Mini-Review |
Address correspondence to Dr. Anthony Brown, The Ohio State University, Neurobiotechnology Center, Rightmire Hall, 1060 Carmack Road, Columbus, OH 43210. Tel.: (614) 292-1205. Fax: (614) 292-5379. E-mail: brown.2302{at}osu.edu
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Abstract |
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Introduction |
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Fast and slow axonal transport |
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Most of what we know about the composition and kinetics of axonal transport comes from studies on laboratory animals using radioisotopic pulse labeling (Grafstein and Forman, 1980). Several decades of work using this experimental paradigm has demonstrated that proteins and other molecules are transported along axons in association with distinct membranous and nonmembranous cargo structures that move at different rates (Tytell et al., 1981). Membranous organelles move most rapidly, in the fast components of axonal transport, whereas cytoskeletal polymers and cytosolic protein complexes move more slowly, in the slow components. The difference in the rate of fast and slow axonal transport has long been assumed to indicate that membranous and nonmembranous cargoes move by fundamentally distinct mechanisms, but direct observations on the movement of these cargoes in living cells now indicate that they are all transported by fast motors and that the principle difference between fast and slow transport is not the rate of movement per se, but rather the manner in which the movement is regulated.
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Movement of membranous organelles |
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Direct observations on the movement of membranous organelles in living axons indicate that many organelles move in a continuous and unidirectional manner at instantaneous rates that are comparable to the maximal rate of fast axonal transport determined by radioisotopic pulse labeling (Table I) and also to the maximal rates reported for microtubule motors in vitro (e.g., Woehlke and Schliwa, 2000). These observations indicate that many membranous organelles move along axons in a highly efficient manner, pausing only infrequently during their journey. One way to express this efficiency is in terms of the duty ratio. In the context of intracellular transport, the duty ratio is the proportion of time that a cargo structure spends actually moving. Thus, we can say that the axonal transport of many membranous organelles is characterized by a high duty ratio.
One exception to the high duty ratio of membranous organelles are mitochondria (Hollenbeck, 1996, Fig. 1). These organelles move less rapidly than most Golgi-derived and endocytic vesicles, forming a kinetically distinct component of axonal transport (Table I). The maximal rate of mitochondrial movement is generally quoted to be 2070 mm/d, which is comparable to the instantaneous rates observed in living cells, but the average rate appears to be considerably slower (Lorenz and Willard, 1978; Grafstein and Forman, 1980). The explanation for this transport behavior appears to be that mitochondria move in an intermittent and bidirectional manner, with differences in the balance of anterograde and retrograde movements and pauses giving rise to a broad range of overall rates (Morris and Hollenbeck, 1993; Ligon and Steward, 2000). Although some axonal mitochondria may move rapidly and continuously for long distances, resulting in the maximal rate of movement, most exhibit frequent pauses and reversals, resulting in much slower rates of movement (Blaker et al., 1981). Though not extensively studied, it appears that endoplasmic reticulum may also be transported along axons in this manner (Ellisman and Lindsey, 1983).
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Movement of cytoskeletal polymers |
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Recently the movement of neurofilaments and microtubules has been observed in axons, and these observations indicate that the solution to the slow axonal transport controversy is disarmingly simple. Cytoskeletal polymers do move in axons, but their movements are not slow after all (Fig. 1). Both neurofilaments and microtubules move at fast rates, approaching the rate of movement of membranous organelles, but the average rate of movement is slow because the movements are both infrequent and bidirectional (Roy et al., 2000; Wang et al., 2000; Wang and Brown, 2001). Remarkably, the key to observing this movement was simply a matter of experimental design; previous studies that did not detect movement were designed with the explicit expectation of a slow and synchronous movement, and it now appears that they were probably not capable of detecting the rapid and asynchronous movement (Wang and Brown, 2001). Thus, the overall speed and direction of neurofilament and microtubule movement is a temporal summation of anterograde and retrograde movements and pauses, perhaps not fundamentally dissimilar from the behavior of mitochondria in axons described above. As is the case for mitochondria, the slow overall rate of movement of neurofilaments and microtubules suggests that these structures move with a low duty ratio, spending most of their time not moving. For example, it has been estimated that neurofilaments in mature axons (such as those used for radioisotopic pulse labeling studies) may spend as much as 99% of their time pausing during their journey along the axon (Brown, 2000). It seems likely that microfilaments may exhibit a similar behavior, but the movement of these cytoskeletal polymers has not yet been observed.
