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USP22 promotes NSCLC tumorigenesis via MDMX up-regulation and subsequent p53 inhibition.

Ding F, Bao C, Tian Y, Xiao H, Wang M, Xie X, Hu F, Mei J - Int J Mol Sci (2014)

Bottom Line: Increasing evidence suggests that ubiquitin-specific protease 22 (USP22) has great clinicopathologic significance in oncology.Additionally, USP22 silencing downregulates MDMX protein expression and activates the p53 pathway.Taken together, our results suggest that USP22 promotes NSCLC tumorigenesis in vitro and in vivo through MDMX upregulation and subsequent p53 inhibition.

View Article: PubMed Central - PubMed

Affiliation: Department of Cardiothoracic Surgery, Xinhua Hospital, School of Medicine, Shanghai Jiaotong University, Shanghai 200092, China. drnail@sina.com.

ABSTRACT
Increasing evidence suggests that ubiquitin-specific protease 22 (USP22) has great clinicopathologic significance in oncology. In this study, we investigated the role of USP22 in human NSCLC tumorigenesis along with the underlying mechanisms of action. First, we determined the expression of USP22 in human NSCLC, as well as normal tissues and cell lines. We then studied the effects of USP22 silencing by shRNA on NSCLC cell growth in vitro and tumorigenesis in vivo, along with the effect on the p53 pathway. We found that USP22 is overexpressed in human NSCLC tissues and cell lines. USP22 silencing by shRNA inhibits proliferation, induces apoptosis and arrests cells at the G0/G1 phases in NSCLC cells and curbs human NSCLC tumor growth in a mouse xenograft model. Additionally, USP22 silencing downregulates MDMX protein expression and activates the p53 pathway. Our co-immunoprecipitation analysis shows that USP22 interacts with MDMX in NSCLC cells. Furthermore, MDMX silencing leads to growth arrest and apoptosis in NSCLC cells, and over-expression of MDMX reverses the USP22 silencing-induced effects. Taken together, our results suggest that USP22 promotes NSCLC tumorigenesis in vitro and in vivo through MDMX upregulation and subsequent p53 inhibition. USP22 may represent a novel target for NSCLC treatment.

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USP22 silencing suppresses proliferation and induces apoptosis and cell cycle arrest in NSCLC cells. (A) USP22 mRNA and protein expressions in A549 and NCl-H460 cells transfected with USP22 shRNA or scrambled control shRNA (SCR shRNA) (n = 3); (B) cell proliferation in transfected A549 and NCl-H460 cells by the MTT assay (n = 3); (C) cell cycle distribution by flow cytometry with PI staining (n = 3). Histograms are representative of three independent experiments; (D) Cell apoptosis by flow cytometry with Annexin V-FITC and PI double staining (n = 3). The values of the Q2 plus Q4 quadrant represent apoptosis rates. Data are expressed as the means ± SD. *p < 0.05 vs. SCR shRNA, **p < 0.01 vs. SCR shRNA.
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ijms-16-00307-f002: USP22 silencing suppresses proliferation and induces apoptosis and cell cycle arrest in NSCLC cells. (A) USP22 mRNA and protein expressions in A549 and NCl-H460 cells transfected with USP22 shRNA or scrambled control shRNA (SCR shRNA) (n = 3); (B) cell proliferation in transfected A549 and NCl-H460 cells by the MTT assay (n = 3); (C) cell cycle distribution by flow cytometry with PI staining (n = 3). Histograms are representative of three independent experiments; (D) Cell apoptosis by flow cytometry with Annexin V-FITC and PI double staining (n = 3). The values of the Q2 plus Q4 quadrant represent apoptosis rates. Data are expressed as the means ± SD. *p < 0.05 vs. SCR shRNA, **p < 0.01 vs. SCR shRNA.

Mentions: We studied the impact of UPS22 silencing on A549 and NCI-H460 cell growth and apoptosis using shRNA transfection. USP22 shRNA transfection down-regulated both the mRNA and protein expressions of USP22 in A549 and NCI-H460 (Figure 2A). We then studied cell proliferation using the MTT assay and cell cycle distribution and apoptosis using flow cytometry. Our data showed that USP22 silencing led to significantly slower cell growth compared with the control (p < 0.01 after 120 h) (Figure 2B). Meanwhile, our flow cytometry analysis revealed that, compared with the control, USP22 shRNA-transfected cells displayed a significantly higher portion of cells at the G0/G1 phases and significantly lower portions of cells at the S and G2/M phases (Figure 2C), indicating that USP22 silencing induces cell cycle arrest. Additionally, cells transfected with USP22 shRNA showed significantly increased apoptosis compared with the control (Figure 2D).


