Methylation of histone H3K9 is a well-conserved epigenetic marker for transcriptional silencing (14). Histone H3K9 methylation and DNA methylation can work together on the establishment and maintenance of heterochromatin (15). The methyltransferases responsible for histone H3K9 methylation are able to catalyze different substrates and lead to various results (16). These include Clr4/SUV39H1 (17), SUV39H2 (18), G9a (19), GLP/Eu-HMTase1 (20), ESET/SETDB1 (21), Riz1/PRDM2 (22) and CLLD8/KMT1F (23).

Among these HKMTs, G9a is critical for gene silencing and embryo development. Currently, aberrant regulation of G9a has been identified in a number of cancers (Fig. 3), and is involved in the control of cancer metabolism by maintaining the serine-glycine biosynthetic pathway (24). In PC3 prostate cancer cells, knockdown of G9a significantly inhibits cell growth and induces cellular senescence, and higher G9a expression is associated with poorer prognosis in cancer patients (25). In addition, knockdown of G9a promotes E-cadherin expression in claudin-low breast cancer (CLBC), and inhibited cell migration and invasion in CLBC and lung cancer (26,27). In ovarian cancer, higher G9a expression predicts a greater mortality of patients (28). In neuroblastoma, our previous study reported the importance of G9a in regulating the autophagy signaling pathway, knockdown of G9a inhibited cell growth and proliferation, and the activation of autophagy occurred (29). In acute myeloid squamous cell carcinoma and pancreatic adenocarcinoma, inhibition of G9a induced autophagy-related cell death (30,31). In glioma cancer, previous studies have identified that G9a-dependent H3K9me2 repressed cluster of differentiation 133 and Sox2 expression and in leukemia, and loss of G9a markedly delayed tumor progression and repressed ATRA-mediated leukemia cell differentiation (32,33). Additionally, in head and neck cancer, G9a inhibited stem cell self-renewal (34). G9a was also upregulated in hepatocellular carcinoma (HCC) tissues, and cooperated with the H3K9 methylation effector protein CDYLb, which is involved in HCC development (35,36). In oesophageal squamous cell carcinoma, G9a may serve as an effective prognostic factor and be used as a biomarker (37).

Another key enzyme is SETDB1 (Fig. 3), which has been reported in numerous types of human cancer. SETDB1 was identified in 1999, and the activity of histone H3-K9-specific methyltransferase was reported in 2002 (38,39). SETDB1 is recruited by various transcription factors to regulate gene expression, and is associated with the H3K9me3-enriched genome regions (40). As a constitutive member of promyelocytic leukemia-nuclear bodies, SETDB1 has been linked to numerous cellular processes, such as apoptosis, DNA damage responses and transcriptional regulation (41). In melanoma, a study in zebrafish regarded SETDB1 as an oncogene, and indicated its role in regulating tumorigenesis (42). SETDB1 was upregulated in cell lines and tissues in a number of human carcinomas, for example, non-small and small lung cancer, glioma and prostate cancer. In non-small and small lung cancer, recent studies have shown that knockdown of SETDB1 reduced lung cancer cell growth in vitro and in vivo, and overexpression of SETDB1 promoted cancer cell invasiveness (43–46). In glioma, suppression of SETDB1 by siRNA significantly reduced cell proliferation (47). In prostate cancer, downregulation of SETDB1 by siRNA inhibited PCa cell proliferation, migration and invasion (48). Another study reported that a microRNA, known as miR-7, directly targeted SETDB1, and inhibited breast cancer stem cell (CSC) invasion and metastasis, and decreased the breast CSC population, which suggested that SETDB1 activity is important in breast cancer (49).

Methylation of histone H3K27 is correlated with transcriptional repression (50). Enhancer of EZH2, as a catalytic component of the polycomb repressive complex 2, catalyzes histone H3K27 tri-methylation. To date, >300 studies have reported a close correlation between EZH2 and 46 types of human cancer (Fig. 4). EZH2 is commonly overexpressed in the majority of common cancers, and high EZH2 expression is a prognostic indicator of poor survival. In breast cancer, downregulation of EZH2 blocks the cell cycle, and suppresses cell growth and survival (51–54). In prostate cancer, knockdown of EZH2 inhibits cell proliferation and invasion (55–57). In glioma stem cells, EZH2 is a known target of the MELK-FOXM1 complex, having a critical role in promoting resistance to radiation (58). In glioma and clear cell renal cell carcinoma, downregulation of EZH2 expression can reduce cell proliferation and increase cell apoptosis (59,60). Furthermore, in non small-cell lung cancer, EZH2 silencing alters the cell cycle by inducing G₂/M arrest (61). In lymphoma, overexpression of EZH2 promotes the proliferation and aggressiveness of neoplastic cells, facilitates malignant tumor cell diffusion and mediated transcriptional silencing (62). Additionally, knockdown of EZH2 induces RUNX expression to further inhibit cell proliferation in gastric, breast, prostate, colon and pancreatic cancer cells (63). Furthermore, EZH2 is important in other types of cancer, including malignant peripheral nerve sheath tumor (64), medulloblastoma (65) and lung adenocarcinoma (66).

