Small-interfering RNA-mediated silencing of the MAPK p42 gene induces dual effects in HeLa cells

Abstract

Abstract. The genesis and progression of cervical cancer involve the mutation or deviant expression of numerous genes, including the activation of oncogenes (Ha-ras, C-myc, C-erbB2 and Bcl-2) and inactivation of tumor-suppressor genes (p53 and Rb). Previous studies showed that small-interfering RNAs (siRNAs) targeting the MAPK p42 gene partly inhibit proliferation and increase apoptosis in human cervical carcinoma HeLa cells. Results of a microarray analysis showed that MAPK p42 siRNA inhibited cell growth through the regulation of cell cycle control and apoptosis and induced interferon-like response in HeLa cells. In order to confirm the dual effects of MAPK p42 siRNA, we compared the roles of siRNA and U0126, an inhibitor of MAPK p42, in HeLa cells. Short 21-mer double-stranded/siRNAs were synthesized to target MAPK p42 mRNA in HeLa cells. The siRNAs were transfected into HeLa cells using Lipofectamine. The cells were treated with siRNA or U0126 at different concentrations for a period of 48 h. The biological effect of siRNA and U0126 on HeLa cells was measured by MTT and flow cytometry. MAPK1, NUP188, P38, STAT1, STAT2, PML and OAS1 were analyzed by real-time quantitative PCR. HeLa cell growth was inhibited by siRNA or U0126, and the effect of siRNA inhibition was greater than that of U0126. Cell cycle phases were different for siRNA or U0126, but HeLa cell growth was arrested at the S phase by siRNA and at G1 phase by U0126. A down-regulation in MAPK p42 expression by siRNA and up-regulation by U0126 were noted. The results of real-time quantitative PCR showed that P38 was up-regulated and NUP188 was down-regulated by siRNA in comparison with the control groups, and the results were consistent with those of U0126. Expression levels of STAT1, STAT2, PML and OAS1 induced by siRNA differed from those induced by U0126. siRNA-mediated silencing and deactivation induced by U0126 in MAPK p42 led to growth inhibition in the HeLa cells. The effects of siRNA on HeLa cell growth were different from those of U0126. Dual effects of MAPK p42 siRNA-2 on HeLa cell growth were noted: one consisted of a specific effect induced by siRNA-mediated p42 MAPK silencing and the other exhibited a non-specific interferon-like response.

Phosphorylation of Ser45 and Ser59 of αB‐crystallin and p38/extracellular regulated kinase activity determine αB‐crystallin‐mediated protection of rat brain astrocytes from C2‐ceramide‐ and staurosporine‐induced cell death

“…Thus, non-specific effects of MAPK siRNA affecting the expression of other MAPKs are possible. For example, it has been reported that siRNA of p42-MAPK reduced the expression of p42 and simultaneously up-regulated the expression of p38 (Yuan et al 2010). Thus, specific inhibitors of MAPK are helpful tools for exploring the involvement of MAPK, albeit with some limitations.…”

“…Thus, non‐specific effects of MAPK siRNA affecting the expression of other MAPKs are possible. For example, it has been reported that siRNA of p42‐MAPK reduced the expression of p42 and simultaneously up‐regulated the expression of p38 (Yuan et al. 2010).…”

Role of the tumour protein P53 gene in human cervical squamous carcinoma cells: Discussing haematopoietic cell‑specific protein 1‑associated protein X‑1‑induced survival, migration and proliferation

“…Understanding the complex process of the regulation of a variety of signalling networks in the development and progression of cervical cancer is of great importance. As one of the key genes promoting apoptosis, tumour protein P53 (p53) serves an important role in controlling the cell cycle, inhibiting tumour formation, maintaining cell genome integrity and responding to various cellular stresses (9)(10)(11)(12). In recent years, accumulating evidence has demonstrated that the p53 protein may translocate from the cytoplasm to the mitochondria and activate mitochondria-related apoptotic pathways.…”