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Cellular Movement and Muscles Cytoskeleton and Motor Proteins All physiological processes depend on movement Intracellular transport Changes in cell shape Cell motility Animal locomotion All movement is due to the same cellular “machinery” Cytoskeleton Protein-based intracellular network Motor proteins Enzymes that use energy from ATP to move Figure 5.1 Use of Cytoskeleton for Movement Cytoskeleton elements Microtubules Microfilaments Three ways to use the cytoskeleton for movement Cytoskeleton “road” and motor protein carriers To reorganize the cytoskeletal network Motor proteins pull on the cytoskeletal “rope” Microtubules Are tubelike polymers of the protein tubulin Similar protein in diverse animal groups Multiple isoforms Are anchored at both ends Microtubule-organization center (MTOC) (–) near the nucleus Attached to integral proteins (+) in the plasma membrane Function of Microtubules Motor proteins can transport subcellular components along microtubules Vesicles and organelles Motor proteins kinesin and dynein For example, rapid change in skin color Also Movement of specialized cell projections such as cilia and flagella Distribution of chromosomes during cell division through formation of mitotic spindle Microtubules: Composition and Formation Microtubules are polymers of the protein tubulin Tubulin is a dimer of a-tubulin and b-tubulin Tubulin forms spontaneously does not require an enzyme Polarity The two ends of the microtubule are different Minus (–) end Plus (+) end Microtubule Assembly Activation of tubulin monomers by GTP Monomers join to form tubulin dimer Dimers form a single-stranded protofilament Many protofilaments form a sheet Sheet rolls up to form a tubule Dimers can be added or removed from the ends of the tubule Asymmetrical growth Growth is faster at + end Cell regulates rates of growth and shrinkage Microtubule Growth and Shrinkage Factors affecting growth/shrinkage are Local concentrations of tubulin High [tubulin] promotes growth Dynamic instability GTP hydrolysis on b-tubulin causes disassembly Microtubule-associated proteins (MAPs) Temperature Low temperature causes disassembly Chemicals that disrupt the dynamics For example, plant poisons such as taxol and colchicine Treadmilling Movement Along Microtubules Motor proteins move along microtubules Direction is determined by polarity and the type of motor protein Kinesin move in (+) direction Dynein moves in (–) direction Movement is fueled by hydrolysis of ATP Rate of movement is determined by the ATPase domain of motor protein and regulatory proteins Dynein is larger than kinesin and moves five times faster Vesicle Traffic in a Neuron Figure 5.7 Cilia and Flagella Cilia Numerous, wavelike motion Flagella Single or in pairs, whiplike movement Composed of microtubules arranged into axoneme Bundle of parallel microtubules Nine pairs of microtubules around a central pair “Nine-plus-two” Asymmetric activation of dynein causes movement animation Microtubules and Physiology - Review Table 5.1 Microfilaments Polymers composed of the protein actin Found in all eukaryotic cells Often use the motor protein myosin Movement arises from Actin polymerization Sliding filaments using myosin More common than movement by polymerization Microfilament Structure and Growth G-actin monomers polymerize to form a polymer called F-actin Spontaneous growth 6–10 times faster at + end Treadmilling Assembly and disassembly occur simultaneously and overall length is constant Capping proteins Increase length by stabilizing – end and slowing disassembly Microfilament (Actin) Arrangement Arrangement of microfilaments in the cell Tangled neworks Microfilaments linked by filamin protein Bundles Cross-linked by fascin protein Networks and bundles of microfilaments are attached to cell membrane by dystrophin protein Maintain cell shape and can be used for movement Movement by Actin Polymerization Two types of amoeboid movement Filapodia are rodlike extensions of cell membrane Neural connections Microvilli