RESUMEN
Background: Hepatocellular carcinoma (HCC) is one of the most lethal human tumors with extensive intratumor heterogeneity (ITH). Serine protease 3 (PRSS3) is an indispensable member of the trypsin family and has been implicated in the pathogenesis of several malignancies, including HCC. However, the paradoxical effects of PRSS3 on carcinogenesis due to an unclear molecular basis impede the utilization of its biomarker potential. We hereby explored the contribution of PRSS3 transcripts to tumor functional heterogeneity by systematically dissecting the expression of four known splice variants of PRSS3 (PRSS3-SVs, V1~V4) and their functional relevance to HCC. Methods: The expression and DNA methylation of PRSS3 transcripts and their associated clinical relevance in HCC were analyzed using several publicly available datasets and validated using qPCR-based assays. Functional experiments were performed in gain- and loss-of-function cell models, in which PRSS3 transcript constructs were separately transfected after deleting PRSS3 expression by CRISPR/Cas9 editing. Results: PRSS3 was aberrantly differentially expressed toward bipolarity from very low (PRSS3Low ) to very high (PRSS3High ) expression across HCC cell lines and tissues. This was attributable to the disruption of PRSS3-SVs, in which PRSS3-V2 and/or PRSS3-V1 were dominant transcripts leading to PRSS3 expression, whereas PRSS3-V3 and -V4 were rarely or minimally expressed. The expression of PRSS3-V2 or -V1 was inversely associated with site-specific CpG methylation at the PRSS3 promoter region that distinguished HCC cells and tissues phenotypically between hypermethylated low-expression (mPRSS3-SVLow ) and hypomethylated high-expression (umPRSS3-SVHigh ) groups. PRSS3-SVs displayed distinct functions from oncogenic PRSS3-V2 to tumor-suppressive PRSS3-V1, -V3 or PRSS3-V4 in HCC cells. Clinically, aberrant expression of PRSS3-SVs was translated into divergent relevance in patients with HCC, in which significant epigenetic downregulation of PRSS3-V2 was seen in early HCC and was associated with favorable patient outcome. Conclusions: These results provide the first evidence for the transcriptional and functional characterization of PRSS3 transcripts in HCC. Aberrant expression of divergent PRSS3-SVs disrupted by site-specific CpG methylation may integrate the effects of oncogenic PRSS3-V2 and tumor-suppressive PRSS3-V1, resulting in the molecular diversity and functional plasticity of PRSS3 in HCC. Dysregulated expression of PRSS3-V2 by site-specific CpG methylation may have potential diagnostic value for patients with early HCC.
RESUMEN
PURPOSE: Metallothionein 2A (MT2A) suppresses the progression of human gastric cancer potentially through an "MT2A-NF-κB pathway" with unclear mechanisms. This study explored the role of a transcription factor, myeloid zinc-finger 1 (MZF1), in MT2A-NF-κB pathway and its clinical significance in gastric cancer. EXPERIMENTAL DESIGN: MZF1 expression and function in gastric cancer were investigated in vitro and in vivo. The relationship between MZF1 and MT2A was determined by gain-of-function and loss-of-function assays in gastric cancer cells and an immortalized gastric cell line GES-1. The prognostic value of MZF1 expression in association with MT2A was evaluated using IHC in two cohorts. RESULTS: MZF1 was epigenetically silenced in human gastric cancer cell lines and primary tumors. Overexpression of MZF1 in gastric cancer cells suppressed cell proliferation and migration, as well as the growth of xenograft tumors in nude mice. Knocking-down of MZF1 transformed GES-1 cells into a malignant phenotype characterized by increased cell growth and migration. Mechanistically, MZF1 was upregulated in both GC and GES-1 cells by MT2A ectopically expressed or induced upon treatment with a garlic-derived compound, diallyl trisulfide (DATS). MZF1 associated with MT2A was colocalized in the nuclei of GES-1 cells to target the promoter of NF-κB inhibitor alpha (NFKBIA). Clinically, MT2A and MZF1 were progressively downregulated in clinical specimens undergoing gastric malignant transformation. Downregulation of MT2A and MZF1 was significantly correlated with poorer patient prognosis. CONCLUSIONS: MT2A exerts its anti-gastric cancer effects by complexing with MZF1 to target NFKBIA. MT2A/MZF1 may serve as a valuable prognostic marker and a novel therapeutic target for human gastric cancer.
