Muscle fibers contain as many as 10,000 nuclei bounded by a single cell membrane, in contrast to most cells of the body which contain only one nucleus and only one copy of the genetic blueprint for the cell. During development, many individual muscle precursor cells fuse together to form each large muscle fiber. A basic problem arises when one considers that muscle fibers are separated into three, largely discrete, functional domains. Most of the length of each fiber contains contractile machinery, but the ends of the fibers are specialized for attachment to tendons while the very small synaptic region in the middle is specialized for attachment to tendons while the very small synaptic region in the middle is specialized for response to neurotransmission. To what extent can nuclei that have fused into one part of a muscle fiber contribute to a specialized function that occurs in another part of the same muscle fiber? The answer to this question is essential to an understanding of the regulation and organization of muscle fibers. The proposed research addresses the nuclear contribution to the small but essential post-synaptic specialization where the muscle receives instruction from the motor neuron. Cells of a muscle cell line in tissue culture can be induced to fuse into multi-nucleated myotubes which spontaneously contract and express elements of the post-synaptic specialization even in the absence of neurons. When muscle cells of different genetic origin are co-cultured, hybrid myotubes form which contain nuclei from both sources. A computerized microscope can then be used to track the different nuclei and the molecules that they contribute to the post-synaptic specialization. With this approach, it will be possible to decide whether an identified nucleus is always associated with a post-synaptic domain or whether it is free to migrate within the myotube, thereby expanding its zone of influence. Careful mapping of the post-synaptic molecules on the myotube surface will also provide an understanding of how fixed and mobile components of the post-synaptic specialization interact.
