Global DNA Hypomethylation following 5-aza-2'-deoxycytidine treatment induces aneuploidy in HCT-116 tumor cells.

Aneuploidy, the alteration of the normal number of chromosomes, is found in most of the human solid tumors and correlated with to defects in the process of chromosome segregation (1). It was also suggested that the alteration of the 5-methylcytosine (5-mC) pattern in the chromosome pericentromeric region, generated to aneuploid cells (2, 3). To investigate the relationship between hypomethylation and whole chromosome aneuploidy, we treated HCT-116 cells, a near diploid line, with the demethylating agent 5-aza- 2'-deoxycytidine (DAC). The treatment with DAC for 24, 48 and 72 hours produced a progressive reduction of DNA methylation as shown by decrease of 5-mC signal. DNA hypomethylation res…

p14ARF re-expression induces apoptosis in aneuploid HCT116 cells

Weakening the Spindle Assembly Checkpoint by reduced expression of its components such as MAD2, BubR1 and MPS1 induces chromosome instability and aneuploidy both hallmarks of cancer cells. p14ARF that is found frequently altered in human cancers, is overexpressed in response to oncogenic stimuli to stabilize p53 halting cell progression. Previously, we determined that lack or reduced expression of p14ARF is involved in the maintenance of aneuploid cells suggesting that it could be part of a pathway controlling proliferation of aneuploidy cells. To investigate further this aspect of p14ARF function it was ectopically expressed in HCT116 cells, a stable near diploid cell line, after MAD2 depl…

Proliferation of aneuploid cells induced by CENP-E depletion is counteracted by the p14ARF tumor suppressor

The spindle assembly checkpoint (SAC) is a cellular surveillance mechanism that ensures the fidelity of chromosomes segregation. Reduced expression of some of its components weakens the SAC and induces chromosome instability and aneuploidy, which are both well-known hallmarks of cancer cells. Centromere protein-E (CENP-E) is a crucial component of the SAC and its function is to facilitate kinetochore microtubule attachment required to achieve and maintain chromosome alignment. The present study investigates the possible role of p14ARF as a controller of aneuploid cells proliferation. We used RNA interference to induce aneuploidy by partial depletion of CENP-E in human primary fibroblasts (I…

p14(ARF) Prevents Proliferation of Aneuploid Cells by Inducing p53-Dependent Apoptosis.

Weakening the Spindle Assembly Checkpoint by reduced expression of its components induces chromosome instability and aneuploidy that are hallmarks of cancer cells. The tumor suppressor p14(ARF) is overexpressed in response to oncogenic stimuli to stabilize p53 halting cell progression. Previously, we found that lack or reduced expression of p14(ARF) is involved in the maintenance of aneuploid cells in primary human cells, suggesting that it could be part of a pathway controlling their proliferation. To investigate this aspect further, p14(ARF) was ectopically expressed in HCT116 cells after depletion of the Spindle Assembly Checkpoint MAD2 protein that was used as a trigger for aneuploidy. …

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

Most human tumors are characterized by aneuploidy that is believed to be the consequence of chromosomal instability (CIN). The mechanism(s) leading to aneuploidy and the pathways that allow its tolerance are not completely understood. The Spindle Assembly Checkpoint (SAC) is a cellular surveillance mechanism working during mitosis, and alterations of genes that encode components of the SAC weakening the mitotic checkpoint, induce aneuploidy by chromosome mis-segregation. We induced aneuploidy in near-diploid tumor cells by simultaneous depletion of the SAC proteins MAD2 and BUBR1 by RNA interference in the attempt to gain further insight on the cellular responses to aneuploidy. Individual r…

The tumor suppressor p14ARF hampers proliferation of aneuploid cells induced by CENP-E partial depletion

The Spindle Assembly Checkpoint (SAC) is a cellular surveillance mechanism that ensures faithfully segregation of chromosomes. Reduced expression of some of its components weakens the SAC and induces chromosome instability and aneuploidy, both hallmarks of tumor cells. Centromere Protein-E (CENP-E) is a crucial component of the SAC and facilitates kinetochore microtubule attachment required to achieve and maintain chromosome alignment. To investigate the possible role of p14ARF on aneuploid cells proliferation we induced aneuploidy in primary human fibroblasts (IMR90) and in near diploid tumor cells (HCT116) by partial depletion of CENP-E obtained by RNA interference. Our results show that …

p14ARFPrevents Proliferation of Aneuploid Cells by Inducing p53-Dependent Apoptosis

Weakening the Spindle Assembly Checkpoint by reduced expression of its components induces chromosome instability and aneuploidy that are hallmarks of cancer cells. The tumor suppressor p14ARF is overexpressed in response to oncogenic stimuli to stabilize p53 halting cell progression. Previously, we found that lack or reduced expression of p14ARF is involved in the maintenance of aneuploid cells in primary human cells, suggesting that it could be part of a pathway controlling their proliferation. To investigate this aspect further, p14ARF was ectopically expressed in HCT116 cells after depletion of the Spindle Assembly Checkpoint MAD2 protein that was used as a trigger for aneuploidy. p14ARF…

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