Methylation of H3K4, as an epigenetic phenomenon conserved from yeast to human, is extremely important for transcriptional initiation. It recruits proteins for transactivation, and has reverse functionally to H3K9 methylation. As a common marker of activated genes, H3K4 tri-methylation provides an epigenetic signature of active enhancers (67). H3K4 methyltransferases include the mixed lineage leukemia (MLL) family (68), SET1A/B (69), absent, small or homeotic disks 1-like (ASH1L) (70), ASH2L (71), SET and MYND domain-containing protein 3 (SMYD3) (72), SET7/9 (73), and SMYD1 (74). However, in mammals, ≥6 H3K4 methyltransferases, including Set1A and Set1B and MLLs 1–4, exhibit histone methyltransferase activity (68).

As cardiac- and muscle-specific histone methyltransferases, except for the regulation of early heart development, the oncogenic role of SMYD3 has been identified in different types of cancer. In colorectal cancer, HCC and esophageal squamous cell carcinoma, knockdown of SMYD3 impairs cell proliferation (75–77). Similarly to CRC and HCC, in breast cancer, silencing of SMYD3 also inhibits cell growth, and SMYD3 promotes breast carcinogenesis by directly regulating the protooncogene WNT10B (78). In cervical carcinoma cells and prostate cancer, reduction of SMYD3 expression by doxycycline or small hairpin RNA is able to significantly inhibit cell proliferation, colony formation and migration/invasion activity (79,80).

H3K36 methylation is an indicator of transcriptional elongation, and H3K36 methyltransferases contain nuclear receptor binding SET domain-containing protein 1 (NSD1), NSD2, NSD3 (81), SMYD2 and SETD2 (82). These are involved in diverse biological processes, including alternative splicing and transcriptional repression, as well as DNA repair and recombination (83).

The NSD family is known to be involved in multiple types of cancer, and knockdown of NSD members would suppress cell proliferation and tumor growth. NSD1 specifically mediates methyl transfer onto H3K36 and H4K20. In prostate cancer, NSD1 can enhance androgen receptor transactivation and is associated with prostate tumorigenesis (84,85). In neuroblastoma, overexpression of NSD1 induces tumor suppressor-like features, such as reduced colony formation density and inhibited cell growth (86). NSD1 has been reported in numerous types of cancer, including multiple myeloma (87), acute myeloid leukemia (88,89), lung cancer (90,91) and ganglioglioma (92). SETD2 is a novel tumor suppressor, which is responsible for H3K36me3 reduction, further resulting in tumor growth inhibition. Mutated SETD2 has been frequently identified in human leukemia, thymic carcinoma (93), renal cell carcinoma (82,94), non-small cell lung cancer (95) and pediatric glioma (96).

In general, H3K79 methylation correlates with gene transcription (97,98). Disrupter of telomeric silencing 1 (DOT1), the only known H3K79 methyltransferase, participates in the regulation of transcription, development, differentiation and proliferation of normal cells (99). However, the role of DOT1L in cancer cells remains to be elucidated. Dot1 has been shown to interact with multiple MLL fusion partners including AF9, 11–19-leukemia protein, AF10 and AF17. In clinically aggressive acute leukemia, it may be involved in cell survival pathways, and loss of Dot1 activity attenuates cell viability and the colony formation ability (100). However, in lung cancer, downregulation of DOT1L reduces cell proliferation and led to cell cycle arrest at the G₁ phase (101).

Methylation of H4K20 has been implicated in multiple biological processes, such as gene transcriptional regulation, cell cycle control, development and genomic integrity maintenance (102,103). Mono- and di-methylated H4K20 have been attributed to DNA replication, DNA damage repair and chromatin compaction. Lack of H4K20me1 results in chromosome condensation in the interphase nucleus. Tri-methylation of H4K20 is required for silencing heterochromatic regions (104).

H4K20 methyltransferases include SUV4-20H1 and SUV4-20H2 (104), PR-Set7/Set8/KMT5A (105), NSD1 (106) and NSD2/WHSC1/MMSET (107). SUV4-20H1 and SUV4-20H2 mediate H4K20me2 and H4K20me3, NSD2 mediates methyl transfer onto H3K4 and H4K20, and PR-Set7 is known as the sole enzyme that catalyzes H4K20me1 (108). In breast cancer cells, overexpression of SUV420H1 and SUV420H2 suppresses cell invasiveness, whereas knockdown of SUV420H2 activates normal mammary epithelial-cell invasion in vitro (109).