Arsenite-induced aneuploidy following short and long-term exposure in mammalian cells

Abstract

We studied the long-term progression of chromosomal instability in V79 cells treated acutely with arsenite (10mM, 24 hr) followed by growth in arsenic-free medium for 120 cell generations. Indirect immunostaining using anti-ß-tubulin antibody showed severe alterations in spindle morphology after only 6 h treatment and cytogenetic investigations carried out at the end of treatment revealed that the percentage of cells with 21 chromosomes (modal number of the cell line) decreased, making way for aneuploid cells. The acquired instability remained and propagated within the cell population. Moreover, we treated V79-derived G12 cells with sub-lethal doses (0.1-1.0 μM) of arsenite for 10 days followed by growth in arsenite-free medium for 40 cell generations. Cytogenetic analysis at the end of treatment showed concentration-dependent increase in aneuploid cells frequency that was even higher after 40 cell generations. In addition to this finding a large amount of cells in anaphase and cells with chromosomes showing premature centromere division was observed. Western blotting analysis showed dose-dependent upregulation of Mad2 that returns to normal after 40 cell generations and persistent aberrant expression of the spindle assembly complex proteins p-BubR1 and Cenp-E. Taken together, these results demonstrate that arsenic induces aneuploidy independently of mode of treatment; in particular, results from chronic exposure to sub-lethal doses raise the possibility that arsenic induces bypass of the spindle assembly checkpoints which may be mechanistically involved in the induction of cell transformation.