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Two fragments of HBV DNA integrated into chrX: 11009033 and its genetic regulation in HepG2.2.15.
Molecular Medicine Reports 2023 May
Hepatitis B virus (HBV) integration into human genome causes hepatocellular carcinoma (HCC). The present study used inverse nested PCR; the full sequence of HBV DNA fragments of the chrX: 111009033 integration site was detected (987 bp), containing two fragments of double‑stranded linear DNA with the same orientation (1,744‑1,094 and 1,565‑1,228 nt). By reverse transcription‑quantitative PCR, HBV‑cell fusion transcript was observed in HepG2.2.15 cells. The mean copy number of this site in cells with H2 O2 treatment (8.73x10‑2 ±1.65x10‑2 copies/cell) was significantly higher than that in the cells without H2 O2 treatment (3.02x10‑2 ±2.33x10‑2 copies/cell; P<0.0001). The mean levels of P21‑activated kinase 3 (PAK3) were 15.67±5.65 copies/cell in HepG2.2.15 cells with H2 O2 treatment, significantly higher than in the cells without H2 O2 treatment (11.34±4.58 copies/cell, P=0.0076) and in HepG2 cells (5.92±1.54 copies/cell, P<0.0001). Significant difference of PAK3 levels was also found between HepG2.2.15 cells without H2 O2 treatment and HepG2 cells (11.34±4.58 vs. 5.92±1.54 copies/cell, P<0.0001). The average copy numbers of the integration site chrX: 111009033 were positively correlated with the average levels of PAK3 (P=0.0013). The overall trend of PAK3 expression was significantly increased in HepG2.2.15 cells with H2 O2 treatment compared with that in HepG2.2.15 cells without H2 O2 treatment (37.63±8.16 and 31.38±7.94, P=0.008) and HepG2 cells (21.67±7.88, P<0.0001). In summary, the chrX: 11009033 integration site may originate from primary human hepatocytes, occurrence and clonal expansion of which may upregulate PAK3 expression, which may contribute to hepatocarcinogenesis.
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