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Query: UMLS:C0155339 (Brown)
 12,436 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Using x-rays from a laboratory source and an area detector, myosin layer lines and the diffuse scattering between them in the moderate angle region have been recorded. At full overlap, incubation of rigor muscles with S-1 greatly reduces the diffuse scattering. Also, three of the four actin-based layer lines lying close to the meridian (Huxley, H. E., and W. Brown, 1967. J. Mol. Biol. 30:384-434; Haselgrove, J. C. 1975. J. Mol. Biol. 92:113-143) increase, suggesting fuller labeling of the actin filaments. These results are consistent with the idea (Poulsen, F. R., and J. Lowy, 1983. Nature [Lond.]. 303:146-152) that some of the diffuse scattering in rigor muscles is due to a random mixture of actin monomers with and without attached myosin heads (substitution disorder). In relaxed muscles, regardless of overlap, lowering the temperature from 24 to 4 degrees C practically abolishes the myosin layer lines (a result first obtained by Wray, J.S. 1987. J. Muscle Res. Cell Motil. 8:62 (a). Abstr.), whilst the diffuse scattering between these layer lines increases appreciably. Similar changes occur in the passage from rest to peak tetanic tension in live frog muscle (Lowy, J., and F.R. Poulsen. 1990. Biophys. J. 57:977-985). Cooling the psoas demonstrates that the intensity relation between the layer lines and the diffuse scattering is of an inverse nature, and that the transition occurs over a narrow temperature range (12-14 degrees C) with a sigmoidal function. From these results it would appear that the helical arrangement of the myosin heads is very temperature sensitive, and that the disordering effect does not depend on the presence of actin. Measurements along the meridian reveal that the intensity of the diffuse scattering increases relatively little and does so in a nearly linear manner: evidently the axial order of the myosin heads is much less temperature sensitive. The combined data support the view (Poulsen, F. R., and J. Lowy. 1983. Nature [Lond.]. 303:146-152) that in relaxed muscles a significant part of the diffuse scattering originates from disordered myosin heads. The observation that the extent of the diffuse scattering is greater in the equatorial than in the meridional direction suggests that the disordered myosin heads have an orientation which is on average more parallel to the filament axis.
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PMID:X-ray studies of order-disorder transitions in the myosin heads of skinned rabbit psoas muscles. 174 54

Cardiac myocytes isolated from adult rat hearts were grown on laminin coated culture dishes for more than a month. During this time, the cells underwent a morphological transformation which has also been referred to by others as cell remodeling (Guo J-X, Jacobson SL, Brown DL: Cell Mot Cytoskeleton 1986;6:291-304). This results in a change in myocyte morphology from its typical in vivo cylindrical shape to one which is more pleiomorphic. Despite the long-term change in morphology, myocytes expressed for differing lengths of time several aspects of the adult phenotype as evidenced by the following: 1) maintenance of cylindrical shape and/or evident cross-striations for the first 24-48 hours in culture, 2) reappearance of cross-striations during the second week in culture, 3) little or no spontaneous contractility for the first 4 days in culture, 4) expression of only the V1 isoform of myosin for at least 7 days, and 5) altered myosin isoform expression in response to changes in environmental conditions. These factors taken together suggest that in culture the adult cardiac myocyte remains a highly differentiated cell (as opposed to possible dedifferentiation) and maintains many of its previous in vivo characteristics. Such highly differentiated adult cells should be suitable as an in vitro system for studying the direct cellular effects of factors which regulate growth and differentiation of the in vivo heart.
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PMID:Differentiation of adult rat cardiac myocytes in cell culture. 246 96

The patterns of myosin isoenzymes in fast- and slow-twitch muscles of the rat hindlimb were studied, by pyrophosphate/polyacrylamide-gel electrophoresis, with hypertrophy (induced by synergist removal) and with spontaneous running exercise of 4 and 11 weeks duration. At 11 weeks, changes with hypertrophy in the slow-twitch soleus, composed of greater than 95% SM2 (slow myosin 2) in normal muscles, were minor, and consisted of an increase in the SM1 and SM1', and a loss of intermediate myosin (IM), an isoenzyme characteristic of Type IIa fibres [Fitzsimons & Hoh (1983) J. Physiol. (London) 343, 539-550]. The changes were dramatic, however, in the fast-twitch plantaris muscle. There was a 3-fold increase in the proportion of SM. In addition, IM became the predominant isoenzyme in the profile of hypertrophied plantaris by 4 weeks. These increases were balanced by decreases in the proportion of FM2 (fast myosin 2), with FM1 completely absent from the profile at 11 weeks. The changes in the plantaris with exercise were similar in direction but not as extensive as those with hypertrophy, and FM1 remained present at control levels throughout the study. When hypertrophy and exercise were combined, the increase in slow myosin was equal to the sum of the increases with each treatment alone. Changes at 4 weeks were intermediate between those of control and 11-week muscles. Peptide mapping of individual myosin isoenzymes showed that the heavy chains of IM were different from either fast or slow heavy chains. Furthermore, IM was found to be composed of a mixture of fast and slow light chains. These changes suggest that a transformation of myosin from fast to slow isoforms was in progress in the plantaris in response to hypertrophy, via a Type-IIa-myosin (IM) intermediate stage, a phenomenon similar to that occurring in chronically stimulated fast muscles during fast-to-slow transformation [Brown, Salmons & Whalen (1983) J. Biol. Chem. 258, 14686-14692].
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PMID:Changes in skeletal-muscle myosin isoenzymes with hypertrophy and exercise. 294 96