of digestive epithelia Lamellapodia are sheetlike extensions of cell membrane Leukocytes Macrophages The Cell Myosin Most actin-based movements involve the motor protein myosin Sliding filament model Myosin is an ATPase Converts energy from ATP to mechanical energy 17 classes of myosin (I–XVII) Multiple isoforms in each class All isoforms have a similar structure Head (ATPase activity) Tail (can bind to subcellular components) Neck (regulation of ATPase) Sliding Filament Model Analogous to pulling yourself along a rope Actin – the rope Myosin – your arm Alternating cycle of grasp, pull, and release Your hand grasps the rope Your muscle contracts to pull rope Your hand releases, extends, and grabs further along the rope Movement will depend on which element is immobile Sliding filament model Sliding Filament Model Two processes Chemical reaction Myosin binds to actin (cross-bridge) Structural change Myosin bends (power stroke) Cross-bridge cycle Formation of cross-bridge, power stroke, release, and extension Need ATP to release and reattach to actin Absence of ATP causes rigor mortis Myosin cannot release actin Actino-Myosin Activity Two factors affect movement Unitary displacement Distance myosin steps during each cross-bridge cycle Depends on Myosin neck length Location of binding sites on actin Helical structure of actin Duty cycle Cross-bridge time/cross-bridge cycle time Typically ~0.5 Use of multiple myosin dimers to maintain contact Muscle Cells (Myocytes) Myocytes (muscle cells) Contractile cell unique to animals Contractile elements within myocytes Thick filaments Polymers of myosin ~300 myosin II hexamers Thin filaments Polymers of ?-actin Ends capped by tropomodulin and CapZ to stabilize Proteins troponin and tropomyosin on outer surface Muscle Cells Two main types of muscle cells are based on the arrangement of actin and myosin Striated (striped appearance) Skeletal and cardiac muscle Actin and myosin arranged in parallel Smooth (do not appear striped) Actin and myosin are not arranged in any particular way Striated Muscle Cell Structure Thick and thin filaments arranged into sarcomeres Repeated in parallel and in series Side-by-side across myocyte Causes striated appearance End-to-end along myocyte Sarcomeres - Structural features Z-disk Forms border of each sarcomere Thin filaments are attached to the Z-disk and extend from it towards the middle of the sarcomere A-band (anisotropic band) Middle region of sarcomere occupied by thick filaments I-band (isotropic band) Located on either side of Z-disk Occupied by thin filament Sarcomeres Thin and thick filaments overlap in two regions of each sarcomere Each thick filament is surrounded by six thin filaments Three-dimensional organization of thin and thick filaments is maintained by other proteins Nebulin Along length of thin filament Titin Keeps thick filament centered in sarcomere Attaches thick filament to Z-disk Figure 6.19 - - H Zone A Band I Band I Band Three-Dimensional Structure of Sarcomere Figure 5.18 Iworx Animation But, how does a muscle contract? Thick and thin filaments (myofilaments) slide past one another, but do not shorten Consequently, I band and H zone shrink Cross-bridges leads to thin filaments being pulled toward the center of the sarcomere Muscle Actinomyosin Activity is Unique Myosin II cannot drift away from actin Structure of sarcomere Duty cycle of myosin II is 0.05 (not 0.5) Each head is attached for a short time Does not impede other myosins from pulling the thin filament Unitary displacement is short Small amount of filament sliding with each movement of the myosin head Contractile Force Contractile force depends on overlap of thick and thin filaments More overlap allows for more force Amount of overlap depends on sarcomere length as measured by distance between Z-disks Maximal force occurs at optimal length Decreased force is generated at shorter or longer lengths Myofibril In muscle cells, sarcomeres are arranged into myofibrils Single, linear continuous stretch of interconnected sarcomeres (i.e., in series) Extends