BACKGROUND: The labile nature of microtubules is critical for establishing cellular morphology and motility, yet the molecular basis of assembly remains unclear. Here we use optical tweezers to track microtubule polymerization against microfabricated barriers, permitting unprecedented spatial resolution. RESULTS: We find that microtubules exhibit extensive nanometer-scale variability in growth rate and often undergo shortening excursions, in some cases exceeding five tubulin layers, during periods of overall net growth. This result indicates that the guanosine triphosphate (GTP) cap does not exist as a single layer as previously proposed. We also find that length increments (over 100 ms time intervals, n = 16,762) are small, 0.81 +/- 6.60 nm (mean +/- standard deviation), and very rarely exceed 16 nm (about two dimer lengths), indicating that assembly occurs almost exclusively via single-subunit addition rather than via oligomers as was recently suggested. Finally, the assembly rate depends only weakly on load, with the average growth rate decreasing only 2-fold as the force increases 7-fold from 0.4 pN to 2.8 pN. CONCLUSIONS: The data are consistent with a mechanochemical model in which a spatially extended GTP cap allows substantial shortening on the nanoscale, while still preventing complete catastrophe in most cases.
BACKGROUND: The labile nature of microtubules is critical for establishing cellular morphology and motility, yet the molecular basis of assembly remains unclear. Here we use optical tweezers to track microtubule polymerization against microfabricated barriers, permitting unprecedented spatial resolution. RESULTS: We find that microtubules exhibit extensive nanometer-scale variability in growth rate and often undergo shortening excursions, in some cases exceeding five tubulin layers, during periods of overall net growth. This result indicates that the guanosine triphosphate (GTP) cap does not exist as a single layer as previously proposed. We also find that length increments (over 100 ms time intervals, n = 16,762) are small, 0.81 +/- 6.60 nm (mean +/- standard deviation), and very rarely exceed 16 nm (about two dimer lengths), indicating that assembly occurs almost exclusively via single-subunit addition rather than via oligomers as was recently suggested. Finally, the assembly rate depends only weakly on load, with the average growth rate decreasing only 2-fold as the force increases 7-fold from 0.4 pN to 2.8 pN. CONCLUSIONS: The data are consistent with a mechanochemical model in which a spatially extended GTP cap allows substantial shortening on the nanoscale, while still preventing complete catastrophe in most cases.
Authors: R A Walker; E T O'Brien; N K Pryer; M F Soboeiro; W A Voter; H P Erickson; E D Salmon Journal: J Cell Biol Date: 1988-10 Impact factor: 10.539
Authors: Pavel Zakharov; Nikita Gudimchuk; Vladimir Voevodin; Alexander Tikhonravov; Fazoil I Ataullakhanov; Ekaterina L Grishchuk Journal: Biophys J Date: 2015-12-15 Impact factor: 4.033
Authors: Nicolas Fernandez; Qiang Chang; Daniel W Buster; David J Sharp; Ao Ma Journal: Proc Natl Acad Sci U S A Date: 2009-04-28 Impact factor: 11.205
Authors: Ram Dixit; Brian Barnett; Jacob E Lazarus; Mariko Tokito; Yale E Goldman; Erika L F Holzbaur Journal: Proc Natl Acad Sci U S A Date: 2009-01-06 Impact factor: 11.205