Abstract
BackgroundModifications of the histone amino-terminal tails affect access of regulatory factors and complexes to chromatin and thereby influence biological processes. Cancer cells are characterized by prominent epigenetic dysregulation, including histone modifications. However, the functional roles of the histone methyltransferases (HMT) in cancer remain unclear.Methodology/Principal FindingsWe studied RNAi-based inhibition (knockdown, KD) of 2 different H3K9 HMTs, SUV39H1 and G9a. Knockdown of the 2 HMTs in PC3 cancer cell line markedly inhibited cell growth and caused profound morphological changes with loss of telomerase activity and shortened telomeres. SUV39H1 KD cells showed substantial increase in G2/M fraction. G9a KD cells showed increased DNA content (1.7-fold in 2 independent clones) compared with FACS analyses to control. Karyotype analyses showed that this was due to an increased number of chromosomes (from 61 to 102) in G9a KD cells compared to parental PC3. Intriguingly, we found abnormal centrosome morphology and number in about 25% of the G9a KD cells, while centrosomes were morphologically normal in control cells. Microarray analyses after KD of SUV39H1 or G9a showed very few genes up-regulated among the 39,000 genes. The silenced tumor-suppressor genes p16 and RASSF1A were not activated in KD cells.Conclusions/SignificanceThese data suggest that the 2 HMTs, SUV39H1 and G9a are required to perpetuate the malignant phenotype. Furthermore, G9a plays a critical role in regulating centrosome duplication presumably through chromatin structure rather than through affecting gene expression in cancer cells. Targeting these histone methyltransferases may be of therapeutic benefit in cancers.
Highlights
Chromatin assembly is a critical process related to DNA replication, gene expression and progression through the cell cycle
The G9a-KD cell nucleus showed selectively impaired broad euchromatic staining of 2MeH3K9 and contained only blurred speckles, which were overlapping with the pericentric region and centromeric region. 3MeH3K9 was detected on the pericentric region and centromeric region in control cells
G9a-KD did not significantly affect 3MeH3K9 patterns of the pericentric region and centromeric region. These findings were consistent with the previous G9a- deficient ES cell studies [19,20]. It appeared that G9a-KD affected H3K9me3 status at euchromatic region
Summary
Chromatin assembly is a critical process related to DNA replication, gene expression and progression through the cell cycle. G9a plays a critical role in regulating centrosome duplication presumably through chromatin structure rather than through affecting gene expression in cancer cells.
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