actomyosin. Fluorescence imaging indicated that non-muscle myosin is enriched in the AB cortex. The cell killing experiments provided evidence that AB but not P₁ is responsible for the pressure difference. Computer simulation clarified that the cell-to-cell heterogeneity of cortical tensions is indispensable for explaining the pressure difference. This study demonstrates that heterogeneity in surface tension results in significant deviations of cell behavior compared to simple soap bubble models, and thus must be taken into consideration in understanding cell shape within embryos.     

A semianalytical model to study the effect of cortical tension on cell rolling

Cell rolling on the vascular endothelium plays an important role in trafficking of leukocytes, stem cells, and cancer cells. We describe a semianalytical model of cell rolling that focuses on the microvillus as the unit of cell-substrate interaction and integrates microvillus mechanics, receptor clustering, force-dependent receptor-ligand kinetics, and cortical tension that enables incorporation of cell body deformation. Using parameters obtained from independent experiments, the model showed excellent agreement with experimental studies of neutrophil rolling on P-selectin and predicted different regimes of cell rolling, including spreading of the cells on the substrate under high shear. The cortical tension affected the cell-surface contact area and influenced the rolling velocity, and modulated the dependence of rolling velocity on microvillus stiffness. Moreover, at the same shear stress, microvilli of cells with higher cortical tension carried a greater load compared to those with lower cortical tension. We also used the model to obtain a scaling dependence of the contact radius and cell rolling velocity under different conditions of shear stress, cortical tension, and ligand density. This model advances theoretical understanding of cell rolling by incorporating cortical tension and microvillus extension into a versatile, semianalytical framework.     

Apparent viscosity and cortical tension of blood granulocytes determined by micropipet aspiration.

Continuous deformation and entry flow of single blood granulocytes into small caliber micropipets at various suction pressures have been studied to determine an apparent viscosity for the cell contents and to estimate the extent that dissipation in a cortical layer adjacent to the cell surface contributes to the total viscous flow resistance. Experiments were carried out with a wide range of pipet sizes (2.0-7.5 microns) and suction pressures (10(2)-10(4) dyn/cm2) to examine the details of the entry flow. The results show that the outer cortex of the cell maintains a small persistent tension of approximately 0.035 dyn/cm. The tension creates a threshold pressure below which the cell will not enter the pipet. The superficial plasma membrane of these cells appears to establish an upper limit to surface dilation which is reached after microscopic "ruffles" and "folds" have been pulled smooth. With aspiration of cells by small pipets (less than 2.7 microns), the limit to surface expansion was derived from the maximal extension of the cell into the pipet; final areas were measured to be 2.1 to 2.2 times the area of the initial spherical shape. For suctions in excess of a threshold, the response to constant pressure was continuous flow in proportion to excess pressure above the threshold with only a small nonlinearity over time until the cell completely entered the pipet (for pipet calibers greater than 2.7 microns). With a theoretical model introduced in a companion paper, (Yeung, A., and E. Evans., 1989, Biophys. J. 56:139-149) the entry flow response versus pipet size and suction pressure was analyzed to estimate the apparent viscosity of the cell interior and the ratio of cortical flow resistance to flow resistance from the cell interior. The apparent viscosity was found to depend strongly on temperature with values on the order of 2 x 10(3) poise at 23 degrees C, lower values of 1 x 10(3) poise at 37 degrees C, but extremely large values in excess of 10(4) poise below 10 degrees C. Because of scatter in cell response, it was not possible to accurately establish the characteristic ratio for flow resistance in the cortex to that inside the cell; however, the data showed that the cortex does not contribute significantly to the total flow resistance.     

