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PubMed1997

alpha-Tocopheryl hemisuccinate administration increases rat liver subcellular alpha-tocopherol levels and protects against carbon tetrachloride-induced hepatotoxicity.

Rats were administered a series of tocopherol analogs 18 h prior to a hepatotoxic dose of carbon tetrachloride (CCl4). Of the compounds tested, only d-alpha-tocopheryl hemisuccinate (TS) provided significant protection against CCl4-induced hepatotoxicity. No protection was observed with either d-alpha-tocopherol (alpha-T) or a tocopherol succinate ether derivative, d-alpha-tocopheryloxybutyric acid (TSE). None of the tocopherol analogs significantly inhibited CYP2E1 activity as measured by oxidation of p-nitrophenol. Liver homogenates and subcellular fractions (cytosol, nuclei, plasma membranes, mitochondria and microsomes) were collected 18 h after tocopherol analog administration in the absence of CCl4. Homogenate and subcellular alpha-T levels were not significantly increased following TSE administration but were increased 2-3 fold following TS and alpha-T administration. Total tocopherol levels (alpha-T+ TS + TSE) in liver homogenates and subcellular fractions were highest in rats supplemented with TS. In these animals, TS was detected in all subcellular fractions and total tocopherol levels were increased from 6-23 fold over those seen in controls and 2-9 fold over alpha-T treated rats. In vitro studies in which liver homogenates and subcellular fractions were peroxidized with ascorbate and ADP/Fe suggest that increasing levels of alpha-T but not TS correlates with increased protection against lipid peroxidation. These results suggest that the ability of TS to protect against CCl4-induced hepatotoxicity relates to its enhanced hepatic accumulation and subsequent hydrolysis to alpha-T.

Alanine Transaminase↗

PubMed1988

Role of phosphatidylserine in the cellular and subcellular lung distribution of quinidine in rats.

The role of phosphatidylserine in the cellular and subcellular lung distribution of quinidine was investigated in rats, since quinidine was found to bind preferentially to phosphatidylserine. The concentration of phosphatidylserine in the cellular and subcellular fractions was determined after separation by two-dimensional thin-layer chromatography. Selective accumulation of quinidine in vivo was observed in the alveolar macrophage at the cellular level and in the plasma membrane at the subcellular level. Both alveolar macrophages and plasma membranes were rich in phosphatidylserine compared to other fractions. When plasma levels were kept at steady state by i.v. infusion, the distribution of quinidine in the lung cellular and subcellular fractions was linearly correlated with the concentration of phosphatidylserine (r = 0.906). These results suggest that the concentration of phosphatidylserine is a dominant determinant of the cellular and subcellular lung distribution of quinidine.

Animals↗

PubMed1988

Further studies on the activity and subcellular distribution of alanine:glyoxylate aminotransferase in the livers of patients with primary hyperoxaluria type 1.

1. The activity of alanine:glyoxylate aminotransferase (AGT; EC 2.6.1.44) has been measured in the unfractionated livers of 20 patients with primary hyperoxaluria type 1 (PH1), three patients with other forms of primary hyperoxaluria and one PH1 heterozygote. The subcellular distribution of AGT activity was examined in four of the PH1 livers and in the liver of the PH1 heterozygote. 2. The mean AGT activity in the unfractionated PH1 livers was 12.6% of the mean control value. The activities of other aminotransferases and the peroxisomal marker enzymes were normal. When corrected for cross-over from glutamate:glyoxylate aminotransferase (GGT; EC 2.6.1.4), the mean AGT activity in the PH1 livers was reduced to 3.3% of the control values. 3. The livers from a patient with primary hyperoxaluria type 2 (D-glycerate dehydrogenase deficiency) and one with an undefined form of primary hyperoxaluria (possibly oxalate hyperabsorption) had normal AGT levels. The livers of a very mild PH1-type variant and a PH1 heterozygote had intermediate levels of AGT activity. 4. Subcellular fractionation of four PH1 livers by sucrose gradient isopycnic centrifugation demonstrated a complete absence of peroxisomal AGT activity. The subcellular distribution of the residual AGT activity was very similar to that of GGT activity (i.e. mainly cytosolic with a small amount mitochondrial). There were no alterations in the subcellular distributions of any of the peroxisomal marker enzymes. The subcellular distribution of AGT activity in the PH1 heterozygote liver was similar to that of the control (i.e. mainly peroxisomal).

