DYNAMIC MICROTUBULES UNDER THE RADIAL AND OUTER TANGENTIAL WALLS OF MICROINJECTED PEA EPIDERMAL-CELLS OBSERVED BY COMPUTER RECONSTRUCTION

By microinjecting rhodamine-conjugated pig brain tubulin into living p ea stem epidermal cells it has been possible to follow cortical microt ubules beneath the outer tangential wa II (OTW) as they re-orientate f rom a transverse to a longitudinal alignment. Earlier immunofluorescen ce studies on fixed material have shown that parallel cortical microtu bules circumnavigate the cell forming apparently continuous arrays whi ch are transverse, oblique or longitudinal to the cell's long axis. If the array re-orientates as a whole then microtubules along the radial walls would be expected to share the alignment of those on the tangen tial walls. There are, however, reports that microtubules beneath the outer tangential wall have a different orientation from microtubules a t the radial cell walls, raising important questions about the constru ction and behaviour of the array. Using computer-rotated stacks of opt ical sections collected by confocal scanning laser microscopy it has b een possible to display the microtubules along radial as well as tange ntial walls of the same microinjected cells. These observations demons trate for living epidermal cells that when microtubules are aligned lo ngitudinally at the outer epidermal wall they remain oblique or transv erse at the radial walls. The array may not therefore re-orientate as a whole but seems to undergo re-organization on only one cell face. Ho wever, despite the differing angles between the OTW and radial walls m icrotubules still form patterns which at the level of the confocal mic roscope are continuous from one cell face to another, around the cell. It is concluded that some organizing principle attempts to establish overall organization at the cellular level but that this can be pertur bed by local re-organization of dynamic microtubules in subcellular do mains. This study emphasizes the importance of the outer epidermal wal l and its associated cytoskeleton in initiating changes in the directi on of cell expansion.