Dynamin: possible mechanism of "Pinchase" action.
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Dynamin is a GTPase playing an essential role in ubiquitous intra cellular processes involving separation of vesicles from plasma membranes and membranes of cellular compartments. Recent experimental progress (. Cell. 93:1021-1029;. Cell. 94:131-141) has made it possible to attempt to understand the action of dynamin in physical terms. Dynamin molecules are shown to bind to a lipid membrane, to self-assemble into a helicoidal structure constricting the membrane into a tubule, and, as a result of GTP hydrolysis, to mediate fission of this tubule (). In a similar way, dynamin is supposed to mediate fission of a neck connecting an endocytic bud and the plasma membrane, i.e., to complete endocytosis. We suggest a mechanism of this "pinchase" action of dynamin. We propose that, as a result of GTP hydrolysis, dynamin undergoes a conformational change manifested in growth of the pitch of the dynamin helix. We show that this gives rise to a dramatic change of shape of the tubular membrane constricted inside the helix, resulting in a local tightening of the tubule, which is supposed to promote its fission. We treat this model in terms of competing elasticities of the dynamin helix and the tubular membrane and discuss the predictions of the model in relation to the previous views on the mechanism of dynamin action. (+info)
In vivo human brachial artery elastic mechanics: effects of smooth muscle relaxation.
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BACKGROUND: The effects of smooth muscle relaxation on arterial wall mechanics are controversial. We used a new, in vivo, noninvasive technique to measure brachial artery wall mechanics under baseline conditions and following smooth muscle relaxation with nitroglycerin (NTG). METHODS AND RESULTS: Eight healthy, normal subjects (6 male, 2 female; age 30+/-3.1 years) participated in the study. The nondominant brachial artery was imaged through a water-filled blood pressure cuff using an external ultrasound wall-tracking system at baseline and following 0.4 mg sublingual NTG. Simultaneous radial artery pressure waveforms were recorded by tonometry. Transmural pressure (TP) was reduced by increasing water pressure in the cuff. Brachial artery area, unstressed area, compliance, stress, strain, incremental elastic modulus (Einc), and pulse wave velocity (PWV) were measured over a TP range from 0 to 100 mm Hg. Baseline area versus TP curves generated 30 minutes apart were not significantly different. NTG significantly shifted area versus TP (P<0.0001) and compliance versus TP (P<0.001) curves upward, whereas the Einc versus TP (P<0.05) and PWV versus TP (P<0. 01) curves were shifted downward. NTG also significantly shifted stress versus strain (P<0.01) and Einc versus strain (P<0.01) curves to the right. CONCLUSIONS: We conclude that brachial artery elastic mechanics can be reproducibly measured over a wide range of TP and smooth muscle tone using a new noninvasive ultrasound technique. Smooth muscle relaxation with NTG increases isobaric compliance and decreases isobaric Einc and PWV in the human brachial artery. (+info)
Myxococcus cells respond to elastic forces in their substrate.
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Elasticotaxis describes the ability of Myxococcus xanthus cells to sense and to respond to elastic forces within an agar gel on which they rest. Within 5 min of the application of stress, each cell begins to reorient its long axis perpendicular to the stress force. The cells then glide in that direction, and the swarm becomes asymmetric. A quantifiable assay for the strength of elasticotaxis is based on the change in swarm shape from circular to elliptic. By using a collection of isogenic motility mutants, it has been found that the ability to respond to stress in agar depends totally on adventurous (A) motility, but not at all on social (S) motility or on the frz genes. In fact, S- mutants (which are moving only by means of A motility) respond to the applied stress more strongly than does the wild type, despite the fact that their spreading rates are slower than that of the wt strain. Based on the swarming and elasticotactic phenotypes of isogenic frizzy strains that were also defective either in A or S motility, frz behaves as if part of the S motility system. (+info)
Cardiac consequences of prolonged exposure to an isolated increase in aortic stiffness.