Adolescent↗

PubMed2000

Random GFP::cDNA fusions enable visualization of subcellular structures in cells of Arabidopsis at a high frequency.

We describe a general approach for identifying components of subcellular structures in a multicellular organism by exploiting the ability to generate thousands of independent transformants in Arabidopsis thaliana. A library of Arabidopsis cDNAs was constructed so that the cDNAs were inserted at the 3' end of the green fluorescent protein (GFP) coding sequence. The library was introduced en masse into Arabidopsis by Agrobacterium-mediated transformation. Fluorescence imaging of 5,700 transgenic plants indicated that approximately 2% of lines expressed a fusion protein with a different subcellular distribution than that of soluble GFP. About half of the markers identified were targeted to peroxisomes or other subcellular destinations by non-native coding sequence (i.e., out-of-frame cDNAs). This observation suggests that some targeting signals are of sufficiently low information content that they can be generated frequently by chance. The potential of the approach for identifying markers with unique dynamic processes is demonstrated by the identification of a GFP fusion protein that displays a cell-cycle regulated change in subcellular distribution. Our results indicate that screening GFP-fusion protein libraries is a useful approach for identifying and visualizing components of subcellular structures and their associated dynamics in higher plant cells.

Arabidopsis↗

PubMed1997

Human N-myristoyltransferase amino-terminal domain involved in targeting the enzyme to the ribosomal subcellular fraction.

N-Myristoyltransferase (NMT) catalyzes the cotranslational acylation with myristic acid of the NH2-terminal glycines of a number of cellular and viral proteins. Most of the in vitro NMT activity (60-85%) in isoosmotic cell homogenates of human lymphoblastic leukemia (i.e. CEM and MOLT-4) and cervical carcinoma (i.e. HeLa) cells was shown to be associated with the ribosomal subcellular fractions by differential centrifugation. Also found in the ribosomal fractions was a approximately 60-kDa protein that was specifically immunoblotted with an anti-human NMT (hNMT) peptide antibody. This approximately 60-kDa protein was stable in the presence of proteolytic enzyme inhibitors but was gradually converted into a approximately 46-kDa species when stored in the absence of protease inhibitors. Sucrose density gradient centrifugation of the ribosomal fraction resulted in the hNMT activity sedimenting exactly coincident with the 260 nm absorption profile and exhibiting A260/A280 absorption ratios >1.8, indicating an association of NMT with putative ribosomal particle(s)/subunit(s). The subcellular targeting of hNMT was also examined by immunoblotting subcellular fractions from HeLa cells transfected with plasmids containing FLAG epitope-tagged hNMT inserts corresponding either to the originally assigned hNMT gene or to an alternative open reading frame initiated from an in-frame start site upstream from the assumed hNMT start site. Anti-FLAG immunoblotting of cells transfected with a plasmid containing the larger insert revealed FLAG-NMT primarily in the ribosomal fraction with an apparent molecular mass similar to the approximately 60-kDa native hNMT. In contrast, immunoblotting of cells transfected with a plasmid containing the smaller insert identified a approximately 50-kDa FLAG-NMT predominantly in the cytosolic fraction. An analysis of mixtures of CEM ribosomes and serial dilutions of purified recombinant FLAG-NMTs demonstrated that the approximately 60-kDa FLAG-NMT binds ribosomes with higher affinity than the approximately 50-kDa FLAG-NMT. These in vivo and in vitro subcellular targeting and recombinant expression experiments identify a native hNMT that is 10-12 kDa larger than the enzyme predicted by the originally assigned hNMT gene and which is apparently translated from an alternative up-stream start site. The data also indicate that although the unique NH2-terminal residues encoded by this larger open reading frame are not required for in vitro catalytic activity, they do provide signal(s) involved in targeting hNMT to the ribosomal subcellular fraction where cotranslational N-myristoylation occurs.

