MCB 252 Topic 21 Muscle Cells and Myosin MotorsProf David Rivier MCB 252 Spring 2015 MCB 252 Cells Tissues and Development Topic 21 Muscle Cells and Myosin Motors Reading Lodish 17 5 17 6 and pp 819 820 Outline Actin Cytoskeleton A Properties of Actin B Role of Actin in Cell Movement C Regulation of Actin Assembly in Cells D Stable Actin Structures E Muscle Cells and Myosin Motors Muscle Cells and Myosin Motors 1 Overview of Muscle Cells and Contraction What we Know How we Know it 2 How Do Myosin Motors Work 3 Additional Proteins and Design Features 4 Regulation of Contraction 5 Additional Uses of Contractile Bundles 6 Myosin Domains 7 Myosin Superfamily Recall from MCB150 Sliding Filament Model What you need to know and do not need to know Muscle Structure Lodish 17 31a Sarcomere Structure EM Cross Section of Muscle Fiber Huxley 1969 Hexagonal arrays of thick and thin filaments Image courtesy Dr John Heuser Wash U Med School Muscle Cells and Myosin Motors 1 Overview of Muscle Cells and Contraction What we Know How we Know it 2 How Do Myosin Motors Work 3 Additional Proteins and Design Features 4 Regulation of Contraction 5 Additional Uses of Contractile Bundles 6 Myosin Domains 7 Myosin Superfamily Some highlights of early biochemical work 1859 K hne isolated a muscle protein that he called myosin 1880 T H Huxley showed that A bands were lost after treatments which removed myosin he concluded that A bands were composed mainly of myosin 1938 Albert Szent Gy rgyi 1937 Nobel Prize for discovery of Vitamin C began his studies on muscle How does contraction work First need an experimental model Albert Szent Gy rgyi s choice Criteria need muscle with long parallel fibers and little connective tissue It is easy to find the muscle which contains the least connective matter for it is the quantity of connective tissue which decides the culinary value of a muscle to which it is inversely proportional The culinary value is expressed numerically on the right side of any menu A significant difference is always found at the level of Musculus psoas Psoas muscle a k a tenderloin Glycerinated muscle fiber prep Szent Gy rgi Dissect out small bundles of muscle 2mm diameter 8 cm long place in 50 glycerol at 0 C then transfer to 20 C store for months Plasma membrane is disrupted soluble components leak out but insoluble components of contractile apparatus remain intact Glycerol prevents formation of ice crystals which could disrupt structure muscle fiber at rest add ATP contracts Went on to show that purified actin myosin filaments could contract after addition of ATP Different Types of Light Microscopy Bright Field Differential Interference Contrast Phase Contrast Lodish Fig 9 9 pages 404 408 Sarcomere structure EM Z disk I band H zone A band M line Z disk I band Lodish 17 31b Some terms defined A band anisotropic I band isotropic H zone from Hell German for bright M line from Mittel German for middle Z disc from Zwischen German for between Phase contrast images of contracting myofibrils Huxley and Hanson 1954 Blended glycerol extracted psoas muscle to generate fine myofibrils 2 m Imaged contraction by phasecontrast microscopy A bands remain constant length during contraction 1 4 Extract myosin A bands disappear 5 6 What you need to know and do not need to know Sliding filament model Huxley and Huxley A F Huxley Hugh Huxley Thick and thin filaments maintain constant length during contraction slide past each other during contraction Experimental evidence from phase contrast microscopy differential interference contrast microscopy electron microscopy X ray diffraction patterns Pollard and Earnshaw 42 11 Actin myosin cross bridges in rigor mortis the stiffness of death T E M Pollard and Earnshaw 42 11c When ATP is depleted myosin heads bind tightly to actin creating crossbridges muscle becomes rigid ATP can be added back and muscle will lose its stiffness Lodish 19 22c Muscle Cells and Myosin Motors 1 Overview of Muscle Cells and Contraction What we Know How we Know it 2 How Do Myosin Motors Work 3 Additional Proteins and Design Features 4 Regulation of Contraction 5 Additional Uses of Contractile Bundles 6 Myosin Domains 7 Myosin Superfamily Myosin II Structure Myosin Movement Lodish 17 26 Lodish 17 26 Movement Results from a Series of Conformational Changes Lodish 17 26 Lodish 17 26 Lodish 17 26 Myosin Movement T21M1 Lodish 1705 myosin crossbridge swf Lodish 17 26 If sarcomeres only contract how can we flex or extend our limbs Opposing motions flexion bending of limb at joint extension straightening of limb Different muscles are responsible for these opposing motions flexor e g biceps extensor e g triceps adductor motion toward midline abductor motion away from midline Widmaier et al Human Physiology Fig 9 31 Muscle Cells and Myosin Motors 1 Overview of Muscle Cells and Contraction What we Know How we Know it 2 How Do Myosin Motors Work 3 Additional Proteins and Design Features 4 Regulation of Contraction 5 Additional Uses of Contractile Bundles 6 Myosin Domains 7 Myosin Superfamily These are the Basic Components of Contraction Other Proteins Provide Added Features Components of the Sarcomere Design Features CapZ and tropomodulin caps stability Titin spring Nebulin ruler to form filaments of fixed size Components of the Sarcomere Titin largest known protein in the human genome 38 000 amino acids 1 2 m long connection to Z disc keeps thick filaments centered during contraction Flexible PEVK domains provide elasticity Pollard and Earnshaw 42 7 2 Components of the Sarcomere Components of the Sarcomere Role of Telethonin was only figured out in 2006 Limb girdle Muscular Dystrophy Muscle Cells and Myosin Motors 1 Overview of Muscle Cells and Contraction What we Know How we Know it 2 How Do Myosin Motors Work 3 Additional Proteins and Design Features 4 Regulation of Contraction 5 Additional Uses of Contractile Bundles 6 Myosin Domains 7 Myosin Superfamily Logic of Regulation Muscles are maintained in a relaxed state contraction occurs when brain or neurons send signal to muscle The relaxed state is maintained by proteins that bind to actin and block the binding of myosin The signal to contract results in movement of the blocking protein which exposes the myosin binding site allowing myosin to bind actin and move the actin filament An electrical signal in the neuron is converted into a chemical signal within the cell Additional Components of the Sarcomere Mediate Regulation Regulation of Contraction