Characterization of the myosin-based source for second-harmonic generation from muscle sarcomeres.

Several biologically important protein structures give rise to strong second-harmonic generation (SHG) in their native context. In addition to high-contrast optical sections of cells and tissues, SHG imaging can provide detailed structural information based on the physical constraints of the optical effect. In this study we characterize, by biochemical and optical analysis, the critical structures underlying SHG from the complex muscle sarcomere. SHG emission arises from domains of the sarcomere containing thick filaments, even within nascent sarcomeres of differentiating myocytes. SHG from isolated myofibrils is abolished by extraction of myosin, but is unaffected by removal or addition of actin filaments. Furthermore, the polarization dependence of sarcomeric SHG is not affected by either the proportion of myosin head domains or the orientation of myosin heads. By fitting SHG polarization anisotropy readings to theoretical response curves, we find an orientation for the elemental harmonophore that corresponds well to the pitch of the myosin rod alpha-helix along the thick filament axis. Taken together, these data indicate that myosin rod domains are the key structures giving SHG from striated muscle. This study should guide the interpretation of SHG contrast in images of cardiac and skeletal muscle tissue for a variety of biomedical applications.

[1]  Watt W Webb,et al.  Interpreting second-harmonic generation images of collagen I fibrils. , 2005, Biophysical journal.

[2]  J. Wray Structure of the backbone in myosin filaments of muscle , 1979, Nature.

[3]  J. Sanger,et al.  Distribution and orientation of rhodamine-phalloidin bound to thin filaments in skeletal and cardiac myofibrils. , 1997, Cell motility and the cytoskeleton.

[4]  Thierry Boulesteix,et al.  Second-harmonic microscopy of unstained living cardiac myocytes: measurements of sarcomere length with 20-nm accuracy. , 2004, Optics letters.

[5]  Oscillatory contraction of single sarcomere in single myofibril of glycerinated, striated adductor muscle of scallop. , 1994, The Japanese journal of physiology.

[6]  Tsung-Han Tsai,et al.  Studies of χ(2)/χ(3) Tensors in Submicron-Scaled Bio-Tissues by Polarization Harmonics Optical Microscopy , 2004 .

[7]  A. Huxley,et al.  The variation in isometric tension with sarcomere length in vertebrate muscle fibres , 1966, The Journal of physiology.

[8]  Tsung-Han Tsai,et al.  Higher harmonic generation microscopy for developmental biology. , 2004, Journal of structural biology.

[9]  Watt W. Webb,et al.  Uniform polarity microtubule assemblies imaged in native brain tissue by second-harmonic generation microscopy , 2003, Proceedings of the National Academy of Sciences of the United States of America.

[10]  K. Beck,et al.  Supercoiled Protein Motifs: The Collagen Triple-Helix and the α-Helical Coiled Coil , 1998 .

[11]  William A Mohler,et al.  Three-dimensional high-resolution second-harmonic generation imaging of endogenous structural proteins in biological tissues. , 2002, Biophysical journal.

[12]  Paul J Campagnola,et al.  Second harmonic generation imaging of endogenous structural proteins. , 2003, Methods.

[13]  Guy Cox,et al.  3-dimensional imaging of collagen using second harmonic generation. , 2003, Journal of structural biology.

[14]  C. Gregorio,et al.  To the heart of myofibril assembly. , 2000, Trends in cell biology.

[15]  R. Colby INTRINSIC BIREFRINGENCE OF GLYCERINATED MYOFIBRILS , 1971, The Journal of cell biology.

[16]  Beop-Min Kim,et al.  Polarization-dependent optical second-harmonic imaging of a rat-tail tendon. , 2002, Journal of biomedical optics.

[17]  Douglas J. Moffatt,et al.  Second-harmonic generation optical activity of a polypeptide α-helix at the air∕water interface , 2005 .

[18]  Bruce J Tromberg,et al.  Imaging coronary artery microstructure using second-harmonic and two-photon fluorescence microscopy. , 2004, Biophysical journal.

[19]  C. C. Wang,et al.  Nonlinear optics. , 1966, Applied optics.

[20]  H. Weber,et al.  Polarisationsoptik und molekularer Feinbau der Q-Abschnitte des Froschmuskels , 1935, Pflüger's Archiv für die gesamte Physiologie des Menschen und der Tiere.

[21]  Leslie M Loew,et al.  Sensitivity of second harmonic generation from styryl dyes to transmembrane potential. , 2004, Biophysical journal.

[22]  B. Millman,et al.  Structure of the cross-striated adductor muscle of the scallop. , 1976, Journal of molecular biology.

[23]  T. L. Mazely,et al.  Second‐order susceptibility tensors of partially ordered molecules on surfaces , 1987 .

[24]  L. Kowalczyk,et al.  The vertebrate skeletal muscle thick filaments are not three-stranded. Reinterpretation of some experimental data. , 2002, Acta biochimica Polonica.

[25]  N. Mora-Diez,et al.  Second-harmonic generation optical activity of a polypeptide alpha-helix at the air/water interface. , 2005, The Journal of chemical physics.

