The mission of our laboratory is to elucidate the cellular and molecular mechanisms by which the microtubule arrays of the neuron are established. Toward this end, we use a variety of techniques which include microscopic, biochemical, and molecular assays.
Microtubules
form the infrastructure of eucaryotic cells, acting as both architectural
elements and as railways for the transport of cytoplasmic constituents.
To serve these functions, microtubules must be organized into a wide variety
of configurations, ranging from the bipolar conformation of the mitotic
spindle to the dense paraxial arrays that occupy elongated cellular processes.
Thus an important question in cell biology is how different cell types organize
their microtubules into these various configurations. Traditionally, cell
biologists have focused on attachment to structures such as the centrosome
as the means by which microtubules are organized within the cytoplasm. The
centrosome nucleates a radial array of microtubules with their plus-ends
outward and their minus-ends inward. This simple mechanism can explain the
interphase microtubule arrays of many cell types, but it cannot explain
how microtubules are organized into more sophisticated patterns such as
those of the mitotic spindle or elongated cellular processes. Most efforts
to understand how these more complex patterns are established, while not
explicitly stated, have worked under the assumption that each cell type
invokes fundamentally different themes in order to establish its specialized
microtubule pattern. While this could be true, it seems more reasonable
that cells would work from a single blueprint, modifying it in order to
meet the specialized need of different cell types. If this theory is correct,
it is logical that the blueprint for organizing microtubules across cell
types would be the mitotic spindle, the most fundamental of all microtubule
arrays.
Neurons are terminally postmitotic cells that no longer organize their microtubules into mitotic spindles. Instead, the microtubules of the neuron are utilized for the formation of elongated processes termed axons and dendrites. These two types of processes contain dense arrays of paraxially-oriented microtubules, none of which are attached to the centrosome or any comparable structure. Despite this, the microtubules within these processes are highly organized with respect to their intrinsic polarity. In the axon, the microtubules are uniformly oriented with their plus-ends distal to the cell body of the neuron. In the dendrite, the microtubules are nonuniformly oriented with roughly similar numbers of microtubules of each orientation. These distinct microtubule patterns are essential for defining the cytoplasmic composition of each type of process, as well as for regulating morphological characteristics such as their relative lengths. In terms of structure and function, the microtubule arrays of the neuron would at first glance appear to be as different from the mitotic spindle as one could image. But appearances can be deceiving. Is it possible that the microtubule arrays of the postmitotic neuron are established using revisions of the same blueprint used to organize the mitotic spindle?
This issue could not have been addressed with any authority had it been raised a decade ago, when substantially less information was available on the means by which mitotic cells and postmitotic neurons organize their microtubules. However, the past several years have seen a revolution with regard to the mechanisms by which the mitotic spindle is organized, and recent work from our laboratory speaks to whether similar themes operate within the neuron. The major advancement in understanding the mitotic spindle came when it was realized that microtubules are not organized into a bipolar configuration simply as a result of their assembly and disassembly. It is now clear that, in addition to these dynamic events, force generation by ATP-driven molecular motor proteins is essential for the formation and functioning of the mitotic spindle. These forces move microtubules, after and during their assembly, into specific locations within the cell and into specific orientations. These forces are essential for separating the duplicated centrosomes in prophase, for the formation of the bipolar spindle in metaphase, and for the separation of the half-spindles in anaphase. Do analogous motor-driven forces exist in the postmitotic neuron, and are these forces utilized for establishing the axonal and dendritic microtubule arrays?
Recent studies from our laboratory strongly suggest that the forces generated by motor proteins are the key to understanding microtubule organization within the neuron, and that the specific motor proteins that generate these forces are the same motors that function during mitosis to organize microtubules into the bipolar spindle. Given the importance of the neuronal microtubule arrays, these motor-driven forces undoubtedly have broad implications. How does the neuron generate a single axon and multiple dendrites? How is one of the processes selected for axonal differentiation and the others for dendritic differentiation? What accounts for the unique morphological specializations of axons and dendrites? Why do axons grow indefinitely or until they reach an appropriate target tissue, whereas dendrites obtain a certain length and then stop growing? How are the distinct complements of cytoplasmic organelles and membrane constituents targeted for axons or dendrites? What accounts for the balance between plasticity and stability that permits the neuron to change in response to its environment? We suspect that the answers to these and other important questions are intimately related to the microtubule systems of the neuron. Thus we feel that elucidating how the neuronal microtubule arrays are established will provide broad insights into the most fundamental issues of cellular neuroscience.