Severin is a protein from Dictyostelium that severs actin filaments in a Ca2+-dependent manner and remains bound to the filament fragments (Brown, S. S., K. Yamamoto, and J. A. Spudich , 1982, J. Cell Biol., 93:205-210; Yamamoto, K., J. D. Pardee , J. Reidler , L. Stryer , and J. A. Spudich , 1982, J. Cell Biol. 95:711-719). Further characterization of the interaction of severin with actin suggests that it remains bound to the preferred assembly end of the fragmented actin filaments. Addition of severin in molar excess to actin causes total disassembly of the filaments and the formation of a high-affinity complex containing one severin and one actin. This severin -actin complex does not sever actin filaments. The binding of severin to actin, measured directly by fluorescence energy transfer, requires micromolar Ca2+, as does the severing and depolymerizing activity reported previously. Once bound to actin in the presence of greater than 1 microM Ca2+, severin is not released from the actin when the Ca2+ is lowered to less than 0.1 microM by addition of EGTA. Tropomyosin, DNase I, phalloidin, and cytochalasin B have no effect on the ability of severin to bind to or sever actin filaments. Subfragment 1 of myosin, however, significantly inhibits severin activity. Severin binds not only to actin filaments, but also directly to G-actin, as well as to other conformational species of actin.
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PMID:Ca2+-dependent binding of severin to actin: a one-to-one complex is formed. 642 34

A method that relates molecular structure to the forces that maintain it and to its X-ray diffraction pattern is described and applied to muscle. In a computer model, the potential energy of the movable components (here the myosin heads) is minimized by letting them move down the steepest gradient in three dimensions from a variety of starting positions. Initial values are assumed for the parameters that determine the forces, and for those that define the structure and arrangement of the fixed components. The X-ray pattern expected from the resulting structures can be calculated in a straightforward manner and compared with relevant observed data. Discrepancies can then be minimized by varying the values initially assumed for the parameters, as in the conventional "trial and error" method. This first application of the present method is concerned with the effects of the hexagonal lattice on the myosin head configuration in thick filaments of the type found in vertebrate skeletal muscle. For that purpose, a very simple model was used with the following main features: smooth cylinders for the thin filaments and for the thick filament backbones, two spherical heads attached by Hookean springs to each point of a 9/3 helix on the surface of the backbone, and repulsive forces of the electrostatic double-layer type acting between each head and all other surfaces. The myosin head configuration was calculated for an isolated thick filament and a study was made of the effects of packing such filaments into a hexagonal lattice of various side spacings in the presence or absence of thin filaments. For the isolated filament, it was found that the 9/3 helical symmetry is maintained in the myosin head configuration and that the two heads of each molecule are splayed azimuthally. When such filaments are packed into the hexagonal lattice with thin filaments present, the 9/3 helical symmetry of the myosin head configuration is lost. As the lattice side spacing is reduced, the myosin heads become increasingly displaced not only in the radial and azimuthal directions but also in the axial direction, although they interact primarily with smooth cylinders. The axial separation of the two heads in each molecule becomes different in one level from that in the other two in the 43 nm axial repeat, thus increasing the repeat in projection onto the axis from 14.3 to 43 nm. This effect may contribute to the "forbidden meridionals" described by Huxley & Brown (1967).(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Application of potential energy calculations to the determination of muscle structure from X-ray data with special reference to the configuration of myosin heads. 671 80