the length of the muscle cell Have parallel arrangement in the cell More myofibrils in parallel can generate more force Regulation of Contraction Excitation-contraction coupling (EC coupling) Depolarization of the muscle plasma membrane (sarcolemma) Elevation of intracellular Ca2+ Contraction Sliding filaments Relaxation Sarcolemma repolarizes and Ca2+ returns to resting levels How is this system regulated? Ca2+ Allows Myosin to Bind to Actin At rest, cytoplasmic [Ca2+] is low Troponin-tropomyosin cover myosin binding sites on actin As cytoplasmic [Ca2+] increases Ca2+ binds to TnC (calcium binding site on troponin) Troponin-tropomyosin moves, exposing myosin-binding site on actin Myosin binds to actin and cross-bridge cycle begins Cycles continue as long as Ca2+ is present Cell relaxes when the sarcolemma repolarizes and intracellular Ca2+ returns to resting levels Regulation of Contraction by Ca2+ Figure 5.22 Ionic Events in Muscle Contraction Figure 5.23 Isoforms Properties of isoforms affect contraction For example, fTnC has a higher affinity for Ca2+ than s/cTnC Muscle cells with the fTnC isoform respond to smaller increases in cytoplasmic [Ca2+] Isoforms differ in the affect of temperature and pH Properties of myosin isoforms affect contraction Multiple isoforms of myosin II in muscle Isoforms can change over time Excitation of Vertebrate Striated Muscle Skeletal muscle and cardiac muscle differ in mechanism of excitation and EC coupling Differences include Initial cause of depolarization Time course of the change in membrane potential (action potential) Propagation of the action potential along the sarcolemma Cellular origins of Ca2+ Action Potentials APs along sarcolemma signal contraction Na+ enters cell when Na+ channels open Depolarization Voltage-gated Ca2+ channel open Increase in cytoplasmic [Ca2+] Na+ channels close K+ leave cell when K+ channels open Repolarization Reestablishment of ion gradients by Na+/K+ ATPase and Ca2+ ATPase Although these events are similar, muscles show important differences in the time course of membrane potential change Time Course of Depolarization Figure 5.24 Key point: length of effective refractory period Important for Function Initial Cause of Depolarization Myogenic (“beginning in the muscle”) Spontaneous For example, vertebrate heart Pacemaker cells Cells that depolarize fastest Unstable resting membrane potential Neurogenic (“beginning in the nerve”) Excited by neurotransmitters from motor nerves For example, vertebrate skeletal muscle Can have multiple (tonic) or single (twitch) innervation sites T-Tubules and Sarcoplasmic Reticulum AP Conductance is Facilitated by: Transverse tubules (T-tubules) Invaginations of sarcolemma Enhance penetration of action potential into myocyte More developed in larger, faster twitching muscles Less developed in cardiac muscle Sarcoplasmic reticulum (SR) Stores Ca2+ bound to protein sequestrin Terminal cisternae increase storage T-tubules and terminal cisternae are adjacent to one another Ca2+ Channels and Transporters Channels allow Ca2+ to enter cytoplasm Ca2+ channels in cell membrane Dihydropyridine receptor (DHPR) – L-type Ca2+ channels in the SR membrane Ryanodine receptor (RyR) Transporters remove Ca2+ from cytoplasm Ca2+ transporters in cell membrane Ca2+ ATPase Na+/Ca2+ exchanger (NaCaX) Ca2+ transporters in SR membrane Ca2+ ATPase (SERCA) Ca2+ Channels and Transporters Figure 5.27 How is DHPR actn linked to RyR actn? Is this enough? Induction of Ca2+ Release From SR Ca2+ enters cell from extracellular fluid In heart, ? [Ca2+] causes RyR to open, allowing release of Ca2+ from SR “Ca2+ induced Ca2+ release” AP along sarcolemma conducted down T-tubules Depolarization opens DHPR In skeletal muscle, change in DHPR shape causes RyR to open, allowing release of Ca2+ from SR “Depolarization induced Ca2+ release” EGTA? Relaxation Repolarization of sarcolemma Reestablish Ca2+ gradients Remove Ca2+ from cytoplasm Ca2+ ATPase in sarcolemma and SR Na+/Ca2+ exchanger (NaCaX) in sarcolemma Very important due to strong driving