Estimating intercellular surface tension by laser-induced cell fusion

Intercellular surface tension is a key variable in understanding cellular mechanics. However, conventional methods are not well suited for measuring the absolute magnitude of intercellular surface tension because these methods require determination of the effective viscosity of the whole cell, a quantity that is difficult to measure. In this study, we present a novel method for estimating the intercellular surface tension at single-cell resolution. This method exploits the cytoplasmic flow that accompanies laser-induced cell fusion when the pressure difference between cells is large. Because the cytoplasmic viscosity can be measured using well-established technology, this method can be used to estimate the absolute magnitudes of tension. We applied this method to two-cell-stage embryos of the nematode Caenorhabditis elegans and estimated the intercellular surface tension to be in the 30-90 μN m(-1) range. Our estimate was in close agreement with cell-medium surface tensions measured at single-cell resolution.     

Role of cortical tension in fibroblast shape and movement

In order to analyze the cellular mechanisms of shape formation, the shape of individual 3T3 cells was perturbed by micromanipulation resulting in the detachment and relaxation of a cellular extension and the bending of the extension to form an "elbow" at a variable angle beta. Finally, the tip of the extension was allowed to reattach to the substrate away from the cell. The cells reacted by drawing the extension tight. If beta less than 90 degrees, the elbow moved laterally for 8-15 min until the extension projected orthogonally at the cell surface. If beta greater than or equal to 90 degrees, the extension remained stationary. Finally, in all cases webs formed between attachment points in the perturbed area. If the tip of the extension was allowed to touch its own cell body, thus forming a loop, the cells invariably closed the loop. The paper interprets the cellular reaction as the result of cortical tension and suggests that it is a major factor in the formation of fibroblast shape and the expressions of fibroblast motility.     

Absence of increasing cortical fMRI activity volume in response to increasing visceral stimulation in IBS patients

Cerebral cortical activity associated with perceived visceral sensation represents registration of afferent transduction and cognitive processes related to perception. Abnormalities of gut sensory function can involve either or both of these processes. Cortical registration of subliminal viscerosensory signals represents cerebral cortical activity induced by stimulation of intestinal sensory neurocircuitry without the influence of perception-related cortical activity, whereas those associated with perception represent both neural circuitry and cognitive processes. Our aims were to determine and compare quantitatively cerebral cortical functional magnetic resonance imaging (fMRI) activity in response to subliminal, liminal, and nonpainful supraliminal rectal distension between a group of irritable bowel syndrome (IBS) patients and age/gender-matched controls. Eight female IBS patients and eight age-matched healthy female control subjects were studied using brain fMRI techniques. Three barostat-controlled distension levels were tested: 1) 10 mmHg below perception (subliminal), 2) at perception (liminal), and 3) 10 mmHg above perception (supraliminal). In control subjects, there was a direct relationship between stimulus intensity and cortical activity volumes, ie., the volume of fMRI cortical activity in response to subliminal (3,226 +/- 335 microl), liminal (5,751 +/- 396 microl), and supraliminal nonpainful stimulation (8,246 +/- 624 microl) were significantly different (P < 0.05). In contrast, in IBS patients this relationship was absent and fMRI activity volumes for subliminal (2,985 +/- 332 microl), liminal (2,457 +/- 342 microl), and supraliminal nonpainful stimulation (2,493 +/- 351 microl) were similar. Additional recruitment of cortical fMRI activity volume in response to increasing stimulation from subliminal to liminal and supraliminal domains is absent in IBS patients, suggesting a difference in the processing of perceived stimulation compared with controls.     

IGF-1 induces human myotube hypertrophy by increasing cell recruitment

Insulin-like growth factor-1 (IGF-1) has been shown in rodents (i) in vivo to induce muscle fiber hypertrophy and to prevent muscle mass decline with age and (ii) in vitro to enhance the proliferative life span of myoblasts and to induce myotube hypertrophy. In this study, performed on human primary cultures, we have shown that IGF-1 has very little effect on the proliferative life span of human myoblasts but does delay replicative senescence. IGF-1 also induces hypertrophy of human myotubes in vitro, as characterized by an increase in the mean number of nuclei per myotube, an increase in the fusion index, and an increase in myosin heavy chain (MyHC) content. In addition, muscle hypertrophy can be triggered in the absence of proliferation by recruiting more mononucleated cells. We propose that IGF-1-induced hypertrophy can involve the recruitment of reserve cells in human skeletal muscle.     

