Multiple signaling pathways and mechanisms regulate p53 activity. transcriptional activity, thus suppressing p53-mediated regulation of the expression of genes involved in the cell cycle (p21, cyclin Nonivamide D1) and apoptosis (Bax, Bcl-2, and PUMA). Collectively, our results, both in vivo and in vitro, indicate that the expression of MAGEA3 contributes to CC cell proliferation and tumor growth and exerts tumor-promoting effects by regulating the KAP1/p53 signaling pathway. Keywords: Melanoma-associated antigen A3 (MAGEA3), p53, KRAB domain-associated protein 1 (KAP1), cervical cancer, cell cycle, apoptosis Introduction As a common malignant tumor, cervical cancer ranks as the fourth leading cause of cancer-related death worldwide, and it remains one of the major causes of cancer-related death in women worldwide . Although cervical cancer treatment options, including radiotherapy, chemotherapy, and surgery, have made great progress, the overall 5-year survival rate of patients with cervical cancer remains unfavorable because of recurrence and metastasis. Therefore, the introduction of new treatment and diagnosis strategies must reduce recurrence and enhance the survival rate. Melanoma-associated antigen A3 (MAGEA3) gene can be a cancer-testis antigen (CTA) gene whose manifestation has been proven in several malignancies, including melanoma, breasts, colorectal, gastric, lung and pancreatic tumor [2-8]. Growing data possess reported that aberrant manifestation of MAGEA3 in lots of tumor types offers been proven to correlate with poor medical outcome [8-11]. Like a cancer-testis antigen, MAGEA3 is indicated in testes and tumor, making it a perfect candidate for tumor immunotherapy provided its potential to focus on particular tumor cell types without influencing normal cells [12,13]. To research whether MAGEA3 can be mixed up in development and tumorigenesis of human being cervical tumor, we previously recognized the manifestation of MAGEA3 by quantitative RT-PCR and immunohistochemical strategies Nonivamide in cervical lesion cells compared with regular tissues. The outcomes showed how the manifestation of MAGEA3 in cervical tumor (CC) cells was significantly greater than that of the standard group. Moreover, the manifestation degree of MAGEA3 was favorably correlated with the medical stage, pathological grade, and lymphatic metastasis of cervical cancer . Based on this, we speculated that MAGEA3 plays a critical role during the development and progression of cervical cancer. To verify our speculation, we performed the present study to determine whether MAGEA3 regulates the proliferation and apoptosis of CC cells. Recently, convincing evidence points to the capability of MAGE-A proteins to control the p53 tumor suppressor and regulate essential Rabbit Polyclonal to MRPL21 pathways associated with cell proliferation . It has been shown that MAGEA3 is involved in the inhibition of apoptosis via p53-dependent suppression of Bax and preservation of surviving . Another study indicated that Knockdown of MAGEA3 caused a reduction in proliferation in gastric cancer cells by regulating the cell cycle and apoptosis-related genes (p21, Bax) . Thus, to investigate the underlying mechanism of MAGEA3 in carcinogenesis, we Nonivamide further explored the regulatory relationship between MAGEA3 and the P53 signaling pathway in cervical cancer. Materials and methods Cell culture Human cervical cancer cell lines (HeLa, SiHa, C33A, and Caski) were purchased from the Cell Resource Center, Institute of Basic Medical Science (IBMS, Beijing, China). The cells were cultured in RPMI-1640 (GIBCO, NY, USA) medium supplemented with 10% fetal bovine serum (GIBCO), 100 U/ml penicillin and 100 g/ml streptomycin (HyClone, UT, USA) and were incubated at 37C with 5% CO2. The culture medium was replaced with fresh medium every 1-2 days. Cells passaging was performed when the cell cultures became 80-90% confluent. Lentivirus transfection The lentivirus for overexpressing MAGEA3 (Lv-MAGEA3) and the negative control lentivirus (Lv-NC) were obtained commercially from GenePharma (Shanghai, China). SiHa cells were seeded into a 6-well plate at 70-80% confluence. Cells were transfected with Lv-MAGEA3 or Lv-NC at an MOI (multiplicity of infection) of 5 and divided into three groups: blank control group (blank, untransfected cells), negative control group (Lv-NC, transfected with negative control lentivirus), and MAGEA3-overexpressing group (Lv-MAGEA3, transfected with lentivirus for overexpressing MAGEA3). Transfection was performed according to the manufacturers instructions. Cells were transfected 48-72 h prior to being harvested for use in subsequent Nonivamide analysis. siRNA and plasmid transfection MAGE-A3 knockdown was performed with siRNA in HeLa.