force on Na+ entry Parvalbumin Cytosolic Ca2+ binding protein buffers Ca2+ Ca2+ dissociates from troponin Tropomyosin blocks myosin binding sites Myosin can no longer bind to actin Relaxation Figure 5.27 Summary of Striated Muscles Table 5.5 Smooth Muscle Slow, prolonged contractions Often found in the wall of “tubes” in the body Blood vessels, intestine, airway, etc. Smooth Muscle Key differences from skeletal muscle No sarcomeres (no striations) Thick and thin filaments are scattered in the cell Attached to cell membrane at adhesion plaques No T-tubules and minimal SR Often connected by gap junctions Function as a single unit Contract in all dimensions Different mechanism of EC coupling Control of Smooth Muscle Contraction Regulated by nerves, hormones, and physical conditions (e.g., stretch) At rest, the protein caldesmon is bound to actin and blocks myosin binding Smooth muscle does not have troponin Stimulation of cell increases intracellular Ca2+ Ca2+ binds to calmodulin Calmodulin binds caldesmon and removes it from actin Cross-bridges form and contraction occurs Calmodulin also causes phosphorylation of myosin Increase in myosin ATPase activity Diversity of Muscle Fibers Different protein isoforms affect EC coupling Ion channels Ion pumps Ca2+-binding proteins Speed of myosin ATPase Variation in other properties of muscle cells Myoglobin content Number of mitochondria Skeletal muscle cells can be classified as “fast,” “slow,” “white,” “red,” “oxidative,” “glycolytic” Changing Fiber Types Developmental (from embryo to adult) Increased proportion of fast muscle isoforms Physiological response For example, exercise Can change both cardiac and skeletal muscle Changes due to hormonal and nonhormonal mechanisms For example, thyroid hormones repress expression of b-myosin II gene and induce a-myosin II gene a-myosin II exhibits the fastest actino-myosin ATPase rates For example, direct stimulation of cell can alter gene expression Nonhormonal Mechanisms - local Figure 5.33 Trans-Differentiation of Muscle Cells Trans-differentiation Cells used for novel functions For example, heater organs of billfish eye Specialized muscle cells Few myofibrils (little actin and myosin) Abundant SR and mitochondria Futile cycle of Ca2+ in and out of the SR High rate of ATP synthesis and consumption Electric organs Invertebrate Muscles Variation in contraction force due to graded excitatory postsynaptic potentials (EPSP) Innervation by multiple neurons EPSPs can summate to give stronger contraction Some nerve signals can be inhibitory Figure 5.35 Asynchronous Insect Flight Muscles Wing beats: 250–1000 Hz Fastest vertebrate contraction ? 100 Hz (toadfish sonic muscle) freesound :: sounds that don't sound like :: mosquito.mp3 Asynchronous Insect Flight Muscles Asynchronous muscle contractions Contraction is not synchronized to nerve stimulation Stretch-activation Sensitivity of the myofibril to Ca2+ changes during contraction/relaxation cycle Intracellular [Ca2+] remains high Contracted muscle is Ca2+ insensitive Muscle relaxes Stretched muscle is Ca2+ sensitive Muscle contracts Mollusc (Bivalve) Catch Muscle Muscle that holds shell closed Capable of long duration contractions with little energy consumption Protein twitchin may stablilize actin-myosin cross-bridges Cross-bridges do not continue to cycle Phosphorylation/dephophorylation of twitchin regulates its function Mollusc (Bivalve) Catch Muscle Figure 5.37 Control of Smooth Muscle Contraction Figure 6.39 Cross-bridge formation Functions like TnC-TnI in Skeletal Muscle; but does not directly bind Ca2+ Control at both thin and thick filaments Table1. Comparison of Muscle types. Property Skeletal Muscle Cardiac Muscle Smooth Muscle Striations? Yes Yes No Relative Speed of Contraction Fast Intermediate Slow Voluntary Control? Yes No No Membrane Refractory Period Short Long Nuclei per Cell Many Single Single Control of Contraction Nerves Beats spontaneously but modulated by nerves Nerves Hormones Stretch Cells Connected by Intercalated Discs or Gap Junctions? No Yes Yes
