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| © Neuroscience-Net Volume 1, Article #10002 | Received April 24, 1996 Accepted for Publication May 14, 1996 Published May 17, 1996 |
Dr. George C. Ruben
Department of Biological Sciences
Dartmouth College
Gilman 8
Hanover, NH 03755
(603) 646-2144
Fax: (603) 646-1347
E-mail: george.c.ruben@dartmouth.edu
There are no high resolution transmission electron microscope methods available that can reliably image the microtubule-associated proteins (MAPs) on the surface of microtubules, visualize lattice arrangements of MAPs or measure their associated filament diameters. We have bypassed a low resolution rotary replication method with ~4-5 nm resolution and implemented a vertical Pt-C replication method with a resolution of 0.7-1.0 nm. Previously we studied the MAP tau and found that it formed 2.1 nm triple-stranded left-hand helical polymers in purified tau preparations that had elastic properties (Ruben et. al., [1991] J. Biol. Chem., 266:22019-22027). This finding led to the suggestion that these polymers could associate axially with the microtubule wall protofilaments and could restore bent microtubules after axons crossing knee and elbow joints were flexed. The 2.1 nm tau polymer filaments have been documented in purified tau preparations but have never been seen in association with microtubules. We have now directly imaged the freeze-dried replicated microtubule surface in the presence and absence of MAPs. We present here our first findings using a new method for visualizing heterogeneous MAP preparations in association with taxol stabilized microtubules. These rat brain MAPs were neither exposed to acid pH nor heating, to ensure their assembly competence. Although we were unable to distinguish between MAP 1, MAP 2 or tau, it appears that very long (197-944 nm) MAP polymers (~2 nm diameter) can be assembled axially along the microtubule surface. The finding is consistent with the hypothesis that polymers of MAPs can associate with the microtubule surface. This study will be extended in the future with purified tau. Since the 2.1 nm tau polymer filaments have been found in Alzheimers neurofibrillary tangles (Ruben et. al, [1992] Brain Res., 590:164-179) and they may be precursor filaments of the paired helical filaments in tangles (Ruben et. al, [1993] Brain Res., 602:1-13), we hope to establish in the future a biological role for the triple-stranded left-hand helical 2.1 nm tau polymer filaments. Our imaging technique employed here will make it possible in future investigations to generally study the relationship of fibrous MAPs and motor proteins, dyneins and kinesins to microtubules. © 1996 Neuroscience-Net.
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