Ultraviolet microbeam irradiation of chromosomal spindle fibres shears microtubules and permits study of the new free ends in vivo

Abstract

Irradiation of birefringent chromosomal spindle fibres in crane-fly spermatocytes in metaphase I or anaphase I produces an area of reduced birefringence (ARB) on the fibre. This ARB moves poleward and is lost at the pole. Ultrastructural and immunofluorescence analysis of ARBs obtained by irradiation with monochromatic ultraviolet light of wavelength 260 nm shows that the microtubules in the irradiated area are depolymerized, though the rest of the spindle appears unaffected. The area of microtubule depolymerization moves poleward with the ARB, and once the ARB reaches the pole the irradiated half-spindle appears normal. The motion of the ARB, therefore, appears to be due to the behaviour of the sheared microtubules in the chromosomal spindle fibre. The relative stability of the sheared microtubules shows that chromosomal fibre microtubules are not dynamically unstable, as are microtubules under certain conditions in vitro. However, ARB motion may be due to a moderated version of dynamic instability. Possible models for ARB motion are discussed.