Acyltransferases↗

PubMed2005

pTARGET [corrected] a new method for predicting protein subcellular localization in eukaryotes.

MOTIVATION: There is a scarcity of efficient computational methods for predicting protein subcellular localization in eukaryotes. Currently available methods are inadequate for genome-scale predictions with several limitations. Here, we present a new prediction method, pTARGET that can predict proteins targeted to nine different subcellular locations in the eukaryotic animal species. RESULTS: The nine subcellular locations predicted by pTARGET include cytoplasm, endoplasmic reticulum, extracellular/secretory, golgi, lysosomes, mitochondria, nucleus, plasma membrane and peroxisomes. Predictions are based on the location-specific protein functional domains and the amino acid compositional differences across different subcellular locations. Overall, this method can predict 68-87% of the true positives at accuracy rates of 96-99%. Comparison of the prediction performance against PSORT showed that pTARGET prediction rates are higher by 11-60% in 6 of the 8 locations tested. Besides, the pTARGET method is robust enough for genome-scale prediction of protein subcellular localizations since, it does not rely on the presence of signal or target peptides. AVAILABILITY: A public web server based on the pTARGET method is accessible at the URL http://bioinformatics.albany.edu/~ptarget. Datasets used for developing pTARGET can be downloaded from this web server. Source code will be available on request from the corresponding author.

Algorithms↗

PubMed2006

BaCelLo: a balanced subcellular localization predictor.

MOTIVATION: The knowledge of the subcellular localization of a protein is fundamental for elucidating its function. It is difficult to determine the subcellular location for eukaryotic cells with experimental high-throughput procedures. Computational procedures are then needed for annotating the subcellular location of proteins in large scale genomic projects. RESULTS: BaCelLo is a predictor for five classes of subcellular localization (secretory pathway, cytoplasm, nucleus, mitochondrion and chloroplast) and it is based on different SVMs organized in a decision tree. The system exploits the information derived from the residue sequence and from the evolutionary information contained in alignment profiles. It analyzes the whole sequence composition and the compositions of both the N- and C-termini. The training set is curated in order to avoid redundancy. For the first time a balancing procedure is introduced in order to mitigate the effect of biased training sets. Three kingdom-specific predictors are implemented: for animals, plants and fungi, respectively. When distributing the proteins from animals and fungi into four classes, accuracy of BaCelLo reach 74% and 76%, respectively; a score of 67% is obtained when proteins from plants are distributed into five classes. BaCelLo outperforms the other presently available methods for the same task and gives more balanced accuracy and coverage values for each class. We also predict the subcellular localization of five whole proteomes, Homo sapiens, Mus musculus, Caenorhabditis elegans, Saccharomyces cerevisiae and Arabidopsis thaliana, comparing the protein content in each different compartment. AVAILABILITY: BaCelLo can be accessed at http://www.biocomp.unibo.it/bacello/.

Algorithms↗

PubMed2002

Subcellular Ca2+ alternans represents a novel mechanism for the generation of arrhythmogenic Ca2+ waves in cat atrial myocytes.

Ca(2+) alternans is a potentially arrhythmogenic beat-to-beat alternation of the amplitude of the action potential-induced [Ca(2+)](i) transient in cardiac myocytes. Despite its pathophysiological significance the cellular mechanisms underlying Ca(2+) alternans are poorly understood. Recent evidence, however, points to the modulation of Ca(2+)-induced Ca(2+) release (CICR) from the sarcoplasmic reticulum (SR) by localized alterations in energy metabolism as an important determinant of Ca(2+) alternans. We therefore studied the subcellular properties of Ca(2+) alternans in field-stimulated cat atrial myocytes employing fast two-dimensional fluorescence confocal microscopy. Ca(2+) alternans was elicited by an increase in stimulation frequency or by metabolic interventions targeting glycolysis. Marked subcellular variations in the time of onset, the magnitude, and the phase of alternans were observed. Longitudinal and transverse gradients of Ca(2+) alternans were found as well as neighbouring subcellular regions alternating out-of-phase. Moreover, focal inhibition of glycolysis resulted in spatially restricted Ca(2+) alternans. When two adjacent regions within a myocyte alternated out-of-phase, steep [Ca(2+)](i) gradients developed at their border giving rise to delayed propagating Ca(2+) waves. The results demonstrate that Ca(2+) alternans is a subcellular phenomenon caused by modulation of SR Ca(2+) release, which is mediated, at least in part, by local inhibition of energy metabolism. The generation of arrhythmogenic Ca(2+) waves by subcellular variations in the phase of Ca(2+) alternans represents a novel mechanism for the development of atrial disrhythmias.

