The roles of cell cycle and BRCA1 in the DNA damage response

Exploiting


Introduction
In cancer stem cells (CSCs), epithelial-mesenchymal transition (EMT) networks play an important role to acquisition of the drug resistance and cancer malignant feature [1]. To reveal the network pathways in EMT and CSCs, gene expression in diffuse-and intestinal-type gastric cancer (GC) have been analyzed. Our previous findings identified several molecular networks and the related microRNAs (miRNAs) in intestinal-and diffuse-type GC [2][3][4][5]. Cell proliferation and regulation of the cell cycle are essential in cancer therapeutic targeting. In this article, we focus on the roles of cell cycle and BRCA1 in the DNA damage response in diffuse-and intestinal-type GC. Cell cycle regulation may play an important role in intestinal-and diffuse-type GC. The mechanism of cancer drug resistance would be highlighted by the involvement of cell cycle in EMT and CSCs.

RefSeq data analysis
The RefSeq data of diffuse-type and intestinal-GC are publicly available in The Cancer Genome Atlas (TCGA) of The cBioPortal for Cancer Genomics database [6][7][8] in NCI Genomic Data Commons (GDC) Data Portal [9]. From the data of stomach adenocarcinoma (TCGA, PanCancer Atlas), intestinal-and diffuse-type GC data, which are noted as chromosomal instability (CIN) and genomically stable (GS), respectively, in TCGA Research Network publication, were compared [8].

Molecular genome network analysis
Data of intestinal-and diffuse-type GC in TCGA cBioPortal Cancer Genomics were uploaded and analyzed through the use of Ingenuity Pathway Analysis (IPA) (QIAGEN Inc., Hilden, Germany) [10].

Data Visualization
The results of gene expression data of RefSeq and network analysis were visualized by Tableau software.

Canonical pathways altered in diffuse-and intestinal-type GC
Canonical pathways altered in diffuse-and intestinal-type GC are shown in Figure 1 and Table  1. The 2815 IDs which are significantly different in diffuse-and intestinal-type GC were analyzed in network analysis, which identified 69 canonical pathways related to the diffuse-and intestinal-type GC. Gene expression data of the diffuse-and intestinal-type GC revealed 36 canonical pathways with activation z-score as shown in Table 1    Estrogen-mediated S-phase Entry -3.5 1.5

Cell Cycle: G1/S checkpoint Regulation pathway was activated in diffuse-type GC
Molecule activity predictor in IPA predicted the activation of Cell Cycle: G1/S Checkpoint Regulation pathway in diffuse-type GC ( Figure 2). In Cell Cycle: G1/S Checkpoint Regulation pathway, DNA damage induces p53, which is expected to be activated in diffuse-type GC. Analysis of direct relationships of miRNAs and targeted molecules in Cell Cycle: G1/S Checkpoint Regulation pathway revealed the relationships between miRNAs and the targeted molecules ( Figure 2c, Table  2).

Cyclins and Cell Cycle Regulation pathway was activated in intestinal-type GC
Molecule activity predictor in IPA predicted the activation of Cyclins and Cell Cycle Regulation pathway in intestinal-type GC (Figure 3). Analysis of direct relationships of miRNAs and targeted molecules in Cyclins and Cell Cycle Regulation pathway revealed the relationships between miRNAs and the targeted molecules (Figure 3c, Table 3).

Role of BRCA1 in DNA Damage Response pathway was activated in intestinal-type GC
Molecule activity predictor in IPA predicted the activation of Role of BRCA1 in DNA Damage Response pathway in intestinal-type GC (Figure 4). Role of BRCA1 in DNA Damage Response pathway was identified as the most significant canonical pathway with p value of 6.6 x 10 -12 . Gene expression of BRCA1 which is associated with G1/S transition has increased in intestinal-type GC. BRCA1 codes a 190kD nuclear phosphorylation protein that maintains genomic stability and functions as a tumor suppressor. It is interesting that p53 and c21CIP1 are activate in intestinal-type GC in the Role of BRCA1 in DNA Damage Response pathway. BRCA1 may be involved in the activation of p53. Analysis of direct relationships of miRNAs and targeted molecules in Role of BRCA1 in DNA Damage Response pathway revealed the relationships between miRNAs and the targeted molecules (Figure 4c, Table 4). Ten miRNAs which have direct relationships between BRCA1 in Role of BRCA1 in DNA Damage Response pathway included miR-125a-3p (miRNAs w/seed CAGGUGA), miR-146a-5p (and other miRNAs w/seed GAGAACU), miR-224-5p (miRNAs w/seed AAGUCAC), miR-3615 (miRNAs w/seed CUCUCGG), miR-4639-3p (and other miRNAs w/seed CACUCUC), miR-5586-3p (miRNAs w/seed AGAGUGA), miR-6516-5p (miRNAs w/seed UUGCAGU), miR-6814-5p (miRNAs w/seed CCCAAGG), miR-6875-3p (miRNAs w/seed UUCUUCC), miR-99a-3p (and other miRNAs w/seed AAGCUCG) (Figure 4d, Table 5).     Figure 5). Analysis of direct relationships of miRNAs and targeted molecules in Cell Cycle: G2/M DNA Damage Checkpoint Regulation pathway revealed the relationships between miRNAs and the targeted molecules (Figure 5c, Table 6).

Conclusion
The several canonical pathways have been found to be altered in diffuse-and intestinal-type GC. Canonical pathway on Cell Cycle: G1/S Checkpoint Regulation was activated in diffuse-type GC, and Cyclins and Cell Cycle Regulation was activated in intestinal-type GC. Canonical pathway related to Role of BRCA1 in DNA Damage Response was activated in intestinal-type GC, where BRCA1 which is related to G1/S phase transition was up-regulated. Cell cycle regulation may be altered in EMT condition in diffuse-type GC.