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In elderly patients, aortic stiffness is a major determinant of increased end-systolic stress leading to left ventricular (LV) hypertrophy with impaired cardiac performance. However, in a rat model of aortic elastocalcinosis (induced by vitamin D(3)-nicotine [VDN] treatment), brief exposure (1 month) to increased aortic stiffness modified neither cardiac function nor cardiac structure. Here we report the impact of longer exposure (3 months) to aortic stiffness. Three months after induction of aortic stiffness, aortic characteristic impedance was measured in awake rats, 8 control and 10 VDN. Stroke volume was measured (electromagnetic probe) at baseline and after acute volume overload. LV weight/body weight ratio, collagen, and myosin heavy chain (MHC) contents were determined. Although aortic characteristic impedance increased (controls, 32+/-2; VDN rats, 50+/-8 10(3) dyne. s/cm(5); P=0.0248), stroke volume was maintained in VDN rats at baseline (controls, 223+/-18; VDN, 211+/-13 microL) and after volume overload (controls, 378+/-14; VDN, 338+/-15 microL). However, LV weight/body weight ratio (controls, 1.54+/-0.07; VDN, 1.73+/-0.05 g/kg; P=0.0397) and LV collagen content (controls, 31+/-4; VDN, 52+/-4 microgram/g dry wt; P=0.0192) increased. A shift from alpha-MHC (controls, 82+/-2%; VDN, 69+/-3%; P=0.0056) to beta-MHC (controls, 18+/-2%; VDN, 31+/-3%; P=0. 0056) was also observed. Three months' exposure to increased aortic stiffness in VDN rats induced LV hypertrophy with moderate interstitial fibrosis and a shift in the MHC-isoform pattern. Such structural adaptation maintains LV performance. (+info)
Left atrial relaxation and left ventricular systolic function determine left atrial reservoir function.
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BACKGROUND: Determinants of left atrial (LA) reservoir function and its influence on left ventricular (LV) function have not been quantified. METHODS AND RESULTS: In an open-pericardium, paced (70 and 90 bpm) pig model of LV regional ischemia (left anterior descending coronary constriction), with high-fidelity LV, LA, and RV pressure recordings, we obtained the LA area with 2D automated border detection echocardiography, LA pressure-area loops, and Doppler transmitral flow. We calculated LV tau, LA relaxation (a-x pressure difference divided by time, normalized by a pressure), and stiffness (slope between x and v pressure points of v loop). Determinants of total LA reservoir (maximum-minimum area, cm(2)) were identified by multiple regression analysis. Different mean rates of LA area increase identified 2 consecutive (early rapid and late slow) reservoir phases. During ischemia, LV long-axis shortening (LAS, LV base systolic descent) and LA reservoir area change decreased (7.3+/-0.3 [SEM] versus 5.6+/-0.3 cm(2), P<0.001) and LA stiffness increased (1.6+/-0.3 versus 3.1+/-0.3 mm Hg/cm(2), P=0.009). Early reservoir area change depended on LA mean ejection rate (LA area at ECG P wave minus minimum area divided by time; multiple regression coefficient=0.9; P<0.001) and relaxation (coefficient=4.9 cm(2)xms/s; P<0.001). Late reservoir area change depended on LAS (coefficient=8 cm/s; P<0.001). Total reservoir filling depended on LA stiffness (coefficient=-0.31 cm(4)/mm Hg; P=0. 001) and cardiac output (coefficient=0.001 cm(2)xmin/L; P=0.002). The strongest predictor of cardiac output was LA reservoir filling (coefficient=301 L/minxcm(2); P<0.001). The v loop area was determined by cardiac output, LV ejection time, tau, and early transmitral flow. CONCLUSIONS: Two (early and late) reservoir phases are determined by LA contraction and relaxation and LV base descent. Acute LV regional ischemia increases LA stiffness and impairs LA reservoir function by reducing LV base descent. (+info)
Rhodamine phalloidin F-actin: critical concentration versus tensile strength.
