Regulation of DNA polymerase POLD4 influences genomic instability in lung cancer.
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.
Huang
Qin Miao
QM
Division of Molecular Carcinogenesis, Nagoya University Graduate School of Medicine, Aichi Cancer Center Research Institute, Nagoya, Japan.
Tomida
Shuta
S
Masuda
Yuji
Y
Arima
Chinatsu
C
Cao
Ke
K
Kasahara
Taka-Aki
TA
Osada
Hirotaka
H
Yatabe
Yasushi
Y
Akashi
Tomohiro
T
Kamiya
Kenji
K
Takahashi
Takashi
T
Suzuki
Motoshi
M
D015153:Blotting, Western; D002289:Carcinoma, Non-Small-Cell Lung / Q000201:enzymology / Q000235:genetics* / Q000473:pathology; D002453:Cell Cycle; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D047369:Chromatin Immunoprecipitation; D053903:DNA Breaks, Double-Stranded; D019175:DNA Methylation; D004258:DNA Polymerase III / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism; D004260:DNA Repair; D004261:DNA Replication; D015972:Gene Expression Regulation, Neoplastic; D042822:Genomic Instability; D006801:Humans; D047468:Immunoprecipitation; D008175:Lung Neoplasms / Q000201:enzymology / Q000235:genetics* / Q000473:pathology; D021122:Protein Subunits; D012333:RNA, Messenger / Q000235:genetics; D034741:RNA, Small Interfering / Q000494:pharmacology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D055752:Small Cell Lung Carcinoma / Q000201:enzymology / Q000235:genetics* / Q000473:pathology
D021122:Protein Subunits; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D004258:DNA Polymerase III