Knocking Down Nucleolin Expression Enhances the Radiosensitivity of Non-Small Cell Lung Cancer by Influencing DNA-PKcs Activity.
Nucleolin (C23) is an important anti-apoptotic protein that is ubiquitously expressed in exponentially growing eukaryotic cells. In order to understand the impact of C23 in radiation therapy, we attempted to investigate the relationship of C23 expression with the radiosensitivity of human non-small cell lung cancer (NSCLC) cells. We investigated the role of C23 in activating the catalytic subunit of DNA-dependent protein kinase (DNA- PKcs), which is a critical protein for DNA double-strand breaks (DSBs) repair. As a result, we found that the expression of C23 was negatively correlated with the radiosensitivity of NSCLC cell lines. In vitro clonogenic survival assays revealed that C23 knockdown increased the radiosensitivity of a human lung adenocarcinoma cell line, potentially through the promotion of radiation-induced apoptosis and adjusting the cell cycle to a more radiosensitive stage. Immunofluorescence data revealed an increasing quantity of γ-H2AX foci and decreasing radiation-induced DNA damage repair following knockdown of C23. To further clarify the mechanism of C23 in DNA DSBs repair, we detected the expression of DNA-PKcs and C23 proteins in NSCLC cell lines. C23 might participate in DNA DSBs repair for the reason that the expression of DNA-PKcs decreased at 30, 60, 120 and 360 minutes after irradiation in C23 knockdown cells. Especially, the activity of DNA-PKcs phosphorylation sites at the S2056 and T2609 was significantly suppressed. Therefore we concluded that C23 knockdown can inhibit DNA-PKcs phosphorylation activity at the S2056 and T2609 sites, thus reducing the radiation damage repair and increasing the radiosensitivity of NSCLC cells. Taken together, the inhibition of C23 expression was shown to increase the radiosensitivity of NSCLC cells, as implied by the relevance to the notably decreased DNA-PKcs phosphorylation activity at the S2056 and T2609 clusters. Further research on targeted C23 treatment may promote effectiveness of radiotherapy and provide new targets for NSCLC patients.
Department of Radiation Oncology, Harbin Medical University, Harbin, Heilongjiang, China E-mail : xuxxyy@sohu.com; 18545169917@163.com.
Jian-Yu
JY
Xu
Shan
S
Lu
Xiang-Ying
XY
Xu
Song-Liu
SL
Hu
Bin
B
Li
Rui-Xue
RX
Qi
Lin
L
Chen
Joe Y
JY
Chang
D000230:Adenocarcinoma / Q000235:genetics*; D017209:Apoptosis / Q000235:genetics / Q000528:radiation effects; D018287:Carcinoma, Large Cell / Q000235:genetics*; D002289:Carcinoma, Non-Small-Cell Lung / Q000235:genetics; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000235:genetics / Q000528:radiation effects; D002470:Cell Survival / Q000235:genetics / Q000528:radiation effects; D053903:DNA Breaks, Double-Stranded / Q000528:radiation effects; D004260:DNA Repair / Q000235:genetics / Q000528:radiation effects; D051747:DNA-Activated Protein Kinase; D055785:Gene Knockdown Techniques; D006801:Humans; D008175:Lung Neoplasms / Q000235:genetics*; D009687:Nuclear Proteins; D010750:Phosphoproteins / Q000235:genetics*; D010766:Phosphorylation; D034622:RNA Interference; D016601:RNA-Binding Proteins / Q000235:genetics*; D011836:Radiation Tolerance / Q000235:genetics*; D000097796:Nucleolin
D051747:DNA-Activated Protein Kinase; D009687:Nuclear Proteins; D010750:Phosphoproteins; D016601:RNA-Binding Proteins; D000097796:Nucleolin; C497263:PRKDC protein, human