RNA interference of MAD2 and BUBR1 genes causes mitotic spindle alterations, aneuploidy and cell cycle arrest p53-dependent.

The Spindle Assembly Checkpoint (SAC) is a cellular surveillance mechanism that functions to ensure faithful chromosome segregation during mitosis. Failure of this checkpoint or alteration in expression of SAC proteins (MAD2, BUB1, BUBR1 and others) can result in aneuploidy, a state of having abnormal numbers of chromosomes. MAD2 haploinsufficiency resulted in aneuploidy in MEFs and colon cancer cells in culture. Thus, spindle checkpoint components might have additional functions not-checkpoint-related functions that when disrupted contribute to tumorigenesis. Here we investigated the effects of MAD2 or BUBR1 transcriptional silencing in HCT-116 cells. Transient reduction of MAD2 (40%) and BubR1 (70%) affected both cell survival and growth, in comparison to MAD2 and BubR1 alone silenced cells. Real time RT-PCR analysis showed p21 and p53 overexpression that might account for the G1/S arrest observed in MAD2/BubR1 co-depleted cells. After MAD2/BubR1 posttrascriptional silencing cells showed spindle alterations and the presence of lagging chromosomes revealed by time-lapse video microscopy . These findings suggest that co-depletion of MAD2 and BubR1 could affect the correct assembly of the mitotic spindle causing aneuploidy and chromosomal alterations, underlining an involvment of the p53/p21 pathway in the block of aneuploid cells.