[26]  Leslie M Loew,et al.  Second-harmonic imaging microscopy for visualizing biomolecular arrays in cells, tissues and organisms , 2003, Nature Biotechnology.

[27]  L. Kowalczyk,et al.  Myosin molecule packing within the vertebrate skeletal muscle thick filaments. A complete bipolar model. , 2002, Acta biochimica Polonica.

[28]  Leslie M Loew,et al.  Second harmonic imaging microscopy. , 2003, Methods in enzymology.

[29]  J. Trinick,et al.  [2] Preparation of myofibrils , 1982 .

[30]  J. Conboy,et al.  Measuring melittin binding to planar supported lipid bilayer by chiral second harmonic generation , 2003 .

[31]  J. Conboy,et al.  Label-free chiral detection of melittin binding to a membrane. , 2003, Journal of the American Chemical Society.

[32]  Alexander L Gaeta,et al.  Delivery of nanojoule femtosecond pulses through large-core microstructured fibers. , 2002, Optics letters.

[33]  Cross-bridge cooperativity during isometric contraction and unloaded shortening of skeletal muscle , 2001, Journal of Muscle Research & Cell Motility.

[34]  H. F. Epstein,et al.  Myosin and paramyosin are organized about a newly identified core structure , 1985, The Journal of cell biology.

[35]  Preliminary three-dimensional model for nematode thick filament core. , 1995, Journal of structural biology.

[36]  F. Gannon,et al.  Fibrodysplasia Ossificans Progressiva Why Do Some People Have Two Skeletons? , 1997, Journal of clinical rheumatology : practical reports on rheumatic & musculoskeletal diseases.

[37]  J. Sanger,et al.  Analysis of myofibrillar structure and assembly using fluorescently labeled contractile proteins , 1984, The Journal of cell biology.

[38]  U. K. Laemmli,et al.  Cleavage of Structural Proteins during the Assembly of the Head of Bacteriophage T4 , 1970, Nature.

[39]  W. Webb,et al.  Nonlinear magic: multiphoton microscopy in the biosciences , 2003, Nature Biotechnology.

[40]  R. Craig,et al.  Purification of native myosin filaments from muscle. , 2001, Biophysical journal.

[41]  Patrick Stoller,et al.  Polarization-modulated second harmonic generation in collagen. , 2002, Biophysical journal.

[42]  Meng Han,et al.  Second-harmonic imaging of cornea after intrastromal femtosecond laser ablation. , 2004, Journal of biomedical optics.

[43]  Brian Seed,et al.  Dynamic imaging of collagen and its modulation in tumors in vivo using second-harmonic generation , 2003, Nature Medicine.

[44]  H. Huxley,et al.  Quantitative studies on the structure of cross-striated myofibrils. II. Investigations by biochemical techniques. , 1957, Biochimica et biophysica acta.

[45]  Nathan Christopher Shaner,et al.  Myofibrillogenesis in skeletal muscle cells. , 2002, Clinical orthopaedics and related research.

[46]  J. Eichler,et al.  Frequency doubling of ultrashort laser pulses in biological tissues. , 1999, Applied optics.

[47]  J. Trinick,et al.  Preparation of myofibrils. , 1982, Methods in enzymology.

[48]  W. Webb,et al.  Live tissue intrinsic emission microscopy using multiphoton-excited native fluorescence and second harmonic generation , 2003, Proceedings of the National Academy of Sciences of the United States of America.

[49]  M. Beckerle,et al.  Striated muscle cytoarchitecture: an intricate web of form and function. , 2002, Annual review of cell and developmental biology.

[50]  C. Cohen,et al.  Paramyosin and the filaments of molluscan "catch" muscles. II. Native filaments: isolation and characterization. , 1971, Journal of molecular biology.

[51]  A. E. Bukatina,et al.  Sarcomere structures in the rabbit psoas muscle as revealed by fluorescent analogs of phalloidin , 2004, Histochemistry.

[52]  C. Gregorio,et al.  Mechanisms of thin filament assembly in embryonic chick cardiac myocytes: tropomodulin requires tropomyosin for assembly , 1995, The Journal of cell biology.

[53]  J. C. Ayoob,et al.  Transfections of primary muscle cell cultures with plasmids coding for GFP linked to full-length and truncated muscle proteins. , 1999, Methods in cell biology.

[54]  Bruce J Tromberg,et al.  Selective corneal imaging using combined second-harmonic generation and two-photon excited fluorescence. , 2002, Optics letters.

[55]  B. Margulis,et al.  The species specifity of the contractile protein composition of the bivalve molluscs. , 1976, Comparative biochemistry and physiology. B, Comparative biochemistry.

[56]  K. Beck,et al.  Supercoiled protein motifs: the collagen triple-helix and the alpha-helical coiled coil. , 1998, Journal of structural biology.

[57]  Y. Nonomura,et al.  Simple and rapid purification of brevin. , 1990, Biochemical and biophysical research communications.