3-Hydroxybutyric Acid↗

PubMed2006

Dosimetric characterization of radionuclides for systemic tumor therapy: influence of particle range, photon emission, and subcellular distribution.

Various radionuclides have been proposed for systemic tumor therapy. However, in most dosimetric analysis of proposed radionuclides the charged particles are taken into consideration while the potential photons are ignored. The photons will cause undesirable irradiation of normal tissue, and increase the probability of toxicity in, e.g., the bone marrow. The aim of this study was to investigate the dosimetric properties according to particle range, photon emission, and subcellular radionuclide distribution, of a selection of radionuclides used or proposed for radionuclide therapy, and to investigate the possibility of dividing radionuclides into groups according to their dosimetric properties. The absorbed dose rate to the tumors divided by the absorbed dose rate to the normal tissue (TND) was estimated for different tumor sizes in a mathematical model of the human body. The body was simulated as a 70-kg ellipsoid and the tumors as spheres of different sizes (1 ng-100 g). The radionuclides were either assumed to be uniformly distributed throughout the entire tumor and normal tissue, or located in the nucleus or the cytoplasm of the tumor cells and on the cell membrane of the normal cells. Fifty-nine radionuclides were studied together with monoenergetic electrons, positrons, and alpha particles. The tumor and normal tissue were assumed to be of water density. The activity concentration ratio between the tumor and normal tissue was assumed to be 25. The radionuclides emitting low-energy electrons combined with a low photon contribution, and the alpha emitters showed high TND values for most tumor sizes. Electrons with higher energy gave reduced TND values for small tumors, while a higher photon contribution reduced the TND values for large tumors. Radionuclides with high photon contributions showed low TND value for all tumor sizes studied. The radionuclides studied could be divided into four main groups according to their TND values: beta emitters, Auger electron emitters, photon emitters, and alpha emitters. The TND values of the beta emitters were not affected by the subcellular distribution of the radionuclide. The TND values of the Auger electron emitters were affected by the subcellular radionuclide distribution. The photon emitters showed low TND values that were only slightly affected by the subcellular radionuclide distribution. The alpha emitters showed high TND values that were only slightly affected by the subcellular radionuclide distribution. This dosimetric characterization of radionuclides may be valuable in choosing the appropriate radionuclides for specific therapeutic applications.

Body Burden↗

PubMed2003

Subcellular localization of a small sporulation protein in Bacillus subtilis.

SpoVM is an unusually small (26-residue-long) protein that is produced in the mother cell chamber of the sporangium during the process of sporulation in Bacillus subtilis. We investigated the subcellular localization of SpoVM, which is believed to be an amphipathic alpha-helix, by using a fusion of the sporulation protein to the green fluorescence protein (GFP). We found that SpoVM-GFP is recruited to the polar septum shortly after the sporangium undergoes asymmetric division and that the fusion protein localizes to the mother cell membrane that surrounds the forespore during the subsequent process of engulfment. We identified a patch of three residues near the N terminus of the proposed alpha-helix that is needed both for proper subcellular localization and for SpoVM function. We also identified a patch of residues on the opposite face of the helix and residues near both ends of the protein that are needed for SpoVM function but not for subcellular localization. Subcellular localization of SpoVM-GFP was found to require an unknown gene(s) under the control of the mother cell transcription factor sigmaE. We propose that the N-terminal patch binds to an unknown anchoring protein that is produced under the control of sigmaE and that other residues important in SpoVM function to recruit an unknown sporulation protein(s) to the mother cell membrane that surrounds the forespore. Our results provide evidence that SpoVM function depends on proper subcellular localization.

