ULTRAVIOLET MICROBEAM IRRADIATIONS OF SPINDLE FIBERS IN CRANE-FLY SPERMATOCYTES AND NEWT EPITHELIAL-CELLS - RESOLUTION OF PREVIOUSLY CONFLICTING OBSERVATIONS

In order to resolve apparent differences in reported experiments. we d irectly compared the effects of ultraviolet (UV) microbeam irradiation s on the behaviour of spindle fibres in newt epithelial cells and cran e-fly spermatocytes, using the same apparatus for both cell types. Thi s work represents the first time that irradiated crane-fly spermatocyt es have been followed using a high-NA objective and video-enhancement of images. In both cell types, irradiation of a kinetochore fibre in m etaphase produced an area of reduced birefringence (ARE), known to be devoid of spindle microtubules (MTs). Subsequently the kinetochore-war d edge of the ARB moved poleward with average velocities of 0.5 mu m/m in (n = 20) in spermatocytes and 1.1 mu m/min (n = 6) in epithelial ce lls. The poleward edge of the ARE rapidly disappeared when viewed usin g a x100. high-NA objective but generally remained visible when viewed with a x32, low-NA objective; this difference suggests that MTs polew ard from the ARE disperse vertically out of the narrow depth of field of the x100 objective but that many remain encompassed by that of the x32 objective. The primary difference in response between the two cell types was in the behaviour of the spindle poles after an ARE formed. In spermatocytes the spindle maintained its original length whereas in epithelial cells the pole on the irradiated side very soon moved towa rds the chromosomes, after which the other pole did the same and a muc h shortened functional metaphase spindle was formed.