Immunofluorescence and electron microscopic investigations of the effects of dithiopyr on onion root tips

Abstract

Light, immunofluorescence, and electron microscopy are used to investigate the effects of dithiopyr [ S,S-dimethyl 2-(difluoromethyl)-4-(2-methylpropyl)-6-(trifluoromethyl)-3,5-pyridine-dicarbothioate] on onion root meristems. At high herbicide concentrations (10 −4, 10 −5 M) all of the dividing cells are in an arrested prometaphase configuration although, at lower concentrations (10 −6, 10 −7 M), multipolar mitosis, along with some apparently normal mitotic figures, also are noted. Immunofluorescence microscopy reveals that nearly all of the microtubules are absent after treatment with high concentrations of herbicide, although small tufts of microtubules remain at the kinetochore. These kinetochore tufts are confirmed to be microtubules (and not just aggregates of tubulin protein) by electron microscopic analysis. Phragmoplast arrays are absent at these higher concentrations and no cell plates are formed. Nuclei that reform after the arrested prometaphase or multipolar division are irregularly formed, either highly lobed or fragmented into multiple nuclei. Branched and undulating phragmoplast microtubule arrays oriented in abnormal directions are observed frequently after 10 −6 M treatment. The walls that result from these abnormal phragmoplasts are also unusual. Branched walls, multiple walls/cell, walls with electron opaque depositions, and incomplete reticulate walls are also noted. Because dithiopyr does not bind tubulin, we propose that this herbicide destabilizes microtubules as they are formed so that only short microtubules result. These shortened, destabilized microtubules are ineffective in chromosome movement during mitosis or other cellular functions involving microtubules. Thus, the net effects of dithiopyr are similar to those compounds that cause complete microtubule loss, such as trifluralin or amiprophosmethyl, even though the mechanism of disruption is different.