Cytoskeleton - Actin Filaments and Myosin Motors

Chapter 17: Cytoskeleton - Actin Filaments

• Actin filaments are the smallest in diameter among the three cytoskeletal filaments.
• They form a very dynamic network of thin filaments.
• Actin filaments are essential for numerous cellular processes:
  • Cell division (contractile ring).
  • Maintenance of cell shape.
  • Cell migration (crawling).
  • Attachment to surfaces.
  • Phagocytosis (engulfment of large particles).
  • Muscle contraction.

Actin Filaments and Cellular Structures

• Actin filaments form various cellular structures:
  • Microvilli: Found in epithelial cells lining the gut.
  • Contractile bundles: Act like tiny muscles in the cell.
  • Cortical actin and filopodia: Found in migrating cells.
  • Contractile ring: Found in dividing cells.

Structure and Polarity of Actin Filaments

• Each filament is a twisted chain of actin monomers.
• Actin exhibits polarity:
  • One side of the actin monomer is distinct from the other.
  • Filaments have a plus end and a minus end.
  • The minus end has an exposed ATP-binding site.

ATP Hydrolysis in Actin Filaments

• ATP-bound actin can be added to the plus end of a filament.
• ATP is quickly hydrolyzed once actin is added to the filament.
• ADP-actin in a filament is less stable.
  • ADP-actin will fall off the minus end.

Actin Treadmilling vs. Microtubule Dynamic Instability

• Actin filaments undergo treadmilling, while microtubules exhibit dynamic instability.
• Treadmilling (Actin):
  • Actin monomers are added to the plus end.
  • ADP-actin dissociates from the minus end.
• Dynamic Instability (Microtubules):
  • Rapid growth at the plus end is associated with a GTP cap.
  • Loss of the GTP cap leads to catastrophic shrinkage.
  • The GTP cap can be re-established.

Regulation of Actin by Actin-Binding Proteins

• Actin is regulated by actin-binding proteins.
• Like microtubule-associated proteins, actin-binding proteins are the effector proteins for cell signaling pathways.

Actin Monomer-Binding Proteins and Plus-End Growth

• Profilin: Promotes the addition of actin to the plus end of a filament.
• Thymosin: Prevents monomers from adding onto a filament.

Nucleating Proteins

• Arp2/3:
  • Nucleates new filaments by branching off existing filaments.
• Formins:
  • Nucleate new individual filaments and promote elongation.

Actin Meshwork and Cell Shape

• Crosslinked actin meshwork is the basic structure of cortical actin.
  • Essential for defining cell shape.
  • Provides mechanical support to the cell membrane.

Actin Filament Bundles

• Parallel bundles:
  • More tightly packed.
  • Greater mechanical strength.
  • Form protrusions such as filopodia and microvilli.
• Contractile bundles:
  • Loosely packed.
  • Filaments are anti-parallel.
  • Spacing allows myosin motor proteins to bind.

Myosin Motor Proteins

• All actin motor proteins belong to the myosin family.

Myosin-I

• A simple motor similar to microtubule motors.
• Head domain:
  • Binds actin filament and ATP.
  • Energy for movement comes from ATP hydrolysis.
• Tail domain:
  • Binds specific cargo.
• Myosin-I motors can transport specific cargo by walking along actin filaments, similar to the activity of microtubule motors; always plus-end directed
• Myosin-I can also anchor by its tail to the cell membrane.
  • Myosin-I activity in this case pulls the membrane into new shapes.
  • This is one way actin influences cell shape.

Myosin-II

• The primary motor protein associated with contractile actin bundles.
• Exists as a dimer:
  • Each monomer has a head and a tail.
  • Each head binds actin and ATP.
  • Tails form a coiled-coil.
• Myosin-II dimers create bipolar filaments.
  • Heads are oriented in opposite directions.
  • Coiled-coil tails interact with one another to form the filament.
  • Myosin-II heads are plus-end directed.
  • Actin filaments in contractile bundles have opposite polarity.
• The myosin-II filament is bipolar:
  • The head domains in each half of the filament are oriented in opposite directions.
  • A bare region of coiled-coil tails separates the two halves of the filament.

