The pp32 (ANP32A) gene acts as a tumor suppressor while its closely related 
homologue pp32r1 (ANP32C) is oncogenic and is overexpressed in breast, prostate 
and pancreatic tumors. The transduction of p53wt cell lines (ACHN and HeLa) with 
pp32r1 or pp32r1Y140H lentivirus increased the proliferation of p53wt cell lines 
compared to the untransduced control cells while transduction of the p53(R248W) 
MiaPaCa2 cell line had no effect. Cell cycle analysis of transduced ACHN cells 
by PI staining and BrdU incorporation illustrated a pronounced shift toward the 
S-phase of the cell cycle in cells overexpressing the pp32r1 and pp32r1Y140H 
proteins. Confocal microscopy and western blotting demonstrated that pp32r1 and 
the pp32r1Y140H mutant protein reside predominantly in the cytoplasm in 
constrast to pp32 which is a nuclear/cytoplasmic shuttling protein. To determine 
the effects of pp32r1 or pp32r1Y140H overexpression at the proteomic level we 
performed a comprehensive proteome analysis on ACHN, ACHN-pp32r1 and 
ACHN-pp32r1Y140H cell lysates using the isotope-coded protein label (ICPL) 
method. Among those proteins with >40% regulation were Macrophage Capping 
protein (CAPG) and Chromodomain Helicase DNA binding protein 4 (CHD4) proteins 
which were significantly upregulated by pp32r1 and pp32r1Y140H overexpression. 
This increase in CHD4 also appears to influence a number of cell cycle regulator 
genes including; p53, p21 and cyclinD1 as judged by western blotting. Silencing 
of CHD4 in ACHN-pp32r1Y140H cells using specific shRNA reverted the cell cycle 
dysregulation caused by pp32r1Y140H expression to that of the untransduced ACHN 
cell line, suggesting that CHD4 is the prominent effector of the 
pp32r1/pp32r1Y140H phenotype.