Downregulated PITX1 Modulated by MiR-19a-3p Promotes Cell Malignancy and Predicts a Poor Prognosis of Gastric Cancer by Affecting Transcriptionally Activated PDCD5.
BACKGROUND/AIMS
PITX1 has been identified as a potential tumor-suppressor gene in several malignant tumors. The molecular mechanism underlying PITX1, particularly its function as a transcription factor regulating gene expression during tumorigenesis, is still poorly understood.


METHODS
The expression level and location of PITX1 were determined by quantitative reverse transcription PCR (qRT-PCR) and immunohistochemical staining in gastric cancer (GC). The effect of PITX1 on the GC cell proliferation and tumorigenesis was analyzed in vitro and in vivo. To explore how PITX1 suppresses cell proliferation, we used PITX1-ChIP-sequencing to measure genome-wide binding sites of PITX1 and assessed global function associations based on its putative target genes. ChIP-PCR, electrophoretic mobility shift assay, and promoter reporter assays examined whether PITX1 bound to PDCD5 and regulated its expression. The function of PDCD5 in GC cell apoptosis was further examined in vitro and in vivo. The relationship between the PITX1 protein level and GC patient prognosis was evaluated by the Kaplan-Meier estimator. Meanwhile, the expression level of miR-19a-3p, which is related to PITX1, was also detected by luciferase reporter assay, qRT-PCR, and western blotting.


RESULTS
The expression level of PITX1 was decreased in GC tissues and cell lines. Elevated PITX1 expression significantly suppressed the cell proliferation of GC cells and tumorigenesis in vitro and in vivo. PITX1 knockdown blocked its inhibition of GC cell proliferation. PITX1 bound to whole genome-wide sites, with these targets enriched on genes with functions mainly related to cell growth and apoptosis. PITX1 bound to PDCD5, an apoptosis-related gene, during tumorigenesis, and cis-regulated PDCD5 expression. Increased PDCD5 expression in GC cells not only induced GC cell apoptosis, but also suppressed GC cell growth in vitro and in vivo. Moreover, PITX1 expression was regulated by miR-19a-3p. More importantly, a decreased level of PITX1 protein was correlated with poor GC patient prognosis.


CONCLUSION
Decreased expression of PITX1 predicts shorter overall survival in GC patients. As a transcriptional activator, PITX1 regulates apoptosis-related genes, including PDCD5, during gastric carcinogenesis. These data indicate PDCD5 to be a novel and feasible therapeutic target for GC.
['Qiao|Fengchang|F|', 'Gong|Pihai|P|', 'Song|Yunwei|Y|', 'Shen|Xiaohui|X|', 'Su|Xianwei|X|', 'Li|Yiping|Y|', 'Wu|Huazhang|H|', 'Zhao|Zhujiang|Z|', 'Fan|Hong|H|']
D000328:Adult D000368:Aged D000369:Aged, 80 and over D000818:Animals D017209:Apoptosis D051017:Apoptosis Regulatory Proteins D045744:Cell Line, Tumor D049109:Cell Proliferation D015536:Down-Regulation D005260:Female D005753:Gastric Mucosa D015972:Gene Expression Regulation, Neoplastic D006801:Humans D008297:Male D008819:Mice, Nude D035683:MicroRNAs D008875:Middle Aged D009363:Neoplasm Proteins D051761:Paired Box Transcription Factors D011379:Prognosis D013270:Stomach D013274:Stomach Neoplasms D015533:Transcriptional Activation
Pitx1 MiR-19a-3p PDCD5 Cell proliferation Cell apoptosis