Microtubules

Microtubules are long, hollow cylinders made of the protein tubulin

• Roles of Microtubules in Cells

  • organelle positioning

  • vesicle trafficking

  • mitotic spindle formation pulling apart chromosomes

  • form stable structures in cilia and flagella

    • movement of cilia and flagella depends on microtubules

• Subunits

  • tubulin: dimeric protein composed of 2 globular proteins held together by non-covalent bonds

    • alpha-tubulin

    • beta-tubulin

  • each subunit binds GTP

    • GTP bound to alpha-tubulin CANNOT be hydrolyzed

    • GTP bounds to beta-tubulin can be hydrolyzed to GDP

• Structure

  • hollow tubes composed of tubulin dimers (alpha and beta-tubulin stack together forming the wall of the MT)

  • hollow tubes made of 13 parallel protofilaments

    • inner diameter = 15nm

    • outer diameter = 25 nm

  • have structural polarity - the two ends of the polymer are different from each other - which is crucial for MT assembly and function

    • beta-tubulin end: + end

    • alpha-tubulin end: - end

  • tubulin dimers can be ADDED and REMOVED at both ends BUT at different rates

    

    	Minus End (Alpha-Tubulin)	Plus end (Beta-Tubulin)
    Characteristics	Does not readilt bind to beta-tubulin in an incoming dimer (not the right conformation)	Addition of subunits causes a conformational change in beta-tubulin that increases binding for more subunits (binds alpha-tubulin of an incoming dimer)
    Growth Rate	slow growing end	fast growing end

Microtubule Assembly

In Vivo

• A. In Vivo (inside the cell): microtubule organizing centres (MTOCs) in cells provide the right conditions for rapid nucleation of microtubules

  • microtubules grow from gamma tubulin rings of the centrosome

  • structure of centrosome

    • pairs of centrioles - in the middle

    • centrosome matrix

    • a-ring-shaped gammtubulin - floats in matrix

  • Minus end of MT is attached to the centrosome

  • microtubules originate from Microtubule Organizing Centers (MTOC) - minus end of MT attached to a MTOC

    • interphase cells

    • ciliated cell

    • dividing cell

      

In Vitro

• B. In Vitro (monomers in a tube) - initiating cytoskeletal polymerization (nucleation) to build microtubule or actin polymers is a slow process in vitro

  • Critical Concentration: the [ ] at which the length of the filament is stable

    • CC = rate of subunit addition = the rate of subunit loss

  • Critical [ ] at 2 ends of a microtubule is diff

    • Cc (minus end) > Cc (plus end)

  • dynamic instability: rapid cycles of growth and shrinkage of microtubules

    • 

      	Active	Inactive
      structure	tubulin dimers that have GTP bound beta-tubulin	tubulin dimers that have GDP bound beta
      affinity for MT	high affinity	low affinity

    

  • GTP tubulin dimers convert to GDP tubulin dimers. The end of a microtubule could be

    1. GTP-bound end: high affinity for GTP-bound tubulin → MT grows

    2. GDP-bound end: low affinity for tubulin → MT disassembles

  • GTP Cap: a region at the end of a polymerizing microtubule where GTP hydrolysis has not yet occurred

    • growing MT have a GTP at + end

  • GTP hydrolysis controls the dynamics of microtubule polymerization

    	Microtubule growth (formation of GTP cap)	Microtubule shrinks
    rate of GTP hydrolysis	addition of tubulin > faster than GTP hydrolysis	GTP hydrolysis > addition of tubulin
    structure of growing MT	straight	curved protofilaments at the end; looks like protofilaments are peeling

• treadmilling: the polymer grows at the plus end and shrinks at the minus end

  

• involves periodic phases of MT catastrophe and MT rescue

Provide examples of how proteins can interact with microtubules or tubulin to influence their structure, which, in turn, will influence function

Learning Objective 6

MAPS (capping protein) at the plasma membrane helps to influence cell shape and to form polarize cells

• function

  1. some MAPS bind microtubules to neighboring microtubules, but MT can still grow → helps establish a network of MT’s going in the same direction

  2. MAPS stabilize microtubules by binding along sides

  3. MAPs bind to the growing plus end and can link microtubules to the plasma membrane