Chapter 19: MICROTUBULES/INTERMEDIATE FILAMENTS
Microtubule structures:
· Subunits - a and b tubulin dimers (19-2); each subunit can bind to GTP, but only b tubulin can hydrolyze GTP; dimers organized into head to tail linear protofilaments.
· Tubules are composed of helically arranged protofilaments (19-2); 13 protofilaments per singlet microtubule (MT), but also form doublet and triplet MTs (19-3); tubules are polarized by having a and b subunits at opposite ends (19-2).
· MTs form transient structures like the mitotic spindle (19-4a) and permanent structures like flagella, sperm tails and axons (19-4b).
· MTs are nucleated at MT Organizing Centers (MTOCs); centrosomes usually contain centrioles (pre-formed MTs) that nucleate MT growth; upon depolymerization by colcemid, MTs grow from centrosomes by adding subunits from (-) end (at MTOC) to (+) end (away) (19-6); basal bodies are MTOCs for cilia and flagella (19-7); g-tubulin is found as ring structures in centrosomes and nucleates MT growth (19-8).
Tubule assembly and disassembly:
· Assembly kinetics is temperature and concentration dependent (19-9); preferential growth and loss at the + end (19-10); GTP-b tubulin is added and GTP is eventually hydrolyzed, leading to GTP caps.
· Assembly of MT occurs in three steps (19-11); morphology of assembling and disassembling MTs (19-12).
· Dynamic Instability – MTs lengthen and shorten under the same cytosolic conditions (19-13); assembly is slow and disassembly is fast; assembly and disassembly are primarily + end phenomena; concentration and GTP association determine lengthening/shortening; colchicine blocks assembly by binding to tubulin dimers; taxol stabilize MTs.
· Microtubule Associated Proteins (MAPS) bind to MTs; assembly MAPs comprised of a basic MT binding domain and an acidic projection arm (T19-1); function to bundle MTs and alter MT dynamics by stabilizing structures.
Intracellular transport:
· Fast Axonal Transport occurs along MTs; movement is anterograde and retrograde; proteins carried to terminus at different rates; cargo moves different directions simultaneously (19-20).
· Cargo – MTs provide tracks for pigment granule movement in response to external signals (19-21); vesicles move along MTs; reformation of Golgi via vesicle movement; ER associated with MTs (19-22).
· Kinesin identifies a class of motor protein; kinesin structure and domain function (19-23); mediates anterograde transport of vesicles towards + end (19-24); related proteins mediate spindle function (T19-2).
· Dynein identifies a second class of motor proteins; dynein structure (19-25); dynein types (T19-2); mediates retrograde movement towards the – end.
Cilia and Flagella:
· Flagellar movement (19-27).
· Cilia and flagella have a central bundle of MTs called an axoneme; 9 + 2 structure (19-28a); A and B tubules; connection with the basal body (19-29), which has A, B and C tubules; MAPs that hold together the axoneme (19-28a).
· Bending of axoneme caused by sliding of doublets along each other (19-31); dynein on A tubule walks along the B tubule to the – end in an ATP dependent manner; dynein has a base that attaches to the A tubule and 2 or 3 globular heads that attach to the B tubule; timing and location of active dynein regulate the bending waveform.
MTs during mitosis:
· Kinetochore attaches spindle to chromosome (19-36b); kinetochore, polar and astral MTs; interdigitation region.
· MTs dynamics during mitosis (19-41); forces mediating chromosome alignment and separation (19-42); chromosome capture (19-44); mechanisms for chromosome movement (19-45); anaphase movement starts by MT depolymerizing at the kinetochore (19-46); spindle poles separate through forces on polar and astral MTs (19-47); asters determine where cytokinesis takes place (19-48).
Intermediate Filaments (IFs):
· Differences from MTs and MFs – extremely stable, no role in motility, formed from a helical rods into ropelike structures, do not bind NTPs.
· There are 5 types (T19-3); where they are expressed.
· Structure and assembly (19-51); head and tail domains ae involved in assembly and binding other proteins; lamins disassembled via phosphorylation.
· IFAPs crosslink IFs to MTs and membranes; support nuclei internally (lamins) and externally (vimentin); attached to desmosomes and hemidesmosomes (19-56); anchor sarcomeres to the PM (19-57); use in cancer diagnosis.