Muscle Cells and Sarcomeres

• In muscle cells, myosin-II forms large filaments.
• The basic contractile unit of a muscle cell is the sarcomere.
• Myosin-II filaments are spaced between actin filaments.
• Actin filaments are anchored at the Z-disc.
  • The plus-end of the filament is anchored.
  • The minus-end of the filament is nearest the myosin filament.
• End-to-end arrangement of sarcomeres forms a single myofibril.
  • Sarcomeres are joined at Z-disks.
• Lateral association of myofibrils creates a muscle fiber (skeletal muscle cell).
• Highly organized sarcomeres form myofibrils.
• Muscle fibers are long, multi- nucleated cells
  • Nuclei are positioned just inside the cell membrane.
  • The bulk of the cell is made up of myofibrils.
  • The regular pattern of sarcomeres gives muscle fibers a striated appearance.
• Simultaneous activity of myosin motors in a filament results in pulling actin filaments to the center of the sarcomere.
  • This causes the sarcomere to shorten.
• Muscles contract by myosin-II motor activity.

Mechanism of Myosin Motors and ATP

• Position 1:
  • Myosin head is attached to actin.
  • No bound ATP or ADP.
• Position 2:
  • ATP binds myosin.
  • ATP binding causes myosin to release the actin filament.
• Position 3:
  • ATP is hydrolyzed, leaving ADP and phosphate bound to myosin.
  • ATP hydrolysis causes a conformational change, moving myosin forward relative to the actin filament.
• Position 4:
  • ADP-myosin binds to the actin filament.
  • Binding causes release of phosphate and tighter binding to the filament.
• Position 5:
  • Tight binding and phosphate release triggers a conformational change in myosin
  • The actin filament is still tightly bound and is shifted by the myosin activity.
  • As this "power stroke" happens, ADP is released.
• Myosin motors use energy from ATP.

Simultaneous Activation of Myosin Motors in Muscle Fibers

• T-tubules: Transverse invaginations of the cell membrane.
  • Relay signal to the sarcoplasmic reticulum.
• Sarcoplasmic reticulum (SR): Specialized ER for $Ca^{2+}$ storage.
  • SR surrounds the myofibrils.

Cell Signaling and Calcium Release in Muscle Fibers

• Acetylcholine binds to an ion channel-coupled receptor on muscle cell membranes.
• Acetylcholine triggers an action potential along the T-tubules.
• The action potential opens a voltage-gated channel on the T-tubule membrane.
  • Linked $Ca^{2+}$ channels in the SR membrane are opened.
• $Ca^{2+}$ acts as a second messenger to trigger contraction.

Calcium's Role in Sarcomere Contraction

• Without $Ca^{2+}$, actin is bound to troponin and tropomyosin
  • Tropomyosin blocks myosin from binding actin filaments
• $Ca^{2+}$ binds troponin and induces a conformational change
  • The shape change in troponin forces tropomyosin to shift position.
  • Myosin binding site is now available.
• $Ca^{2+}$ triggers sarcomere contraction by allowing myosin access to actin filaments

Diseases Related to Muscle Contraction Defects

• Myasthenia gravis:
  • Caused by antibodies that block acetylcholine receptors on muscle fibers.
  • Symptoms: droopy eyelids and mouth, general muscle weakness, difficulty breathing and swallowing.
• Tourette’s Syndrome:
  • Motor tics and vocal tics
  • Causes are neurological, but poorly understood.
  • Likely problems with neurotransmitters like dopamine or GABA
  • Both tend to relax muscles.
• Many are grouped as myopathies – diseases of the muscles
• Hypertrophic cardiomyopathy is caused by mutations in a heart- specific myosin.
  • Leads to hypercontractility in the heart muscle.
• Defects in actin or actin binding proteins can also cause disease.