MCB 252 Topic 19 Regulation of Actin AssemblyProf David Rivier MCB 252 Spring 2015 MCB 252 Cell Biology Topic 19 Regulation of Actin Assembly Reading Lodish 17 2 17 3 17 7 MCB 252 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 Actin Cytoskeleton Properties of Actin Actin assembly Role of ATP Treadmilling Role of actin in cell movement Actin is required for movement Actin assembly can drive movement Regulation of actin assembly in cells Proteins involved in actin assembly disassembly Turning the protein involved in assembly and disassembly ON and OFF Building stable actin structures How Does a Cell Regulate Where Assembly Occurs What are the properties of Actin polymerization in cells 1 Nucleation is regulated so that polymer formation only occurs is specific regions of the cell 2 Dissociation of monomers from the polymer is regulated so that dissociation primarily occurs at the minus end 3 Association of monomers with the polymer is regulated so that growth occurs almost exclusively at the plus end 4 Regulation is mediated by proteins that interact with Gactin or with the ends of F actin How is assembly of actin restricted to particular regions of the cell of the cytoplasm 1 Regulation is mediated by proteins that interact with Gactin or with the ends of F actin 2 Signals are received by one side of a cell but not another Signaling regulates the function of the proteins involved in actin assembly First we will look at the mechanisms by which proteins modulate F actin assembly disassembly Then we will look at regulation how the proteins that modulate assembly are themselves turned ON or OFF in response to signaling Proteins that Facilitate Various aspects of F actin assembly A Parts List STEP Sequestering G actin Nucleation Elongation Disassembly PROTEIN S Thymosin 4 Formin Arp2 3 complex Formin Profilin Cofilin Gelsolin PREVENTING ASSEMBLY AND DISSASEMBLY Capping Proteins Plus end CapZ Minus end Tropomodulin Question How can cells assemble actin polymers and move faster than they can produce actin Answer The concentration of G actin in the cell is 100 x greater than the critical concentration BUT G actin in sequestered in a pool that is not in a form that can be polymerized Binding of Thymosin 4 to G actin holds G actin in a form that does not polymerize To form a polymer G actin must be released from Thymosin 4 Thymosin b 4 Binding to Actin How is Sequestration of ATP actin by Thymosin 4 Regulated Sequestered Free Answer We do not know Possibility 1 signaling Possibility 2 equilibrium buffering Profilin Role 1 Nucleotide exchange factor Profilin Profilin binds G actin so that the surface that binds the plus end is exposed and the surface that binds the minus end is blocked Profilin binds at hinge Cofilin Facilitates disassembly of ADP actin minus end Cofilin induces torsional stress to ADP actin polymers thereby promoting dissociation Gelsolin Family Severs actin filaments Binds plus end at point of severing creates new minus end prevents grow at new plus end allows disassembly from new minus end Gelsolin regulate by Ca Facilitation of Treadmilling Some aspects of this figure are misleading individual actin molecules don t necessarily go through every step in order Proteins that Facilitate Various aspects of F actin assembly A Parts List STEP Sequestering G actin Nucleation Elongation Disassembly PROTEIN S Thymosin 4 Formin Arp2 3 complex Formin Profilin Cofilin Gelsolin PREVENTING ASSEMBLY AND DISSASEMBLY Capping Proteins Plus end CapZ Minus end Tropomodulin Formin facilitates nucleation and elongation Note this is only part of the Formin protein the FH2 domain Formin facilitates assembly of unbranched linear polymers Formin Nanotechnology T19M1Formin mov Formin accelerate the rate at which equilibrium is reached Formin is a Machine Profilin Role 2 increase the rate of actin assembly at end Formin arms bind profilin to further facilitate assembly at end Proteins that Facilitate Various aspects of F actin assembly A Parts List STEP Sequestering G actin Nucleation Elongation Disassembly PROTEIN S Thymosin 4 Formin Arp2 3 complex Formin Profilin Cofilin Gelsolin PREVENTING ASSEMBLY AND DISSASEMBLY Capping Proteins Plus end CapZ Minus end Tropomodulin Actin Related Proteins Arps Arp2 3 Complex a Second Way to Nucleate Arp2 3 complex nucleates branched actin polymers Another view of nucleation by the Arp2 3 complex Repeated Rounds of Arp2 3 nucleation results in branched actin network Another view of nucleation by the Arp2 3 complex Proteins that Facillitate Various aspects of F actin assembly A Parts List STEP Sequestering G actin Nucleation Elongation Disassembly PROTEIN S Thymosin 4 Formin Arp2 3 complex Formin Profilin Cofilin Gelsolin PREVENTING ASSEMBLY AND DISSASEMBLY Capping Proteins Plus end CapZ Minus end Tropomodulin Capping Proteins Four possible consequences of capping 1 Prevent further growth 2 Restrict growth or shrinkage to one end 3 Form a stable filament not growing shrinking or t milling 4 Stop polymerization in regions where you don t want it Proteins work in coordination Lodish 17 41d How does polymerization generate force on the membrane We will come back to this question later Actin Cytoskeleton Properties of Actin Actin assembly Role of ATP Treadmilling Role of acting in cell movement Actin is required for movement Actin assembly can drive movement Regulation of actin assembly in cells Proteins involved in actin assembly disassembly Turning the protein involved in assembly and disassembly ON and OFF Building stable actin structures How does a cell regulate where assembly occurs Logic of Actin Regulation In Movement Cells can respond quickly to signals Cells can rearrange their actin cytoskeleton quickly To achieve these quick responses cells have all the components and machinery synthesized but the most of the components are in an inactive form Signaling results in activation of the components 3 main types of actin filaments involved in this type of motion Filopodia Filopodia and the Lamellopod are at leading edge branched Stress fibers will pull the tail end forward talk about later How does signaling set up and maintain cell polarity RAS superfamily Alberts Table 15 5 Families often named for the first member identified Activation of Ras Family Members is Similar Signaling triggers activation GAP resets the signal making the