Amino Acid Sequence↗

PubMedDate not supplied

Acetylcholinesterase and nonspecific cholinesterase activities in rat liver: subcellular localization, molecular forms, and some extraction properties.

Subcellular distribution and some extraction properties of acetylcholinesterase (AchE) (EC 3.1.1.7) and nonspecific cholinesterase (ChE) (EC 3.1.1.8) were studied in rat liver employing subcellular fractionation techniques. All purified subcellular fractions were enriched in total cholinesterase activity over the homogenate. Plasma membrane and Golgi fractions showed a significant enrichment in AchE activity, while ChE activity was enriched in both rough and smooth endoplasmic reticulum. Subcellular fractions were subjected to conditions that selectively release proteins having varying degrees of association to membranes. High-pH treatment (known to release peripheral and soluble proteins) extracted ChE activity, but more than 90% of AchE activity remained associated to the pellet. Solubility properties and molecular forms of AchE and ChE in this tissue were studied by extraction in high-salt medium with and without Triton X-100, followed by velocity sedimentation centrifugation. Most of AchE activity (88%) (41% G4 and 59% G2 + G1) was detergent soluble; 42% of ChE activity (detected only as G2 + G1) was high-salt soluble, whereas remaining ChE activity was detergent soluble. These results indicate not only a different subcellular location for both enzymes, but also point to a differential association to membranes. AchE behaves as an integral membrane protein and ChE behaves as a peripheral or a luminal soluble protein.

Acetylcholinesterase↗

PubMed1989

Oxygen tolerance in neonatal rats: role of subcellular superoxide generation.

We compared the superoxide anion generating capacity of subcellular fractions from the lungs of neonatal and adult rats. Microsomal and mitochondrial fractions from adult rats produced approximately three times more superoxide (nanomoles per minute per milligram protein) than fractions from neonatal rats in the presence of 100% O2. Subcellular superoxide anion generating capacity was also examined in adult and neonatal rats exposed to greater than 95% fractional concentration of O2 in inspired gas. The O2- produced by mitochondrial and microsomal fractions of adult and neonatal rats increased above control levels for the first 24 h and declined below control values after 48 h of exposure in adults, whereas the elevated O2- production was sustained in microsomal fractions of neonates through 60 h. During the course of hyperoxic exposure, the largest difference in the superoxide generating capacity between adult and neonate was observed after 8-24 h of hyperoxia. The microsomal and mitochondrial fractions from adult rats produced three to seven times more O2- compared with neonatal rats. Cu,Zn superoxide dismutase (SOD) increased during the course of hyperoxia only in neonates at 8, 24, and 48 h of exposure. No change was observed in the activity of Mn SOD. The ratio of SOD activity (units per lung) to subcellular superoxide generating capacity (nanomoles per minute per lung) was calculated for the normal adults and neonates. The ratio for adult rats averaged 23 and 17 for mitochondrial and microsomal fractions, respectively, and 51 for neonatal rats for both subcellular fractions under normoxic conditions. These results suggest that O2- tolerance of neonates may be explained by the favorable balance between antioxidant defenses and subcellular superoxide generating capacity. The role of increased activity of Cu,Zn SOD as an accompanying or a causative phenomenon in O2 tolerance of neonates could not be determined from these experiments.

Aging↗

PubMed2002

Subcellular distribution of 3 functional platelet SNARE proteins: human cellubrevin, SNAP-23, and syntaxin 2.

