Microtubule treadmilling in vivo V.I.Rodionov and G.G. Borisy Science, vol. 275: 215-218, 1997.

In vivo, cytoplasmic microtubules are nucleated and anchored by their minus ends at the centrosome and are believed to turn over by a mechanism termed dynamic instability,--depolymerization and repolymerization at their plus ends. In cytoplasmic fragments of fish melanophores, microtubules were shown to detach from their nucleation site and depolymerize from their minus ends. Free microtubules moved toward the periphery by treadmilling,--growth at one end and shortening from the opposite end. Frequent release from nucleation sites may be a general property of centrosomes and permit a minus end mechanism of microtubule turnover and microtubule treadmilling.

	Movie 1 (1100k) - Fish melanophores translocate pigment granules to the center (aggregation) or to the periphery (redispersion) along a radial array of microtubules (MTs). Remarkably, melanophore fragments retained the capacity to aggregate pigment and organize a radial MT array of correct polarity orientation (minus ends at the center) in the apparent absence of the centrosome. To study MT dynamics, we fluorescently tagged MTs in melanophores by microinjection of labeled tubulin, microsurgically dissected fragments from the parental cells, and induced pigment aggregation and formation of the MT aster. Images of labeled MTs were then sequentially acquired in the living fragments at short time intervals (3 s) for extended periods (10 min). The video sequence illustrates characteristic patterns of MT behavior in melanophore fragments. Some of the MTs remain static with one end at the pigment aggregate and the other at the plasma membrane. Others grow to the periphery from the aggregate which apparently has the capacity to nucleate MTs, or release from the aggregate and depolymerize from their proximal ends. MTs show only short excursions at their ends as if dynamic instability is suppressed. In addition to growing and shortening, short MT of constant length move away from the aggregate.
	Movie 2 (875k) - Movement of MTs could be achieved either by transport (MT dependent motors bound to a cytoplasmic matrix) or by treadmilling (polymerization at the plus end and depolymerization at the minus end). To definitively distinguish between transport and treadmilling mechanisms, a reference mark was placed on translocating MTs by photobleaching a narrow zone with a laser microbeam. If MTs were transported, the zone of bleaching would move with the MT so that the distances between the zone and the ends of the MT would remain constant. Alternatively, if MTs treadmilled, the zone would remain stationary but the distance from the zone to the leading end would increase and the distance to the trailing end would decrease. The movie shows that when a moving MT was irradiated with a laser microbeam, its leading end advanced away from the photobleached zone whereas its trailing end approached the zone, ultimately crossing it and appearing on the other side which demonstrated that MTs moved by a treadmilling mechanism.
	Movie 3 (974k) - This movie shows another example of bleaching experiment (see the legend to Fig. 2).
	Movie 4 (2500k) - This movie shows another example of bleaching experiment (see the legend to Fig. 2).