Analysis of cyclin E co-expression genes reveals nuclear transcription factor Y subunit alpha is an oncogene in gastric cancer

Abstract

Objective: To explore genes potentially co-expressed with cyclin E in gastric cancer and discover possible targets for gastric cancer treatment.

Methods: The Cancer Genome Atlas (TCGA) stomach adenocarcinoma sequencing data were used to predict genes co-expressed with cyclin E. Co-expression genes predicted by cBioPortal online analysis with Pearson correlation coefficient ≥0.4 were analyzed by gene ontology (GO) enrichment annotation using the PANTHER online platform (Ver. 7). Interactions between proteins encoded by these genes were analyzed using the STRING online platform (Ver. 10.5) and Cytoscape software (Ver. 3.5.1). Genes displaying a high degree of connection were analyzed by transcription factor enrichment prediction using FunRich software (Ver. 3). The significant transcription factor and cyclin E expression levels and their impact on gastric cancer progression were analyzed by Western blotting and Kaplan-Meier survival curve analysis.

Results: After filtering the co-expression gene prediction results, 78 predicted genes that included 73 protein coding genes and 5 non-coding genes with Pearson correlation coefficient ≥0.4 were selected. The expressions of the genes were considered to be correlated with cyclin E expression. Among the 78 genes co-expressed with cyclin E, 19 genes at the central of the regulatory network associated with cyclin E were discovered. Nuclear transcription factor Y subunit alpha (NF-YA) was identified as a significant transcription factor associated with cyclin E co-expressing genes. Analysis of specimen donors' clinical records revealed that high expression of NF-YA tended to be associated with increased cyclin E expression. The expression of both was associated with progression of gastric cancer. Western blotting results showed that compared with normal tissues, NF-YA and cyclin E were highly expressed in tumor tissues (P < 0.001). Survival curve analysis clearly demonstrated relatively poor overall survival of gastric cancer patients with high cyclin E or high NF-YA expression level, compared to patients with low cyclin E or NF-YA expression (P < 0.05).

Conclusions: NF-YA may promote gastric cancer progression by increasing the transcription of cyclin E and other cell cycle regulatory genes. NF-YA might be a potential therapeutically useful prognostic factor for gastric cancer.

Keywords: Cyclin E; Gastric cancer; Nuclear transcription factor Y subunit alpha; Oncogene.

Fig. 1

Gene Ontology (GO) enrichment analysis of 78 predicted cyclin E co-expressing genes. GO terms of cellular components (red), molecular functions (green), and biological processes (blue) are plotted in different colors.

Fig. 2

Predicted protein–protein interaction network within the 78 predicted cyclin E co-expressing genes.

Fig. 3

Transcription factor enrichment analysis of 19 predicted cyclin E co-expressing genes. Percentage of genes enriched for each transcription factor is plotted on the left axis and -log₁₀P on the right axis. The dotted horizontal line represents the P = 0.05 threshold.

Fig. 4

Western blotting analysis of cyclin E and NF-YA protein expression levels in 22 pairs of gastric cancer specimens (CA) and their non-cancerous counterparts (NC). (A) Representative results of cyclin E and NF-YA protein expression levels in CA and NC samples from one patient. (B) Statistical analysis of cyclin E and NF-YA expression in 22 pairs of CA and NC samples. The gray scale analysis of each band was performed using ImageJ software, and gray scale of each band was normalized to the mean value of that in NC group. P value was calculated automatically. ^aCompared with NC, P < 0.001. (C) Correlation analysis of cyclin E and NF-YA protein expression based on Western blotting results.

Fig. 5

Kaplan–Meier survival estimates. (A) Kaplan–Meier curve analysis of overall survival of patients with different cyclin E expression levels, P = 0.0417. (B) Kaplan–Meier curve analysis of overall survival of patients with different NF-YA expression levels, P = 0.0325.