Myosin VIIa is a newly identified member of the myosin superfamily of actin-based motors. Recently, the myosin VIIa gene was identified as the gene defective in shaker-1, a recessive deafness in mice [Gibson, F., Walsh, J., Mburu, P., Varela, A., Brown, K.A., Antonio, M., Beisel, K.W., Steel, K.P. & Brown, S.D.M. (1995) Nature (London) 374, 62-64], and in human Usher syndrome type 1B, an inherited disease characterized by congenital deafness, vestibular dysfunction, and retinitis pigmentosa [Weil, D., Blanchard, S., Kaplan, J., Guilford, P., Gibson, F., Walsh, J., Mburu, P., Varela, A., Levilliers, J., Weston, M.D., Kelley, P.M., Kimberling, W.J., Wagenaar, M., Levi-Acobas, F., Larget-Piet, D., Munnich, A., Steel, K.P., Brown, S.D.M. & Petit, C. (1995) Nature (London) 374, 60-61]. To understand the normal function of myosin VIIa and how it could cause these disease phenotypes when defective, we generated antibodies specific to the tail portion of this unconventional myosin. We found that myosin VIIa was expressed in cochlea, retina, testis, lung, and kidney. In cochlea, myosin VIIa expression was restricted to the inner and outer hair cells, where it was found in the apical stereocilia as well as the cytoplasm. In the eye, myosin VIIa was expressed by the retinal pigmented epithelial cells, where it was enriched within the apical actin-rich domain of this cell type. The cell-specific localization of myosin VIIa suggests that the blindness and deafness associated with Usher syndrome is due to lack of proper myosin VIIa function within the cochlear hair cells and the retinal pigmented epithelial cells.
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PMID:Expression in cochlea and retina of myosin VIIa, the gene product defective in Usher syndrome type 1B. 756 24

Myo2 protein (Myo2p), an unconventional myosin in the budding yeast Saccharomyces cerevisiae, has been implicated in polarized growth and secretion by studies of the temperature-sensitive myo2-66 mutant. Overexpression of Smy1p, which by sequence is a kinesin-related protein, can partially compensate for defects in the myo2 mutant (Lillie, S. H. and S. S. Brown, 1992. Nature (Lond.). 356:358-361). We have now immunolocalized Smy1p and Myo2p. Both are concentrated in regions of active growth, as caps at incipient bud sites and on small buds, at the mother-bud neck just before cell separation, and in mating cells as caps on shmoo tips and at the fusion bridge of zygotes. Double labeling of cells with either Myo2p or Smy1p antibody plus phalloidin was used to compare the localization of Smy1p and Myo2p to actin, and by extrapolation, to each other. These studies confirmed that Myo2p and Smy1p colocalize, and are concentrated in the same general regions of the cell as actin spots. However, neither colocalizes with actin. We noted a correlation in the behavior of Myo2p, Smy1p, and actin, but not microtubules, under a number of circumstances. In cdc4 and cdc11 mutants, which produce multiple buds, Myo2p and Smy1p caps were found only in the subset of buds that had accumulations of actin. Mutations in actin or secretory genes perturb actin, Smy1p and Myo2p localization. The rearrangements of Myo2p and Smy1p correlate temporally with those of actin spots during the cell cycle, and upon temperature and osmotic shift. In contrast, microtubules are not grossly affected by these perturbations. Although wild-type Myo2p localization does not require Smy1p, Myo2p staining is brighter when SMY1 is overexpressed. The myo2 mutant, when shifted to restrictive temperature, shows a permanent loss in Myo2p localization and actin polarization, both of which can be restored by SMY1 overexpression. However, the lethality of MYO2 deletion is not overcome by SMY1 overexpression. We noted that the myo2 mutant can recover from osmotic shift (unlike actin mutants; Novick, P., and D. Botstein. 1985. Cell. 40:405-416). We have also determined that the myo2-66 allele encodes a Lys instead of a Glu at position 511, which lies at an actin-binding face in the motor domain.
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PMID:Immunofluorescence localization of the unconventional myosin, Myo2p, and the putative kinesin-related protein, Smy1p, to the same regions of polarized growth in Saccharomyces cerevisiae. 818 49