EDD induces cell cycle arrest by increasing p53 levels

Tight regulation of p53 is essential for its central role in maintaining genome stability and tumor prevention. Here, EDD/ UBR5/hHyd, hereafter called EDD, is identified as a novel regulator of p53. Downregulation of EDD results in elevated p53 protein levels both in transformed and untransformed cells. Concomitant with a rise in p53, the levels of p21, a critical p53 target, are also elevated in these conditions. Surprisingly, EDD knockdown does not affect p53 protein stability, and p53 mRNA levels do not increase significantly upon EDD depletion. Consistent with the function of p53, EDD downregulation triggers a senescent phenotype in fibroblasts at later time points. In addition, the increased p53 levels upon EDD depletion cause a G₁ arrest, as co-depletion of EDD and p53 completely rescues this effect on cell cycle progression.     

Modulation of membrane dynamics and cell motility by membrane tension

The plasma membrane of most cells is drawn tightly over the cytoskeleton of the cell, resulting in a significant tension being developed in the membrane. The tension in the membrane can be calculated from the force required to separate it from the cytoskeleton; and the force itself can be measured rapidly by using laser tweezers. Recent observations indicate that decreasing membrane tension stimulates endocytosis and increasing tension stimulates secretion. Thus, membrane tension provides a simple physical mechanism to control the area of the plasma membrane. Here, we speculate that tension is a global parameter that the cell uses to control physically plasma membrane dynamics, cell shape and cell motility.     

Redistribution of cell surface receptors induced by cell-cell contact

Cell surface lectin receptors underwent rapid redistribution after embryonic Xenopus myotomal muscle cells were manipulated into contact in culture. Soybean agglutinin (SBA) receptors became highly concentrated at the contact area and concanavalin A (Con A) and ricin receptors were depleted at the same region. The accumulation of SBA receptors was greatly reduced by the presence of SBA specific sugars in the incubating medium, by precontact binding of SBA to the surface and by lowering the temperature, but it was unaffected by prolonged treatments with metabolic inhibitors. It is culture-age dependent: older cultures showed a markedly reduced extent of accumulation, and the high accumulation resulting from contact made in younger cultures disappeared with time in culture. These findings are consistent with the notion that specific molecular interaction between the contacting surfaces results in a redistribution of preexisting rapidly diffusing surface receptors. In support of this notion, ligand-free SBA and Con A receptors were shown to be laterally mobile in the membrane, and at least a subpopulation of the SBA receptors contains physically distinct molecules from the Con A receptors. We suggest that such contact- induced redistribution of various surface components may play a role in the interaction between embryonic cells.     

Myosin I contributes to the generation of resting cortical tension.

The amoeboid myosin I's are required for cellular cortical functions such as pseudopod formation and macropinocytosis, as demonstrated by the finding that Dictyostelium cells overexpressing or lacking one or more of these actin-based motors are defective in these processes. Defects in these processes are concomitant with changes in the actin-filled cortex of various Dictyostelium myosin I mutants. Given that the amoeboid myosin I's possess both actin- and membrane-binding domains, the mutant phenotypes could be due to alterations in the generation and/or regulation of cell cortical tension. This has been directly tested by analyzing mutant Dictyostelium that either lacks or overexpresses various myosin I's, using micropipette aspiration techniques. Dictyostelium cells lacking only one myosin I have normal levels of cortical tension. However, myosin I double mutants have significantly reduced (50%) cortical tension, and those that mildly overexpress an amoeboid myosin I exhibit increased cortical tension. Treatment of either type of mutant with the lectin concanavalin A (ConA) that cross-links surface receptors results in significant increases in cortical tension, suggesting that the contractile activity of these myosin I's is not controlled by this stimulus. These results demonstrate that myosin I's work cooperatively to contribute substantially to the generation of resting cortical tension that is required for efficient cell migration and macropinocytosis.     

