🔬Biophysics Unit 10 – Molecular Motors and Cytoskeletal Mechanics

Introduction to Molecular Motors and Cytoskeleton

• Molecular motors are specialized proteins that convert chemical energy into mechanical work, enabling crucial cellular processes such as transport, motility, and force generation
• The cytoskeleton is a dynamic network of protein filaments that provides structural support, organizes cellular components, and serves as tracks for molecular motors
• Three main types of cytoskeletal filaments are microfilaments (actin filaments), microtubules, and intermediate filaments
• Molecular motors and the cytoskeleton work together to facilitate intracellular transport, cell division, muscle contraction, and cell migration
• The study of molecular motors and cytoskeletal mechanics is essential for understanding fundamental cellular processes and their implications in health and disease

Types of Molecular Motors

• Myosins are a diverse family of motor proteins that interact with actin filaments and are involved in muscle contraction, cell migration, and intracellular transport
  • Myosin II is the primary motor protein responsible for muscle contraction
  • Myosin V and myosin VI are processive motors that transport cargo along actin filaments
• Kinesins are microtubule-associated motor proteins that primarily move towards the plus end of microtubules and are involved in intracellular transport and cell division
  • Kinesin-1 (conventional kinesin) transports organelles and vesicles towards the cell periphery
  • Kinesin-5 (Eg5) is essential for spindle formation and chromosome segregation during mitosis
• Dyneins are microtubule-associated motor proteins that move towards the minus end of microtubules and are involved in intracellular transport, cell division, and cilia/flagella movement
  • Cytoplasmic dyneins transport cargo from the cell periphery towards the cell center
  • Axonemal dyneins power the beating of cilia and flagella
• RNA polymerases are molecular motors that transcribe DNA into RNA, moving along the DNA template during the process

Structure and Function of Cytoskeletal Components

• Microfilaments (actin filaments) are thin, flexible polymers of globular actin (G-actin) subunits that form double-stranded helical structures
  • Actin filaments have a polarized structure with a plus (barbed) end and a minus (pointed) end
  • They are involved in cell motility, muscle contraction, and the formation of cellular structures like microvilli and stress fibers
• Microtubules are hollow, cylindrical polymers composed of α-tubulin and β-tubulin dimers that form protofilaments
  • Microtubules have a polarized structure with a plus end and a minus end, and they exhibit dynamic instability (rapid growth and shrinkage)
  • They are involved in intracellular transport, cell division (mitotic spindle), and the formation of cilia and flagella
• Intermediate filaments are rope-like fibers composed of various proteins (e.g., keratins, lamins, and vimentins) that provide mechanical strength and resistance to shear stress
  • They are involved in maintaining cell shape, anchoring organelles, and providing structural support in tissues like skin, hair, and neurons
• Accessory proteins regulate the assembly, disassembly, and crosslinking of cytoskeletal filaments, modulating their dynamics and organization

Mechanics of Motor Proteins

• Motor proteins undergo conformational changes driven by the hydrolysis of ATP (adenosine triphosphate), which is coupled to their mechanical movement along cytoskeletal filaments
• The mechanochemical cycle of motor proteins involves ATP binding, hydrolysis, and product release, which lead to changes in the affinity of the motor for its cytoskeletal track
• The power stroke is the conformational change that generates force and displacement, propelling the motor protein along the filament
• Motor proteins exhibit processivity, which refers to the number of steps a motor can take before dissociating from its track
  • Highly processive motors (e.g., kinesin-1 and myosin V) can take multiple steps before dissociating, enabling efficient long-distance transport
• The duty ratio is the fraction of time a motor spends attached to its filament during the mechanochemical cycle, influencing its processivity and collective behavior
• Motor proteins can work individually or in teams, with multiple motors cooperating to generate higher forces and transport larger cargoes

Energy and Force Generation in Molecular Motors

• Molecular motors convert the chemical energy of ATP hydrolysis into mechanical work, which is used to generate force and movement
• The free energy released from ATP hydrolysis ($\Delta G_{ATP}$) is approximately -30 kJ/mol under cellular conditions, providing the energy for motor protein function
• The efficiency of energy conversion in molecular motors is relatively high, typically ranging from 30-60%
• The force generated by individual motor proteins is in the piconewton (pN) range, with myosin II and kinesin-1 generating forces of ~5-7 pN per motor
• The collective action of multiple motor proteins can generate higher forces, enabling the transport of large cargoes or the contraction of muscle fibers
• The force-velocity relationship describes how the speed of a motor protein varies with the applied load, providing insights into the motor's mechanochemical properties and adaptations to different cellular tasks