The complex of Ni(II) and the tripeptide Gly-Gly-His catalyzes, in the presence of monoperoxyphthalic acid, a zero-length protein-protein cross-linking via an oxidative radical pathway involving mainly aromatic amino acids and not at all nucleophilic residues [Brown, K. C., Yang, S.-H., and Kodadek, T. (1995) Biochemistry 34, 4733-4739]. We have taken advantage of this unprecedented cross-linking system to directly and selectively probe the solution structure and functioning of the hydrophobic interface between F-actin and skeletal myosin subfragment 1 (S-1) at the level of its aromatic components, in the absence and in the presence of nucleotides (ATP and ADP) or nucleotide analogs (AMPPNP, PPi, and ADP. AlF4). Following verification of the structure of the Ni(II)-peptide chelate and of its oxidized active form by electrospray mass spectrometry, complexes of F-actin and S-1 or proteolytic S-1 derivatives and complexes of S-1 and proteolytic F-actin derivatives were readily cross-linked under various controlled conditions without apparent alteration of the acto-S-1 recognition. The covalent adducts were identified on electrophoretic gels using specific protein labeling with the oxidation-resistant fluorophor, monobromobimane, combined with immunochemical staining. Two types of actin-heavy chain conjugates were produced. One, with a mass of 180 kDa, was formed in the rigor state or with ADP bound; the other one, with a mass of 200 kDa, was generated from the ternary complexes comprising a gamma-P-containing ligand. They were accumulated with an efficiency of 8 and 6%, respectively. For each reversible complex, the 180 kDa:200 kDa band ratio was essentially as predicted from the nucleotide-dependent A to R equilibrium mechanism of the acto-S-1 interaction in solution [Geeves, A. M., and Conibear, P. B. (1995) Biosphys. J. 68, 194s-201s]. Both covalent species resulted from the cross-linking of an actin monomer to the central 50 kDa segment, and their distinct mobilities reflect gamma-P-mediated structural changes at or near the actin-50 kDa fragment interface. Peptide mapping showed the cross-linking to take place between the 506-561 S-1 segment and the 48-113 actin stretch. The localization of these regions in the atomic F-actin-S-1 model implies that nucleotide-modulated close contacts, involving aromatic residues, are operating between the C-terminal helix of the hydrophobic strong actin-binding motif of S-1 bound to the primary actin monomer and the top portion of the adjacent lower actin subunit. The specificity of the nickel-peptide cross-linking, as assessed with the acto-S-1 complex, makes it a candidate for potential general use in investigations of the hydrophobic interactions within other protein motor-based assemblies.
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PMID:Probing the hydrophobic interactions in the skeletal actomyosin subfragment 1 and its nucleotide complexes by zero-length cross-linking with a nickel-peptide chelate. 924 2

We have previously reported that a defect in Myo2p, a myosin in budding yeast (Saccharomyces cerevisiae), can be partially corrected by overexpression of Smy1p, which is by sequence a kinesin-related protein (Lillie, S.H., and S.S. Brown. 1992. Nature. 356:358- 361). Such a functional link between putative actin- and microtubule-based motors is surprising, so here we have tested the prediction that Smy1p indeed acts as a microtubule-based motor. Unexpectedly, we found that abolition of microtubules by nocodazole does not interfere with the ability of Smy1p to correct the mutant Myo2p defect, nor does it interfere with the ability of Smy1p to localize properly. In addition, other perturbations of microtubules, such as treatment with benomyl or introduction of tubulin mutations, do not exacerbate the Myo2p defect. Furthermore, a mutation in SMY1 strongly predicted to destroy motor activity does not destroy Smy1p function. We have also observed a genetic interaction between SMY1 and two of the late SEC mutations, sec2 and sec4. This indicates that Smy1p can play a role even when Myo2p is wild type, and that Smy1p acts at a specific step of the late secretory pathway. We conclude that Smy1p does not act as a microtubule-based motor to localize properly or to compensate for defective Myo2p, but that it must instead act in some novel way.
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PMID:Smy1p, a kinesin-related protein that does not require microtubules. 947 39

The shaker-1 (Myo7a) mouse deafness locus is encoded by an unconventional myosin gene: myosin VIIA [Gibson, Walsh, Mburu, Varela, Brown, Antonio, Biesel, Steel and Brown (1995) Nature (London) 374, 62-64]. The myosin VIIA gene is expressed in hair cells in the cochlea, where it is thought to function in the development of the critical neuroepithelium where auditory transduction takes place. In order to understand better the function of myosin VIIA, we have determined the complete sequence of the mouse myosin VIIA cDNA and employed the wild-type sequence for mutational analysis of a number of shaker-1 alleles. Analysis of the mouse myosin VIIA tail sequence demonstrates a large internal repeat with regions of similarity to myosins IV, X and XII as well as members of the band 4.1 family. In addition, the myosin VIIA repeats are similar along their entire length to a tail domain from a plant kinesin. The mouse myosin VIIA tail also contains a putative Src homology 3 (SH3) domain. Along with three previously reported shaker-1 mutations, mutations for seven shaker-1 alleles in total have now been identified. The mutational changes have been analysed in terms of their predicted effect on both myosin motor head and tail domain function and the predictions related to the known phenotypes of the shaker-1 alleles. Five of the mutations lie in the motor head, and analysis of their likely effect on myosin head structure correlates well with the known severity of the shaker-1 alleles. Of the two mutations in the tail, one is a missense mutation within the kinesin and myosin IV, X and XII homology domains that substitutes a conserved amino acid and leads to a severe deafness phenotype. This and other data suggest that myosin VIIA may have properties of a myosin-motor-kinesin-tail hybrid and be involved in membrane turnover within the actin-rich environment of the apical hair cell surface.
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PMID:Mutation analysis of the mouse myosin VIIA deafness gene. 968 Feb 94


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