Untersuchungen zu den molekularen Mechanismen der Interaktionen zwischen Endoplasmatischem Reticulum und Mikrotubuli unter Verwendung von Vaccinia Virus-Replikations-Komplexen als Modellsystem

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Although microtubules (MTs) are known to play an important role in structural organization of the endoplasmic reticulum (ER) and ER-dynamics, relatively little is known about the molecular details underlying these interactions. The current proposal aims to use a novel in vitro system, based on mammalian cells, to study the molecular mechanisms of MT/ER interactions. It takes advantage of the observation that the cytoplasmic sites of vaccinia virus (vv) replication become entirely wrapped by the ER and these ER-enclosed sites move in a MT-dependent fashion towards the perinuclear region of the cell. Since the ER-enwrapped replication-sites can be isolated from infected cells, I propose to reconstitute their MT-interactions in vitro. Using a light microscopy assay, I will study first binding to, and then motility along MTs of isolated replication sites. With this assay I expect to gain important new insight into cellular as well as possibly viral factors underlying ER/MT interactions. Finally, the fact that the minus-end directed motility of the replication sites does not depend on the dynactin/dynein complex, suggests that a novel microtubule motor machinery needs to be identified.