Akt maintains cell size and survival by increasing mTOR-dependent nutrient uptake

In multicellular organisms, constituent cells depend on extracellular signals for growth, proliferation, and survival. When cells are withdrawn from growth factors, they undergo apoptosis. Expression of constitutively active forms of the serine/threonine kinase Akt/PKB can prevent apoptosis upon growth factor withdrawal. Akt-mediated survival depends in part on the maintenance of glucose metabolism, suggesting that reduced glucose utilization contributes to growth factor withdrawal-induced death. However, it is unclear how restricting access to extracellular glucose alone would lead to the metabolic collapse observed after growth factor withdrawal. We report herein that growth factor withdrawal results in the loss of surface transporters for not only glucose but also amino acids, low-density lipoprotein, and iron. This coordinated decline in transporters and receptors for extracellular molecules creates a catabolic state characterized by atrophy and a decline in the mitochondrial membrane potential. Activated forms of Akt maintained these transporters on the cell surface in the absence of growth factor through an mTOR-dependent mechanism. The mTOR inhibitor rapamycin diminished Akt-mediated increases in cell size, mitochondrial membrane potential, and cell survival. These results suggest that growth factors control cellular growth and survival by regulating cellular access to extracellular nutrients in part by modulating the activity of Akt and mTOR.     

Coupling between line tension and domain contact angle in heterogeneous membranes

The compositional differences between domains in phase-separated membranes are associated with differences in bilayer thickness and moduli. The resulting packing deformation at the phase boundary gives rise to a line tension, the one dimensional equivalent of surface tension. In this paper we calculate the line tension between a large membrane domain and a continuous phase as a function of the thickness mismatch and the contact angle between the phases. We find that the packing-induced line tension is sensitive to the contact angle, reaching a minimum at a specific value. The difference in the line tension between a flat domain (that is within the plane of the continuous phase) and a domain at the optimal contact angle may be of order 40%. This could explain why previous calculations of the thickness mismatch based line tension tend to yield values that are higher than those measured experimentally.     

The impact of line tension on the contact angle of nanodroplets

The line tension for a Lennard–Jones (LJ) fluid on a (9, 3) solid of varying strength was calculated using Monte Carlo simulations. A new perturbation method was used to determine the interfacial tension between liquid–vapour, solid–liquid and solid–vapour phases for this system to determine the Young's equation contact angle. Cylindrical and spherical nanodroplets were simulated for comparison. The contact angles from the cylindrical drops and Young's equation agree very well over the range of surface strengths and cylindrical drop sizes, except on a very weak surface. Tolman length effects were not observable for cylindrical drops. This shows that quite small systems can reproduce macroscopic contact angles. For spherical droplets, a deviation between the contact angle of spherical droplets and Young's equation was evident, but decreased with increasing interaction strengths to be negligible for contact angles less than 90°. Linear fitting of the contact angle data for varying droplet sizes showed no clear effect by line tension on contact angle. All calculated line tension values have a magnitude less than 4 × 10^? 12 J/m with both negative and positive signs. The best estimate of line tension for this system of LJ droplets was 1 × 10^? 13 J/m, which is smaller than the reported estimations in the literature, and is too small to be conclusively positive or negative in value.     

Coupling between line tension and domain contact angle in heterogeneous membranes

The compositional differences between domains in phase-separated membranes are associated with differences in bilayer thickness and moduli. The resulting packing deformation at the phase boundary gives rise to a line tension, the one dimensional equivalent of surface tension. In this paper we calculate the line tension between a large membrane domain and a continuous phase as a function of the thickness mismatch and the contact angle between the phases. We find that the packing-induced line tension is sensitive to the contact angle, reaching a minimum at a specific value. The difference in the line tension between a flat domain (that is within the plane of the continuous phase) and a domain at the optimal contact angle may be of order 40%. This could explain why previous calculations of the thickness mismatch based line tension tend to yield values that are higher than those measured experimentally.     

Activation of HERV-K(HML-2) disrupts cortical patterning and neuronal differentiation by increasing NTRK3

1. Download : Download high-res image (176KB)
2. Download : Download full-size image