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Cytoskeletal polymers as carrier structures |
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The idea that cytoskeletal polymers are carrier structures for slow axonal transport was initially met with skepticism, but it is now clear that cytoskeletal polymers do move in axons and that many of the cytosolic proteins that are conveyed by slow axonal transport can bind, directly or indirectly, to cytoskeletal polymers. For example, tau protein and spectrin both move in slow component a along with neurofilament proteins and tubulin. Spectrin is known to interact with neurofilaments (Macioce et al., 1999), and tau is a well characterized microtubule-associated protein. Thus, it is possible that neurofilaments and microtubules could be carrier structures for slow component a of axonal transport. A prediction of this hypothesis is that other proteins that move in this rate component, which have yet to be identified, will also be found to bind to these cytoskeletal polymers. By the same logic, it is also possible that microfilaments could be carrier structures for slow component b. In support of this hypothesis, microfilaments are known to interact with a wide range of different cytosolic proteins, including many that are not traditionally thought of as cytoskeleton-associated proteins (e.g., Knull and Walsh, 1992). However, it is unlikely that the several hundred different proteins that move in slow component b all bind directly to microfilaments. More probably, many of these proteins form functional complexes that in turn associate with the moving filaments. It is also possible that some cytosolic protein complexes may move by binding directly to motor proteins. The identification and characterization of these various protein complexes is likely to provide fundamental insights into the supramolecular interactions that organize the cytosolic compartment of cytoplasm, not just in axons, but in all eukaryotic cells.
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Beyond fast and slow: a unified perspective |
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According to this unified perspective, membranous and nonmembranous cargoes are all transported along axons by the same underlying mechanism but they move at different rates due to differences in their duty ratio. Membranous organelles on the secretory and endocytic pathways, which function primarily to deliver membrane and protein components to sites along the axon and at the axon tip, move rapidly in a unidirectional manner, pausing for only brief periods of time. The high duty ratio of these organelles ensures that they are delivered rapidly to their destination. In contrast, cytoskeletal polymers, mitochondria, and possibly also endoplasmic reticulum, move in an intermittent and bidirectional manner, pausing more often and for longer periods of time, and sometimes reversing during their journey along the axon. Although we refer to these structures as cargoes, they are not simply the luggage of intracellular transport; these organelles and macromolecular assemblies are preassembled functional units that fulfill their architectural, physiological, and metabolic roles in the axon during their transit. For these cargoes, the journey is perhaps more important than the ultimate destination, and this may explain their unique motile behavior.
Based on these considerations, a central question underlying the difference between fast and slow axonal transport is the mechanism by which the movement of membranous and nonmembranous cargoes is regulated. For example, what determines whether a particular cytoskeletal polymer or membranous organelle moves or pauses, or how frequently it does so? And when movement does occur, what determines its direction and duration? Since the motile behavior of axonally transported cargoes determines the efficiency with which they are transported and the manner in which they are distributed along the axon, the regulation of this behavior is likely to be critical for many aspects of axonal structure and function. For example, in the case of mitochondria, the balance of anterograde and retrograde movements and pauses is regulated during axon growth in order to recruit these organelles to sites of metabolic demand (Morris and Hollenbeck, 1993). Likewise, in the case of neurofilaments and microtubules, the balance of anterograde and retrograde movements and pauses is likely to be the principal determinant of their steady-state distribution along the axon, and thus the regulation of the axonal transport of these structures is probably essential for local and long-range remodeling of the neuronal cytoskeleton during axon growth and maturation. Since axonal transport continues throughout the life of the neuron, it is likely that active regulation of the movement of its membranous and nonmembranous components is an ongoing process as fundamental to the biology of axons as metabolism itself.
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Acknowledgments |
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The work in the author's laboratory is supported by the National Institute of Neurological Disorders and Stroke.
Submitted: 2 December 2002
Revised: 28 January 2003
Accepted: 29 January 2003
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References |
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