THE PERINUCLEAR CENTRIOLE-CONTAINING CENTROSOME IS NOT THE MAJOR MICROTUBULE-ORGANIZING CENTER IN SERTOLI CELLS

Microtubules are abundant in Sertoli cells and are predominantly arran ged parallel to the long axis of the cell, In addition, the centrioles occur in a perinuclear position in the basal one third of the cell, I n this study we investigate the importance of the perinuclear centriol e-containing centrosome as a microtubule organizing center (MTOC) in S ertoli cells. In all experiments, rat testes were perfusion fixed and then processed for electron and/or fluorescence microscopy, For fluore scence microscopy, fragments or dissected pieces of seminiferous epith elium were labeled for tubulin and actin, The three-dimensional patter n of microtubules in Sertoli cells was determined using data collected with a confocal microscope and analyzed using the NIH-Image program ( written by Wayne Rasband at the NIH), The detailed arrangement of micr otubules around, and the number of microtubule ends associated with, t he centrosome were determined from composite projections constructed f rom serial thin sections, The nucleating potential of the perinuclear centrosome was determined by perfusing testes for 6 h with 10 mu g/ml nocodazole and then for up to 3 h with control buffer prior to fixatio n and analysis with confocal and standard fluorescence microscopy, Mic rotubules are not organized around a focal perinuclear site and few mi crotubule ends are associated with the centrosome. Moreover, in cells recovering from nocodazole treatment, microtubules first appear in api cal (peripheral) processes, Our data indicate that the centriole-conta ining perinuclear centrosome is not a significant MTOC in Sertoli cell s. Rather, microtubules are nucleated in peripheral regions and projec t basally. Based on the observations that microtubules appear to 'cuff ' the nucleus, intermediate filaments are concentrated around the nucl eus, microtubules project into the perinuclear intermediate filament n etwork, and microtubules and intermediate filaments are often coaligne d, we suggest that microtubules are anchored into position at the base of the cell via linkages with the intermediate filament network, Our nucleation-anchorage model of microtubule organization in Sertoli cell s may be applicable to other epithelial systems.