Regulation of DNA Polymerase POLD4 Influences Genomic Instability in Lung Cancer

Abstract

Genomic instability is an important factor in cancer susceptibility, but a mechanistic understanding of how it arises remains unclear. We examined hypothesized contributions of the replicative DNA polymerase δ (pol δ) subunit POLD4 to the generation of genomic instability in lung cancer. In examinations of 158 lung cancers and 5 mixtures of 10 normal lungs, cell cycle-and checkpoint-related genes generally showed mRNA expression increases in cancer, whereas POLD4 showed reduced mRNA in small cell lung cancer (SCLC). A fraction of non-small cell lung cancer patients also showed low expression comparable with that in SCLC, which was associated with poor prognosis. The lung cancer cell line ACC-LC-48 was found to have low POLD4 expression, with higher histone H3K9 methylation and lower acetylation in the POLD4 promoter, as compared with the A549 cell line with high POLD4 expression. In the absence of POLD4, pol δ exhibited impaired in vitro DNA synthesis activity. Augmenting POLD4 expression in cells where it was attenuated altered the sensitivity to the chemical carcinogen 4-nitroquinoline-1-oxide. Conversely, siRNA-mediated reduction of POLD4 in cells with abundant expression resulted in a cell cycle delay, checkpoint activation, and an elevated frequency of chromosomal gap/break formation. Overexpression of an engineered POLD4 carrying silent mutations at the siRNA target site rescued these phenotypes, firmly establishing the role of POLD4 in these effects. Furthermore, POLD4 overexpression reduced intrinsically high induction of γ-H2AX, a well-accepted marker of double-stranded DNA breaks. Together, our findings suggest that reduced expression of POLD4 plays a role in genomic instability in lung cancer. Cancer Res; 70(21); 8407-16. ©2010 AACR.

The tail that wags the dog: p12, the smallest subunit of DNA polymerase δ, is degraded by ubiquitin ligases in response to DNA damage and during cell cycle progression

“…siRNA depletion of p12 in cultured cells led to cell cycle delay, checkpoint activation, and an increase in chromosomal gap/break formation. 63 Additional studies using shRNA depletion of p12 revealed an increase in formation of karyomere-like cells.…”

4-Nitroquinoline-1-oxide effects human lung adenocarcinoma A549 cells by regulating the expression of POLD4

“…DNA polymerase δ (Pol δ) is essential for DNA replication, which consists of four subunits, which are p125, p50, p68 and p12. The expression of the smallest DNA Pol δ subunit, POLD4, was low in certain non-small cell lung cancers and small cell lung cancers in our previous study, and the lack of POLD4 protein expression or downregulation of its expression by siRNA weakened DNA replication and the NER capability (9).…”

“…In previous studies, POLD4 expression in small cell lung cancer and a small section of non-small cell lung cancers is lower when compared to normal samples (9). Clinical data showed that almost all small cell lung cancers have a smoking history, indicating the possibility of a correlation between smoking-induced lung cancer and POLD4.…”

“…In our previous study (9), the low expression of POLD4 in small cell and non-small cell lung cancer suggested that there is a possible association between smoking and POLD4 genes. In the present study, 4NQO decreased the POLD4 protein expression in the lung cancer cell line A549, and the effect could be reversed by the proteasome inhibitor MG132.…”

“…In our previous study, we proved that regulation of DNA polymerase POLD4 influenced genomic instability in lung cancer, and found that downregulation of POLD4 in Calu6 cells results in G 1 -S blockage through suppression of the AKT-Skp2-p27 pathway (9,16,17).…”