Regulation of Motor Activity

• The activity of molecular motors is tightly regulated to ensure spatial and temporal control over cellular processes
• Phosphorylation is a common mechanism for regulating motor protein activity, with kinases and phosphatases modulating the motor's affinity for its cytoskeletal track or its cargo
  • Myosin II is regulated by the phosphorylation of its light chain, which controls its interaction with actin filaments
  • Kinesin-1 is regulated by the phosphorylation of its tail domain, which controls its autoinhibition and cargo binding
• Calcium signaling plays a crucial role in regulating motor protein activity, particularly in muscle contraction and neurotransmitter release
  • Calcium binding to the troponin complex in muscle cells triggers the exposure of myosin-binding sites on actin filaments, enabling muscle contraction
• Accessory proteins and cofactors can modulate motor protein activity by altering their localization, cargo binding, or mechanochemical properties
  • Dynactin is a multiprotein complex that enhances the processivity and cargo-binding ability of cytoplasmic dynein
• Cellular signaling pathways, such as those involving small GTPases (e.g., Rab proteins), regulate the recruitment and activation of motor proteins at specific locations within the cell

Cytoskeletal Dynamics and Remodeling

• The cytoskeleton is a highly dynamic structure that undergoes constant remodeling in response to cellular needs and external stimuli
• Actin filaments exhibit dynamic turnover through the processes of polymerization (addition of G-actin subunits) and depolymerization (removal of subunits)
  • Actin-binding proteins (e.g., profilin, cofilin, and Arp2/3 complex) regulate actin dynamics by controlling filament nucleation, elongation, and severing
• Microtubules undergo dynamic instability, rapidly switching between phases of growth (polymerization) and shrinkage (depolymerization)
  • Microtubule-associated proteins (MAPs) and plus-end tracking proteins (+TIPs) regulate microtubule dynamics and interactions with other cellular components
• Intermediate filaments are more stable than actin filaments and microtubules but can still undergo assembly and disassembly in response to cellular signals or mechanical stress
• Cytoskeletal remodeling is essential for various cellular processes, such as cell migration, neurite outgrowth, and tissue morphogenesis
  • During cell migration, actin polymerization drives the formation of protrusions (lamellipodia and filopodia) at the leading edge, while myosin II contraction facilitates rear retraction
• Motor proteins contribute to cytoskeletal remodeling by transporting and organizing cytoskeletal components, as well as by generating forces that influence filament dynamics and organization

Applications and Relevance in Cell Biology

• Understanding molecular motors and cytoskeletal mechanics is crucial for elucidating the mechanisms behind various cellular processes and diseases
• Intracellular transport defects caused by mutations in motor proteins or cytoskeletal components are associated with neurodegenerative diseases, such as Alzheimer's, Parkinson's, and Huntington's disease
• Studying the role of molecular motors and the cytoskeleton in cell division is essential for understanding the mechanisms of chromosome segregation and the development of anti-cancer therapies targeting mitotic motors (e.g., Eg5 inhibitors)
• Investigating the mechanics of muscle contraction, which relies on the interaction between myosin II and actin filaments, is crucial for understanding muscle function and disorders, such as myopathies and cardiomyopathies
• Cytoskeletal abnormalities and motor protein dysfunction are implicated in various developmental disorders, such as lissencephaly (abnormal neuronal migration) and ciliopathies (defects in cilia formation and function)
• Biophysical techniques, such as single-molecule imaging, optical tweezers, and atomic force microscopy, have revolutionized the study of molecular motors and cytoskeletal mechanics, providing unprecedented insights into their function and regulation
• Engineered molecular motors and cytoskeletal systems have potential applications in nanotechnology, such as the development of molecular shuttles, nanoscale cargo delivery systems, and artificial muscle fibers
• Comparative studies of molecular motors and cytoskeletal components across different species provide insights into the evolution of cellular motility and the adaptation of these systems to diverse cellular environments