DNA Methyltransferase1 post-transcriptional silencing induces aneuploidy and cell cycle arrest in human cells.

The regulation of chromatin structure is a dynamic and complex process that is modulated by epigenetic mechanisms. Malfunctioning of these processes can cause gene expression alteration and could compromise important events such as chromosome condensation and segregation. Imbalance in cytosine methylation and deregulation of DNA-methyltransferases (DNMTs), and of DNMT1 in particular, is frequent in human cancers. To investigate DNMT1 implication in the generation of aneuploidy we evaluated the effects of its depletion by RNA-interference both in primary human cells (IMR90) and in near diploid human tumor (HCT116) cells. Posttranscriptional silencing of DNMT1 induced aneuploidy, cell prolife…

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 …

pRb loss and chromosomal instability in human cells.

pRb loss and chromosomal instability in human cells. Recent studies suggest that Retinoblastoma tumor suppressor (RB) plays important roles in the prevention of chromosomal instability by regulating genes that control cell cycle progression and mitotic events. We investigated the effects of stable post-transcriptional silencing of RB in primary human fibroblasts (IMR90) and in near-diploid colon cancer cells (HCT116) focusing on chromosome missegregation mechanisms. Stable depletion of pRb was achieved by infection with the retroviral vector MSCV-LMP670 encoding a microRNA (miR670) targeting RB transcript. Cytogenetic, immunofluorescence microscopy and time-lapse video-microscopy analyses s…

Simultaneous reduction of MAD2 and BUBR1 expression induces mitotic spindle alterations associated with p53 dependent cell cycle arrest and death