The correct targeting and localization of transmembrane proteins is an essential aspect of cellular organization, particularly in large cells such as muscle which contains distinct membrane domains. The Golgi complex is the subcellular organelle responsible for this task. In muscle, the organization of the Golgi complex changes dramatically during differentiation and regeneration. Very little is known of the mechanism of these changes. The organization of the Golgi complex of muscle also depends on the pattern of contractile activity and this regulation is not understood at all. Our goal is to understand how the distribution of the Golgi complex in muscle is regulated and is linked to the functional needs of muscle. In the past we have uncoverered basic aspects of the changes that take place during differentiation. We have established that the Golgi complex of each myoblast fragments into hundreds of smaller independent Golgi complexes placed around the nuclei and throughout the cytoplasm of the multinucleated myotubes and muscle fibers. These small Golgi elements are not distributed randomly but are retained next to endoplasmic reticulum sites specialized in the export of proteins to the Golgi complex. This specific localization suggested that the fragmentation of the Golgi complex during muscle differentiation resembles the fragmentation that the Golgi complex undergoes when microtubules are depolymerized. We have indeed demonstrated the similarity between these processes, therefore identifying changes in the microtubule cytoskeleton as a key factor in the changes that take place during differentiation. Patterned activity is an important regulator of muscle metabolism and contraction but it had never been linked to the organization of the protein secretory pathway. We have discovered that the distribution of Golgi complex, endoplasmic reticulum exit sites and microtubules is plastic in mature muscle fibers and responds to the pattern of contractile activity, causing a fiber type-dependent organization. We hypothesize that this plasticity is important and allows muscle to fulfill different metabolic demands depending on patterned activity. Our past work gives, for the first time, a description of the organization of the Golgi complex in muscle. In order to understand how it is controlled by factors such as patterned activity, we need to determine which of the several vesicular and cytoskeletal elements involved is the trigger that organizes the others. For example, it is fundamental to determine whether microtubule changes cause the reorganization of the endoplasmic reticulum exit sites during differentiation or whether the two take place independently. A large effort has been geared towards the observation of endoplasmic reticulum exit sites in live cells as a function of microtubule status. This has been done by the simultaneous observation of fluorescent constructs of two different colors in the C2 mouse muscle cell line, together with treatment with pharmacological agents that disrupt microtubules. The results are consistent, so far, with a model in which microtubules organize the ER exit sites by acting on the ER itself. All our results, so far, thus point to the primary importance of microtubules as regulators of the Golgi complex distribution in muscle. It becomes then important to understand how microtubules themselves are organized. An investigation of the changes in microtubule nucleation during differentiation is showing that nucleation, in differentiated muscle, takes place at three types of sites which differ from the myoblast centrosomes, although all seem to involve the protein g-tubulin. Therefore, new modes of recruitment of g-tubulin from a cytoplasmic pool may be responsible for the changes in microtubule organization during differentiation. Microtubules are essential integrators and regulators of subcellular architecture. Our results suggest that they may be play another important role as sensors of patterned contractile activity.
