Microtubules

Cytoskeleton

Microscopic network of protein filaments and tubules in the cytoplasm of many living cells giving them shape

Cytoskeleton Functions

1. Maintain and organise cell shape

2. Anchor organelles in place

3. Facilitate cell movement (entire cell or components within the cell)

4. Enables cytoplasmic streaming

5. Drives muscle contraction

6. Involved in endocytosis and exocytosis

7. Supports organelle movement

8. plays a role in cytokinesis

Microtubules

Largest of the eukaryotic cytoskeletal elements. Function in cellular processes centered around movement. 2 types: Cytosolic and Axonemal

Structure

Straight hollow tube composed of protofilaments. Typically 13 protofilaments align side by side to form a single microtubule. Each protofilament consists of repeating units of alpha and beta tubulin heterodimers. The 3D structure of alpha and beta tubulin are nearly identical but they share only about 40% amino acid sequence identity. Microtubules exhibit polarity, with distinct plus and minus ends.

alpha tubulin

bind GTP at a non exchangeable site called the N side. GTP does not undergo hydrolysis and is not exchanged for GDP. The N site is buried at the interface between alpha and beta tubulin within the dimer. Provides structural stability to the tubulin dimer

Beta tubulin

Binds GTP at an exchangeable site known as the E-site. GTP at this site can be hydrolysed to GDP. GDP can be exchanged for GTP, allowing dynamic instability. The E-site is exposed to the solvent on the dimer surface. Hydrolysis at the E-site regulates microtubule assembly and disassembly.

Assembly

Tubulin dimer. Oligomers. Protofilaments. Sheets of protofilaments. Closing microtubules. Elongation of microtubules.

Critical concentration

is when the tubulin heterodimer concentration at which MT assembly is exactly balanced with disassembly.

Kinetics (In vitro)- amount of light scattered by a solution containing GTP tubulin after it is warmed from 0 degrees celsius to 37 degrees celsius. Assembly is inhibited by cold and activated by warming.

Microtubule treadmiling

Plus and minus ends of microtubules have different growth rates. This reflects different critical concentrations for tubulin additions. Plus ends has a lower critical concentration. Minus end has a higher critical concentration. When free tubulin concentration is above the critical concentration of the plus ends assembly occurs. Below the critical concentration of the minus ends disassembly occurs.

Simultaneous addition of tubulin at the plus end. Simultaneous loss of tubulins at the minus end. Phenomenon observed in cells, physiological significance is not known.

Dynamic Instability model of microtubules

Growing microtubules have a cap of GTP-bound tubulin at the plus ends. GTP tubulin has a higher affinity for neighbouring subunits compared to GDP tubulin. This GTP cap stabilises the microtubule and promotes the continued addition of alpha and beta tubulin heterodimers. Shrinking microtubules have reduced levels of GTP tubulin at the plus ends. The rate of GTP tubulin addition decreases while GTP hydrolysis exceeds the rate of GTP tubulin addition. As a result, the GTP cap is lost, leading to microtubule instability and the rapid loss of GDP tubulin subunits from the tip.

Cytosolic microtubules

Maintain axons in nerve cells. Responsible for intracellular movement. Form mitotic spindles for movement of chromosomes during mitosis and meiosis. Simple tubes: 13 protofilaments - singlets

Axonemal microtubules

Found in subcellular structures for cellular movement (cilia, flagella and basal bodies). The axoneme of a cillium and a flagellum consists of a highly ordered bundle of axonemal microtubules and associated protein complexes. Doubles (cilia and flagella) and triplets (centrioles and basal bodies)

Originating

Originate from MTOC (microtubule-organizing centres). During interphase the MTOC is known as the centrosome. During cell division centrosomes are duplicated. The duplication of centrosomes results in the formation of new MTOC for each daughter cell. Abnormalities in centrosome duplication or function can lead to chromosomal instability.

y gamma tubulin ring complex

y-tubulin is a specialised tubulin isoform found primarily in the nucleation of microtubules. y-tubulin rings. y tubulin associates with several proteins called y tubulin ring proteins. Together they form……, large ring shaped structures. They act as templates for the assembly and organisation of new microtubules.

Stabilising proteins

1. Maps - Microtubule associated proteins

2. Tips - Tubulin interacting proteins

MAPs

Microtubule associated proteins. Found binding along the walls of MTs. Allow interactions with other cellular structures. Increase MT stability and affect MT bundle density. Have a domain that binds to the microtubules and another domain that is projected outwards. The length of the projecting domain determines how close the microtubules will associate.

TIPs

Tubulin-interacting proteins. Stabilize MTs by capturing their plus ends. Associate with the plus ends and stabilise them, reducing the likelihood of MT catastrophe.

Destabalising proteins

Promote MT depolymerization in one of 2 ways:

1. Bind to tubulin heterodimers and prevent them from polymerising i.e stathmin

2. Bind at the ends of MTs promoting the depolymerisation of subunits i.e catastrophins