L312/Spring 2007 Lecture 15 Drummond March 1 For today: Focus on Chapter 17, 579-592: Actin 592-607 We won’t cover cilia and flagellar movement section (590-591) Look at microtubule movies today for review! Review sessions? Last time Monday at 7, Tuesday at 4. Change? So far: Intermediate filaments (10 nm bundles) single proteins (~50 nm) form long coiled coils (strength!) 8 filaments form a ‘seven around one’ strong fiber (twisted cable) crucial for structural integrity (cell, nuclear lamina) Reversible! Microtubules Hollow, assembled from tubulin heterodimers (a/b) Grow out of organizing body (centrosome; basal body) POLAR (+/- ends) filaments, support directional movement kinesins and dyneins are motor proteins that track along microtubules kinesins (- to +) and dyneins (+ to -) for moving cargo crucial for organizing cell Reversible! Today: Actin: Shares features of each structural cortex for cell polymeric globular protein with a polarity motors proteins (e.g. myosins) can bind and utilize
~10 nm What does: Colchicine or Taxol do to Microtubules? Why are these Anticancer agents?
Kinesins and dyneins are directional motor proteins Organelle Transport Movie Describe heavy and light chains + cargo binding regions for transport specificity But how does this “walk” What is the step size?
Why is the squid axon a good model system for studying transport Red = mitochondrion Blue = vesicle
Movement of mitochondria along microtubules why don’t motor proteins smack into each other? What is carrying these organelles along the microtubule? 400 msec (roughly 1/2 second) Intervals between images The cell is a dynamic entity!!!
Microtubules as cellular organizers (reversible!!!) ER Golgi Green = microtubules Blue = ER microtubules
Actin filaments: Where: How big? What do they contribute? How do they compare with intermediate filaments and microtubules?
Contexts for finding actin filament function (how can a 7 nm fiber carry out all these roles?) Lamellipodia (sheets) Filopodia (fingers) Contractile bundles Villi support Contractile rings
Actin filaments contain a single, polar protein monomer (not +/- charge) Why do the filaments coil?
How do the filaments polymerize? cytochalasins block polymerization jasplakinolinds block depolymerization How does this compare with microtubule polymerization?
Comparing actin filaments with microtubule filaments Microtubules actin Diameter ……………………… Number of proteins in polymer…. Identity of protein in polymer …. NTP associated …………………… Relative fiber stability with NDP (greater or less) … End where monomers may add … This binds monomer …………… (blocks polymerization) This binds polymer ……………. (blocks dissociation) Polymer can affect cell shape …. Polymer can support cell migration ……………… Key roles of polymer in cell …….
How is actin polymerization and structure modulated? Thymosin profilin
A global view of cellular migration based on actin polymerization What is the role Of integrins? What must they Look like?
EM pictures of lamellipodia--note density and branches Platinum shadowing to reveal structure
Different extracellular signals trigger different actin responses What is the experiment here? What is the value of an integrated response?
Anchoring points to a glass slide (human fibroblast) Reflection/interference fluorescent antibody to actin
The actin superhighway Key questions: how does myosin-1 resemble kinesin? How does actin resemble microtubules? What is the key rule about direction of movement? Why vesicles? (now granules with mRNA)
Add ATP Release Step/reattach Dissociate ADP Football game analogy