USP22 promotes NSCLC tumorigenesis via MDMX up-regulation and subsequent p53 inhibition.

Ding F, Bao C, Tian Y, Xiao H, Wang M, Xie X, Hu F, Mei J - Int J Mol Sci (2014)

USP22 silencing suppresses proliferation and induces apoptosis and cell cycle arrest in NSCLC cells. (A) USP22 mRNA and protein expressions in A549 and NCl-H460 cells transfected with USP22 shRNA or scrambled control shRNA (SCR shRNA) (n = 3); (B) cell proliferation in transfected A549 and NCl-H460 cells by the MTT assay (n = 3); (C) cell cycle distribution by flow cytometry with PI staining (n = 3). Histograms are representative of three independent experiments; (D) Cell apoptosis by flow cytometry with Annexin V-FITC and PI double staining (n = 3). The values of the Q2 plus Q4 quadrant represent apoptosis rates. Data are expressed as the means ± SD. *p < 0.05 vs. SCR shRNA, **p < 0.01 vs. SCR shRNA.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4307248&req=5

ijms-16-00307-f002: USP22 silencing suppresses proliferation and induces apoptosis and cell cycle arrest in NSCLC cells. (A) USP22 mRNA and protein expressions in A549 and NCl-H460 cells transfected with USP22 shRNA or scrambled control shRNA (SCR shRNA) (n = 3); (B) cell proliferation in transfected A549 and NCl-H460 cells by the MTT assay (n = 3); (C) cell cycle distribution by flow cytometry with PI staining (n = 3). Histograms are representative of three independent experiments; (D) Cell apoptosis by flow cytometry with Annexin V-FITC and PI double staining (n = 3). The values of the Q2 plus Q4 quadrant represent apoptosis rates. Data are expressed as the means ± SD. *p < 0.05 vs. SCR shRNA, **p < 0.01 vs. SCR shRNA.
Mentions: We studied the impact of UPS22 silencing on A549 and NCI-H460 cell growth and apoptosis using shRNA transfection. USP22 shRNA transfection down-regulated both the mRNA and protein expressions of USP22 in A549 and NCI-H460 (Figure 2A). We then studied cell proliferation using the MTT assay and cell cycle distribution and apoptosis using flow cytometry. Our data showed that USP22 silencing led to significantly slower cell growth compared with the control (p < 0.01 after 120 h) (Figure 2B). Meanwhile, our flow cytometry analysis revealed that, compared with the control, USP22 shRNA-transfected cells displayed a significantly higher portion of cells at the G0/G1 phases and significantly lower portions of cells at the S and G2/M phases (Figure 2C), indicating that USP22 silencing induces cell cycle arrest. Additionally, cells transfected with USP22 shRNA showed significantly increased apoptosis compared with the control (Figure 2D).

Bottom Line: Increasing evidence suggests that ubiquitin-specific protease 22 (USP22) has great clinicopathologic significance in oncology.Additionally, USP22 silencing downregulates MDMX protein expression and activates the p53 pathway.Taken together, our results suggest that USP22 promotes NSCLC tumorigenesis in vitro and in vivo through MDMX upregulation and subsequent p53 inhibition.

View Article: PubMed Central - PubMed

Affiliation: Department of Cardiothoracic Surgery, Xinhua Hospital, School of Medicine, Shanghai Jiaotong University, Shanghai 200092, China. drnail@sina.com.

ABSTRACT
Increasing evidence suggests that ubiquitin-specific protease 22 (USP22) has great clinicopathologic significance in oncology. In this study, we investigated the role of USP22 in human NSCLC tumorigenesis along with the underlying mechanisms of action. First, we determined the expression of USP22 in human NSCLC, as well as normal tissues and cell lines. We then studied the effects of USP22 silencing by shRNA on NSCLC cell growth in vitro and tumorigenesis in vivo, along with the effect on the p53 pathway. We found that USP22 is overexpressed in human NSCLC tissues and cell lines. USP22 silencing by shRNA inhibits proliferation, induces apoptosis and arrests cells at the G0/G1 phases in NSCLC cells and curbs human NSCLC tumor growth in a mouse xenograft model. Additionally, USP22 silencing downregulates MDMX protein expression and activates the p53 pathway. Our co-immunoprecipitation analysis shows that USP22 interacts with MDMX in NSCLC cells. Furthermore, MDMX silencing leads to growth arrest and apoptosis in NSCLC cells, and over-expression of MDMX reverses the USP22 silencing-induced effects. Taken together, our results suggest that USP22 promotes NSCLC tumorigenesis in vitro and in vivo through MDMX upregulation and subsequent p53 inhibition. USP22 may represent a novel target for NSCLC treatment.

Show MeSH
Related in: MedlinePlus