Apc and p53 interaction in DNA damage and genomic instability in hepatocytes.

Abstract

Disruption of Apc within hepatocytes activates Wnt signaling, perturb differentiation and ultimately lead to neoplasia. Apc negatively regulates Wnt signaling but is also involved in organizing the cytoskeleton and may play a role in chromosome segregation. In vitro studies have implicated Apc in the control of genomic stability. However, the relevance of this data has been questioned in vivo as Apc is lost earlier than the onset of genomic instability. Here, we analyse the relationship between immediate loss of Apc and the acquisition of genomic instability in hepatocytes. We used Cre-lox technology to inactivate Apc and in combination with p53 in vivo, to define the consequences of gene loss upon cell-cycle regulation, proliferation, death and aneuploidy. We show that whilst Apc loss leads to increased proliferation, it also leads to increased apoptosis, the accumulation of p53, p21 and markers of DSBs and DNA repair. Flow cytometry revealed an increased 4N DNA content, consistent with a G2 arrest. Levels of anaphase bridges were also elevated, implicating failed chromosome segregation. This was accompanied by an increase in centrosome number which demonstrates a role for Apc in maintaining euploidy.To address the role of p53 in these processes, we analyzed combined loss of Apc and p53, which led to a further increase in proliferation, cell death, DNA damages and repair and a bypass of G2 arrest than was observed with Apc loss. However we observed only a marginal effect on anaphase bridges and centrosome number which could be due to increased cell death. Our data therefore establishes, in an in vivo setting, that APC loss leads to a DNA damage signature and genomic instability in the liver and that additional loss of p53 leads to an increase in the DNA damage signal but not to an immediate increase in the genomic instability phenotype.