MITOTIC DISASSEMBLY OF THE GOLGI-APPARATUS IN-VIVO

Populations enriched in prophase cells were obtained either by using a cell line with a temperature-sensitive mutation in the mitotic kinase , p34(cdc2), or by treating cells with olomoucine, an inhibitor of thi s kinase, Both methods resulted in efficient and reversible block of t he cells at the G(2)/M boundary, After cells were released from the ce ll cycle block, the morphological changes to the Golgi apparatus were characterised using both quantitative conventional electron microscopy and immune-gold microscopy, The early mitotic phases were divided int o six stages (G(2) to pro-metaphase) based on the morphology of the nu cleus. During prophase the cross-sectional length of Golgi stacks decr eased prior to unstacking, At the same time, small vesicular profiles, typically 50-70 nm in diameter, accumulated in the vicinity of the st acks. The disappearance of Golgi stacks was accompanied by the transie nt appearance of tubular networks, By the time cells entered prometaph ase, the stacks had completely disassembled and only clusters consisti ng of Golgi vesicles and short tubular elements were left. When cells were released from the G(2)/M boundary and pulsed briefly with [AlF4]( -) to prevent uncoating of transport vesicles, vesicular profiles with a morphology reminiscent of COP-coated vesicles appeared, These vesic ular profiles were either associated with Golgi stacks or, at later st ages, with clusters, but were formed at all stages of disassembly. Tog ether these results provide further support for our model that continu ed budding of vesicles from the rims of Golgi cisternae is at least pa rtly responsible for the disassembly of the Golgi apparatus.