Bacterial toxins reportedly have specific cytotoxic effects on target cells including tumour cells, and they have received increasing attention in the development of anti-rumour drugs. LukS-PV is the S component of PVL secreted by S. aureus. We previously reported that LukS-PV induces apoptosis in AML cells mediated by C5aR and inhibit AML cell proliferation in vitro and in vivo and have no obvious side effects on mice[2, 25, 26, 37]. However, the precise mechanisms underlying histone modification regulation by LukS-PV exerts its anti-leukaemia effects in AML cells have remained poorly understood.
AML is complex disease with a diverse genetic landscape. Recent studies have revealed that epigenetic dysregulation plays an important role in leukaemia pathogenesis[6, 11]. Further histone-modifying enzymes reportedly contribute to leukaemia pathogenesis. For example, MLL1 is a member of the SET family of methyltransferases and has been reported affect stem cell differentiation in leukaemia patients[10]. SETD2 is a histone methyltransferase that regulates leukaemia pathogenesis[30, 39]. This phenomenon is precisely based on the reversibility of epigenetic modifications may facilitate targeted leukaemia therapy[12]. For example, azacitidine and decitabine are DNA methyltransferase inhibitors approved for clinically treating AML[8, 15]. EZH2 and LSD1 are reportedly involved in leukaemia pathogenesis, and these targeted drugs have entered clinical trials[14, 22]. In summary, histone modifications are potentially promising for targeted therapy for leukaemia.
SET8 is a member of the SET domain-containing methyltransferase family specifically targeting H4K20me1[18]. SET8 is involved in vital cellular processes, including transcriptional regulation[5, 20], S-phase progression[28], genome replication and stability[4, 16], and DNA repair[36]. Aberrant SET8 expression has been reported in numerous tumours. High SET8 levels are also associated with poor survival in cancer patients. Zhou et al reported that SET8 and H4K20me1 levels were higher in AML cells than in control cells, and that SET8 can regulate the Wnt signalling pathway and is associated with leukaemia pathogenesis[38]. Our observation showed that SET8 is downregluate targeted by LukS-PV revealed a mechanism underlying SET8 regulation, thus underscoring the importance of SET8 regulation in AML.
Previous studies have reported that SET8 is a specific monomethylation catalytic enzyme for H4K20me1[18]. Therefore, we further found that LukS-PV reduced H4K20me1 levels by downregulating SET8 in a dose- and time-dependent manner. To further assess the downstream mechanism of LukS-PV in exerting anti-leukaemia effects through the SET8-H4K20me1 axis, we performed ChIP-seq to identify H4K20me1 downstream target genes. Finally, a potential target gene PIK3CB was identified by classifying target genes, and we tested its effect on apoptosis through PIK3CB inhibitor GSK2636771. Together, our study indicate that LukS-PV induces cell apoptosis through downregulate expression of the target gene PIK3CB mediated by SET8 in AML cells.
The phosphatidylinositol 3-kinase (PI3K) pathway plays pivotal roles in cell growth, proliferation, and survival by integrating extracellular growth signals[3]. PI3K proteins fall into three classes, of which class IA are heterodimeric lipid kinases, consisting of a p85 regulatory subunit and a p110 catalytic subunit. The p110 catalytic subunits are p110α, β, and δ. PIK3CB (PI3-kinase p110β) is a member of the PI3K pathway, and hyperactivation of this pathway contributes to cancer progression in humans[9]. Akt, a serine/threonine protein kinase, is one of the best-characterized targets of the PI3K pathway. Recent studies have reported numerous Akt substrates including BAD, CREB, GSK-3β, mTOR, FOXO1, and PTEN[23]. Initiation of these signalling cascades regulates cellular proliferation, motility, and survival in cancer cells. The present ChIP-seq experiments and bioinformatics analysis revealed that PIK3CB is the transcriptional target gene for LukS-PV-mediated H4K20me1. Furthermore, our results show that LukS-PV decreases AKT phosphorylation and increases FOXO1 levels, thus inducing apoptosis by downregulating anti-apoptotic protein Bcl2 and upregulating pro-apoptotic protein Bax in HL-60 and NB-4 cells.
In conclusion, these results show that LukS-PV induces apoptosis in AML cells via the PI3K/AKT/FOXO1 signal transduction pathway by targeting SET8, implying that SET8 is a potential target for LukS-PV-mediated treatment of AML.