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The mechanic and elastic properties of rhodamine phalloidin F-actin were investigated as a function of the ionic strength and in the absence of Mg2+. By increasing ionic strength from 3 to 19 mM, critical concentration decreased from 146 to 36 nM and the yield strength increased from 5.6 pN to 28.6 pN. At the ionic strength of 12-13 mM, the elastic modulus by stretching increased by 330-430 kP. nm-1 up to the break point, where it was 38-44.2 MP. The work required to break the filament, 403-439 kJ.M-1 provides an estimate of the free energy of annealing of rhodamine phalloidin F-actin, the annealing constant being 2.8 x 1074 M-1. (+info)
TGF-beta receptor expression and binding in rat mesangial cells: modulation by glucose and cyclic mechanical strain.
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BACKGROUND: Transforming growth factor-beta (TGF-beta) is a causal factor in experimental glomerulosclerosis, and it mediates the increased extracellular matrix (ECM) accumulation that occurs in cultured mesangial cells (MCs) exposed to high glucose concentrations and cyclic mechanical strain. This change is associated with increased levels of TGF-beta, but may also involve alterations in receptor expression and binding. METHODS: Rat MCs cultured in media containing either 8 or 35 mM glucose were seeded into culture plates with elastin-coated flexible bottoms. Thereafter, they were subjected to cyclic stretch or static conditions and then examined for 125I-TGF-beta1 binding and expression of TGF-beta receptors at the gene and protein levels. RESULTS: Kinetic studies showed that MCs bound TGF-beta1 in a time- and concentration-dependent manner, expressing 6800 high-affinity receptors per cell, with an apparent dissociation constant (Kd) of 15.4 pM, while cross-linking analysis identified three TGF-beta receptors (betaR) corresponding to betaRI, betaRII, and betaRIII of 54, 73, and 200 kDa, respectively. Immunocytochemical studies of betaRI and betaRII protein revealed MC expression in a homogeneous, punctate distribution, whereas Northern analysis demonstrated the presence of the corresponding mRNAs. Exposure to cyclic stretching significantly increased (10%) the overall number of TGF-beta receptors, whereas ligands associated with betaRs I, II, and III also increased (25 to 50%). The finding of increased (30 to 40%) betaRI and betaRII transcript levels and immunoreactive protein (163 and 59%, respectively) in the absence of significant changes in the apparent Kd indicated that stretch-induced binding was the result of increased receptor synthesis and expression and not due to a change in binding affinity. In a similar, but more dramatic fashion, exposure to high glucose also elevated (50%) the receptor number, as well as the amount of ligands associated with betaRs I, II, and III (100 to 250%). This same treatment also increased the levels of betaRI and betaRII mRNA (30 to 40%) and the immunoreactive protein (82 and 82%, respectively), without significantly altering the binding affinity of the receptor. A concerted or synergistic effect of both stimuli was not evidenced. CONCLUSION: These results suggest that the modulation of TGF-beta receptors may be an additional control point in mediating the glucose- and mechanical force-induced increase in ECM deposition by MCs. (+info)
I-band titin in cardiac muscle is a three-element molecular spring and is critical for maintaining thin filament structure.
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In cardiac muscle, the giant protein titin exists in different length isoforms expressed in the molecule's I-band region. Both isoforms, termed N2-A and N2-B, comprise stretches of Ig-like modules separated by the PEVK domain. Central I-band titin also contains isoform-specific Ig-motifs and nonmodular sequences, notably a longer insertion in N2-B. We investigated the elastic behavior of the I-band isoforms by using single-myofibril mechanics, immunofluorescence microscopy, and immunoelectron microscopy of rabbit cardiac sarcomeres stained with sequence-assigned antibodies. Moreover, we overexpressed constructs from the N2-B region in chick cardiac cells to search for possible structural properties of this cardiac-specific segment. We found that cardiac titin contains three distinct elastic elements: poly-Ig regions, the PEVK domain, and the N2-B sequence insertion, which extends approximately 60 nm at high physiological stretch. Recruitment of all three elements allows cardiac titin to extend fully reversibly at physiological sarcomere lengths, without the need to unfold Ig domains. Overexpressing the entire N2-B region or its NH(2) terminus in cardiac myocytes greatly disrupted thin filament, but not thick filament structure. Our results strongly suggest that the NH(2)-terminal N2-B domains are necessary to stabilize thin filament integrity. N2-B-titin emerges as a unique region critical for both reversible extensibility and structural maintenance of cardiac myofibrils. (+info)