Wenzhe Si

BACKGROUND: Metabolic reprogramming and epigenetic alterations contribute to the aggressiveness of pancreatic ductal adenocarcinoma (PDAC). Lactate-dependent histone modification is a new type of histone mark, which links glycolysis metabolite to the epigenetic process of lactylation. However, the role of histone lactylation in PDAC remains unclear. METHODS: The level of histone lactylation in PDAC was identified by western blot and immunohistochemistry, and its relationship with the overall survival was evaluated using a Kaplan-Meier survival plot. The participation of histone lactylation in the growth and progression of PDAC was confirmed through inhibition of histone lactylation by glycolysis inhibitors or lactate dehydrogenase A (LDHA) knockdown both in vitro and in vivo. The potential writers and erasers of histone lactylation in PDAC were identified by western blot and functional experiments. The potential target genes of H3K18 lactylation (H3K18la) were screened by CUT&Tag and RNA-seq analyses. The candidate target genes TTK protein kinase (TTK) and BUB1 mitotic checkpoint serine/threonine kinase B (BUB1B) were validated through ChIP-qPCR, RT-qPCR and western blot analyses. Next, the effects of these two genes in PDAC were confirmed by knockdown or overexpression. The interaction between TTK and LDHA was identified by Co-IP assay. RESULTS: Histone lactylation, especially H3K18la level was elevated in PDAC, and the high level of H3K18la was associated with poor prognosis. The suppression of glycolytic activity by different kinds of inhibitors or LDHA knockdown contributed to the anti-tumor effects of PDAC in vitro and in vivo. E1A binding protein p300 (P300) and histone deacetylase 2 were the potential writer and eraser of histone lactylation in PDAC cells, respectively. H3K18la was enriched at the promoters and activated the transcription of mitotic checkpoint regulators TTK and BUB1B. Interestingly, TTK and BUB1B could elevate the expression of P300 which in turn increased glycolysis. Moreover, TTK phosphorylated LDHA at tyrosine 239 (Y239) and activated LDHA, and subsequently upregulated lactate and H3K18la levels. CONCLUSIONS: The glycolysis-H3K18la-TTK/BUB1B positive feedback loop exacerbates dysfunction in PDAC. These findings delivered a new exploration and significant inter-relationship between lactate metabolic reprogramming and epigenetic regulation, which might pave the way toward novel lactylation treatment strategies in PDAC therapy.

Jumonji domain-containing 6 (JMJD6) is a member of the Jumonji C domain-containing family of proteins. Compared to other members of the family, the cellular activity of JMJD6 is still not clearly defined and its biological function is still largely unexplored. Here we report that JMJD6 is physically associated with the tumor suppressor p53. We demonstrated that JMJD6 acts as an α-ketoglutarate- and Fe(II)-dependent lysyl hydroxylase to catalyze p53 hydroxylation. We found that p53 indeed exists as a hydroxylated protein in vivo and that the hydroxylation occurs mainly on lysine 382 of p53. We showed that JMJD6 antagonizes p53 acetylation, promotes the association of p53 with its negative regulator MDMX, and represses transcriptional activity of p53. Depletion of JMJD6 enhances p53 transcriptional activity, arrests cells in the G1 phase, promotes cell apoptosis, and sensitizes cells to DNA damaging agent-induced cell death. Importantly, knockdown of JMJD6 represses p53-dependent colon cell proliferation and tumorigenesis in vivo, and significantly, the expression of JMJD6 is markedly up-regulated in various types of human cancer especially in colon cancer, and high nuclear JMJD6 protein is strongly correlated with aggressive clinical behaviors of colon adenocarcinomas. Our results reveal a novel posttranslational modification for p53 and support the pursuit of JMJD6 as a potential biomarker for colon cancer aggressiveness and a potential target for colon cancer intervention.

BACKGROUND: Melanoma, originated from melanocytes located on the basal membrane of the epithelial tissue, is the most aggressive form of skin cancer that accounts for 75% of skin cancer-related death. Although it is believed that BRAF mutation and the mitogen-activated protein kinase (MAPK) pathway play critical roles in the pathogenesis of melanoma, how the MAPK signaling is regulated in melanoma carcinogenesis is still not fully understood. METHODS: We characterized JMJD6 expression in melanoma tissue array by immunohistochemistry analysis. We used human melanoma A375, 451Lu and SK-MEL-1 cell lines for in vitro proliferation and invasion experiments, and xenograft transplanted mice using murine melanoma B16F10 cells by bioluminescence imaging for in vivo tumor growth and pulmonary metastasis assessments. Endothelial tube formation assay, chicken yolk sac membrane assay and matrigel plug assay were performed to test the effect of JMJD6 on the angiogenic potential in vitro and in vivo. RESULTS: Here we report that the jumonji C domain-containing demethylase/hydroxylase JMJD6 is markedly up-regulated in melanoma. We found that high expression of JMJD6 is closely correlated with advanced clinicopathologic stage, aggressiveness, and poor prognosis of melanoma. RNA-seq showed that knockdown of JMJD6 affects the alternative splicing of a panel of transcripts including that encoding for PAK1, a key component in MAPK signaling pathway. We demonstrated that JMJD6 enhances the MAPK signaling and promotes multiple cellular processes including melanogenesis, proliferation, invasion, and angiogenesis in melanoma cells. Interestingly, JMJD6 is transcriptionally activated by c-Jun, generating a feedforward loop to drive the development and progression of melanoma. CONCLUSIONS: Our results indicate that JMJD6 is critically involved in melanoma carcinogenesis, supporting the pursuit of JMJD6 as a potential biomarker for melanoma aggressiveness and a target for melanoma intervention.