Morphologic studies have demonstrated a process by which alpha-granule contents are released from platelets. Studies aimed at defining the molecular mechanisms of this release have demonstrated that SNARE proteins are required for alpha-granule secretion. These observations raise the possibility that morphologic features of alpha-granule secretion may be influenced by the subcellular distribution of SNARE proteins in the platelet. To evaluate this possibility, we analyzed the subcellular distribution of 3 functional platelet SNARE proteins-human cellubrevin, SNAP-23, and syntaxin 2. Exposure of streptolysin O-permeabilized platelets to antihuman cellubrevin antibody inhibited Ca(++)-induced alpha-granule secretion by approximately 50%. Inhibition of alpha-granule secretion by antihuman cellubrevin was reversed by a blocking peptide. Syntaxin 2 and SNAP-23 have previously been demonstrated to mediate platelet granule secretion. The subcellular localization of the 3 SNARE proteins was determined by ultrastructural studies, using a pre-embedding immunonanogold method, and by immunoblot analysis of subcellular fractions. Immunonanogold localization demonstrated that approximately 80% of human cellubrevin in resting platelets was localized to platelet granule membranes. In contrast, SNAP-23 localized predominantly to plasma membrane, whereas syntaxin 2 was more evenly distributed among membranes of alpha-granules, the open canalicular system, and plasma membrane. Thus, each of these SNARE proteins has a distinct subcellular distribution in platelets, and each of these membrane compartments demonstrates a unique SNARE protein composition. This distribution provides a basis for several characteristics of alpha-granule secretion that include homotypic alpha-granule fusion and the fusion of alpha-granules with the open canalicular system and plasma membrane.

Antibodies↗

PubMed1990

Adrenal autoantibodies bind to adrenal subcellular fractions enriched in cytochrome-c reductase and 5'-nucleotidase.

A quantitative assay for human adrenal autoantibodies has been developed to aid in the detection and isolation of human adrenal antigens. To define the subcellular location(s) of the antigen(s) capable of binding with these antibodies, we have quantitated both antibody binding to various adrenal subcellular fractions and the adrenal autoantibody binding inhibition caused by each subcellular fraction. To further define the subcellular location of the autoantibody binding, each fraction was assayed for organelle-specific marker enzyme activities. Enzyme activities were correlated to adrenal autoantibody binding to each fraction by linear regression. Of the materials tested, both antibody binding and inhibition of binding were most highly correlated with adrenal subcellular fractions enriched with cytochrome-c reductase and 5'-nucleotidase (r = 0.98; P less than 0.05). Thus, our data support the localization of adrenal autoantigen(s) in the microsomes, plasma membrane, or both.

5'-Nucleotidase↗

PubMed1998

Characteristic subcellular distribution, in brain, heart and lung, of biperiden, trihexyphenidyl, and (-)-quinuclidinyl benzylate in rats.

The subcellular distribution of biperiden (BP), trihexyphenidyl (TP) and (-)-quinuclidinyl benzylate (QNB) in brain, heart and lung following high dose (3.2 mg/kg) i.v. administration was investigated in rats. The subcellular distribution of BP or TP used clinically conformed with that of QNB, a typical potent central muscarinic antagonist. The concentration-time courses of the brain subcellular fractions for these drugs were of two types which decreased slowly and in parallel to the plasma concentration. The subcellular distribution in the brain and heart was dependent on the protein amount of each fraction. The percent post-nuclear fraction (P2) of the total concentration in the lung was characteristically about 3-5 times larger than that in the heart. It was elucidated that the distribution in the lung differs from that in the brain and heart, with high affinity which is not dependent on the protein amount in the P2 fraction containing lysosomes. On the other hand, at a low dose (650 ng/kg) of 3H-QNB, each fraction as a percentage of the total concentration in the brain increased in synaptic membrane and synaptic vesicles and decreased in nuclei and cytosol as compared with the high dose. These results show that although the tissue concentration-time courses of anticholinergic drugs appear to decrease simply in parallel to plasma concentration, the subcellular distribution exhibits a variety of patterns among various tissues.

Animals↗

PubMed2004

Signaling in small subcellular volumes. I. Stochastic and diffusion effects on individual pathways.

