2520_F24_L29_Microtubules

Lecture Overview

• Course: BIOL 2520, Cell Biology, Fall 2024

• Instructor: Sreeparna Vappala

• Suggested Readings: Chapter 17, pp. 602-610

Learning Objectives (LOs)

By the end of the lesson, students should be able to:

• Review the general location and functions of microtubules.

• Describe how microtubule-organizing center (MTOC) location and orientation dictate microtubule arrangement.

• Describe the structure and assembly of microtubules from tubulin dimers.

• State the polarity of microtubules and summarize its effects on assembly and function.

• Describe the structure and function of centrosomes as microtubule-organizing centers.

• Explain the necessity of organizing centers for microtubule nucleation.

• Describe dynamic instability and its relevance to microtubule function.

• Summarize the control of dynamic instability via GTP hydrolysis.

• Contrast effects of colchicine and Taxol in cancer treatment.

• Recall cell modifications of dynamic instability in microtubules.

• Compare diffusion and microtubule-guided transport of cell components.

• Outline microtubule participation in cell polarization.

• Describe the structure and function of motor proteins.

• Differentiate between kinesins and cytoplasmic dyneins regarding microtubule movement.

• Explain the "hand-over-hand" walking mechanism of motor proteins.

• Explain motor protein-cargo interactions.

Microtubules: Function and Structure

• Microtubules are hollow tubes of protein that can rapidly disassemble and reassemble.

• They grow from the centrosome, a MTOC in animal cells.

• Functions include:

  • Positioning organelles within the cell.

  • Creating transport tracks for vesicles and organelles.

  • Forming mitotic spindles, cilia, and flagella.

Structure of Microtubules

• Subunit: Dimer of

  • α-tubulin

  • β-tubulin

• Tubulin dimers assemble to form protofilaments through non-covalent bonds.

• Protofilaments: Thirteen associate to form the hollow cylindrical structure.

• Microtubules display structural polarity:

  • Plus end: β-tubulin

  • Minus end: α-tubulin

• The rapid addition of tubulin dimers occurs at the plus end, influencing intracellular transport directionality.

Centrosome: The Microtubule-Organizing Center

• Structure includes a pair of centrioles and a protein matrix.

• The matrix contains γ-tubulin, which serves as a nucleation site for microtubule growth.

• αβ-tubulin dimers attach to γ-tubulin rings, with growth occurring at the plus end extending into the cytoplasm.

• Nucleation centers are essential for initiating polymerization.

Microtubule Arrangements in Cell Types

• Fibroblasts: Single centrosome near nucleus, creating a starburst array.

• Yeast: MTOCs in the nuclear envelope.

• Plant cells: MTOCs in nuclear envelope and cell cortex.

• Epithelial cells: Microtubules aligned with plus ends toward the basal membrane to support protein transport.

• Neurons: Microtubule bundles support axon and dendrite extensions.

Dynamic Instability of Microtubules

• Microtubules exhibit dynamic instability, switching between growth and shrinkage.

• Shrinkage occurs by tubulin dimer loss from the plus end.

• Stability is achieved through linkage to cellular structures.

• Dynamic instability is advantageous for exploratory cellular responses.

Microtubule Growth and Shrinkage

• GTP Hydrolysis:

  • Each dimer has GTP bound to β-tubulin, which hydrolyzes to GDP after addition to the microtubule.

  • GTP-bound dimers promote stable growth (forming a "GTP cap"), while GDP-bound dimers favor disassembly.

• GTP hydrolysis occurs more slowly than the addition of new dimers, contributing to microtubule stability.

Regulation of Microtubule Function

• Cells modulate dynamic instability for specific functions:

  • Mitotic spindle: Rapidly growing and shrinking.

  • Differentiated cells: Suppress instability to maintain structure.

• Microtubules in axons are oriented with plus ends towards terminals, facilitating bidirectional transport.

Transport Mechanisms: Free Diffusion vs. Microtubule Transport

• Axonal transport via microtubules can exceed 10 cm/day, much faster than diffusion.

• For instance, long axons require significantly longer for proteins to diffuse without the assistance of microtubules.

Microtubule-Associated Proteins (MAPs)

• MAPs modulate microtubule dynamics:

  • Stimulating growth (γ-tubulin ring complex, augmin).

  • Stabilizing or severing microtubules.

  • Mediating interactions with cellular structures.

Drugs Modifying Microtubule Dynamics

• Colchicine & Nocodazole: Bind to free tubulin dimers preventing polymerization.

• Taxol (Paclitaxel): Binds to microtubules, stabilizing them and inhibiting subunit loss.

• All three inhibit mitosis and are utilized in cancer treatment.

Motor Proteins and Intracellular Transport

• Motor proteins like kinesins and dyneins drive the transport of organelles and vesicles.

• Kinesin moves toward the plus end; dyneins toward the minus end.

• Structure: dimers with two globular heads and one tail for cargo interaction.

• Energy for movement is ATP hydrolysis-driven.

Mechanism of Motor Proteins

• The hand-over-hand mechanism involves:

  • ATP hydrolysis releasing the motor head from the microtubule.

  • Conformational change allows the rear head to swing forward, continuing the transport cycle.

Case Study in Kinesin Function

• Experiment using a non-hydrolyzable ATP analog revealed effects on kinesin movement, leading to the conclusion based on options provided for test case understanding.

Conclusion

• Role of microtubules is central in various cellular functions through dynamic instability, structure, and interaction with motor proteins, contributing significantly to intracellular transport and cellular organization.