Many cellular signaling events occur in small subcellular volumes and involve low-abundance molecular species. This context introduces two major differences from mass-action analyses of nondiffusive signaling. First, reactions involving small numbers of molecules occur in a probabilistic manner which introduces scatter in chemical activities. Second, the timescale of diffusion of molecules between subcellular compartments and the rest of the cell is comparable to the timescale of many chemical reactions, altering the dynamics and outcomes of signaling reactions. This study examines both these effects on information flow through four protein kinase regulatory pathways. The analysis uses Monte Carlo simulations in a subcellular volume diffusively coupled to a bulk cellular volume. Diffusion constants and the volume of the subcellular compartment are systematically varied to account for a range of cellular conditions. Each pathway is characterized in terms of the probabilistic scatter in active kinase levels as a measure of "noise" on the pathway output. Under the conditions reported here, most signaling outcomes in a volume below one femtoliter are severely degraded. Diffusion and subcellular compartmentalization influence the signaling chemistry to give a diversity of signaling outcomes. These outcomes may include washout of the signal, reinforcement of signals, and conversion of steady responses to transients.

Calcium↗

PubMed1975

Subcellular distribution of phospholipids in different types of skeletal muscle.

Subcellular distribution of choline and non-choline phosphatides has been studied in tetanic (fast-twitch) and tonic (slow-twitch) muscles of rabbits. The choline phosphatide content of the subcellular fraction including the sarcolemma was greater in the tetanic than in the tonic muscle. On the other hand, the choline phosphatide content of the mitochondria-free sarcoplasmic fraction was greater in the tonic than in the tetanic muscle. A greater amount of non-choline phosphatide was found in each subcellular fraction of the tonic muscle as compared with those of the tetanic one. There was more fatty aldehyde in the non-choline phosphatides of each subcellular fraction of the tetanic muscle, than in those of the tonic one, of this type being much smaller in the tetanic muscle. There is not such an expressed difference in the fatty aldehyde contents of choline phosphatides of the subcellular fractions between the two kinds of muscle.

Aldehydes↗

PubMed1998

Subcellular tension fields and mechanical resistance of the lamella front related to the direction of locomotion.

Keratocytes derived from the epidermis of aquatic vertebrates are now widely used for investigation of the mechanism of cell locomotion. One of the main topics under discussion is the question of driving force development and concomitantly subcellular force distribution. Do cells move by actin polymerization-driven extension of the lamella, or is the lamella edge extended at regions of weakness by a flow of cytoplasm generated by hydrostatic pressure? Thus, elasticity changes were followed and the stiffness of the leading front of the lamella was manipulated by local application of phalloidin and cytochalasin D (CD). In scanning acoustic microscopy (SAM), elasticity is revealed from the propagation velocity of longitudinal sound waves (1 GHz). The lateral resolution of SAM is in the micrometer range. Using this method, subcellular tension fields with different stiffnesses (elasticity) can be determined. A typical pattern of subcellular stiffness distribution is related to the direction of migration. Cells forced to change their direction of movement by exposure to DC electric fields of varying polarity alter their pattern of subcellular stiffness in relationship to the new direction. The cells spread into the direction of low stiffness and retract at zones of high stiffness. The pattern of subcellular stiffness distribution reveals force distribution in migrating cells; i.e., if a cell moves exactly in a direction perpendicular to its long axis, then the contractile forces are largest along the long axis and decrease toward the short axis. Locomotion in any angle oblique to this axis requires an asymmetric stiffness distribution. Inhibition of actomyosin contractions by La3+ (2 mM), which inhibits Ca2+ influx, reduces cytoplasmic stiffness accompanied by an immediate cessation of locomotion and a change of cell shape. Local release of CD in front of a progressing lamella activates a cell to follow the CD gradient: The lamella thickens locally and is extended toward the tip of the microcapillary. Release of phalloidin stops extension of the lamella, and the cell turns away from the releasing microcapillary. The response to CD is assumed to be the result of local weakening of the cytoplasm due to severing of the actin fibrils. Phalloidin is supposed to stabilize the leading front by inhibition of F-actin depolymerization. These observations are in favor of the assumption that migration is due to an extension of the cell into the direction of minimum stiffness, and they are consistent with the hypothesis that local release of hydrostatic pressure provides the driving force for the flux of cytoplasm.

Animals↗