Yitao Wang

PRR11 regulates late-S to G2/M phase progression and induces premature chromatin condensation (PCC)

Recently, we have demonstrated that proline-rich protein 11 (PRR11) is a novel tumor-related gene product likely implicated in the regulation of cell cycle progression as well as lung cancer development. However, its precise role in cell cycle progression remains unclear. In the present study, we have further investigated the expression pattern and functional implication of PRR11 during cell cycle in detail in human lung carcinoma-derived H1299 cells. According to our immunofluorescence study, PRR11 was expressed largely in cytoplasm, the amount of PRR11 started to increase in the late S phase, and was retained until just before mitotic telophase. Consistent with those observations, siRNA-mediated knockdown of PRR11 caused a significant cell cycle arrest in the late S phase. Intriguingly, the treatment with dNTPs further augmented PRR11 silencing-mediated S phase arrest. Moreover, knockdown of PRR11 also resulted in a remarkable retardation of G2/M progression, and PRR11-knockdown cells subsequently underwent G2 phase cell cycle arrest accompanied by obvious mitotic defects such as multipolar spindles and multiple nuclei. In addition, forced expression of PRR11 promoted the premature Chromatin condensation (PCC), and then proliferation of PRR11-expressing cells was massively attenuated and induced apoptosis. Taken together, our current observations strongly suggest that PRR11, which is strictly regulated during cell cycle progression, plays a pivotal role in the regulation of accurate cell cycle progression through the late S phase to mitosis.

NID1, a new regulator of EMT required for metastasis and chemoresistance of ovarian cancer cells

Nidogen-1 (NID1) has been identified as a novel candidate diagnostic biomarker of ovarian cancer in our previous study. Nevertheless, the role of NID1 in the pathogenesis of ovarian cancer is unclear. In the present study, we demonstrated that NID1 was a mesenchymal associated gene and its high expression was significantly correlated with shorter overall survival of ovarian cancer patients. The ectopic expression of NID1 in OVCAR-3 cells revealed a epithelial-mesenchymal transition (EMT) phenotype accompanied by enhancement of motility, invasiveness and cisplatin resistance, whereas the knockdown of NID1 was sufficient to convert HEY cells into epithelial phenotype with decreased capability of motility, invasiveness and cisplatin resistance. Mechanistic studies disclosed that NID1 activated ERK/MAPK signaling pathway to promote EMT. Collectively, our findings have uncovered the molecular mechanisms of NID1 in promoting ovarian cancer metastasis and chemoresistance, and provide a rationale for the therapeutic potential of NID1 suppression in ovarian cancer.

B-Myb Is Up-Regulated and Promotes Cell Growth and Motility in Non-Small Cell Lung Cancer

B-Myb is a transcription factor that is overexpressed and plays an oncogenic role in several types of human cancers. However, its potential implication in lung cancer remains elusive. In the present study, we have for the first time investigated the expression profile of B-Myb and its functional impact in lung cancer. Expression analysis by quantificational real-time polymerase chain reaction (qRT-PCR) and immunohistochemistry demonstrated that B-Myb expression is aberrantly overexpressed in non-small cell lung cancer (NSCLC), and positively correlated with pathologic grade and clinical stage of NSCLC. A gain-of-function study revealed that overexpression of B-Myb significantly increases lung cancer cell growth, colony formation, migration, and invasion. Conversely, a loss-of-function study showed that knockdown of B-Myb decreases cell growth, migration, and invasion. B-Myb overexpression also promoted tumor growth in vivo in a NSCLC xenograft nude mouse model. A molecular mechanistic study by RNA-sequencing (RNA-seq) analysis showed that B-Myb overexpression causes up-regulation of various downstream genes (e.g., COL11A1, COL6A1, FN1, MMP2, NID1, FLT4, INSR, and CCNA1) and activation of multiple critical pathways (e.g., extracellular signal-regulated kinases (ERK) and phosphorylated-protein kinase B (Akt) signaling pathways) involved in cell proliferation, tumorigenesis, and metastasis. Collectively, our results indicate a tumor-promoting role for B-Myb in NSCLC and thus imply its potential as a target for the diagnosis and/or treatment of NSCLC.

Alkaline ceramidase 2 is a novel direct target of p53 and induces autophagy and apoptosis through ROS generation

ACER2 is a critical sphingolipid metabolizing enzyme, and has been shown to be remarkably up-regulated following various stimuli such as DNA damage. However, the transcriptional regulatory mechanism of ACER2 gene and its potential role in the regulation of autophagy remain unknown. In this study, we have for the first time identified the human ACER2 gene promoter, and found that human ACER2 transcription is directly regulated by p53 and ACER2 is implicated in the induction of autophagy as well as apoptosis. A series of luciferase reporter assay demonstrated that ACER2 major promoter is located within its first intron where the consensus p53-binding sites exist. Consistently, forced expression of p53 significantly stimulated ACER2 transcription. Notably, p53-mediated autophagy and apoptosis were markedly enhanced by ACER2. Depletion of the essential autophagy gene ATG5 revealed that ACER2-induced autophagy facilitates its effect on apoptosis. Further studies clearly showed that ACER2-mediated autophagy and apoptosis are accompanied by ROS generation. In summary, our present study strongly suggests that ACER2 plays a pivotal role in p53-induced autophagy and apoptosis, and thus might serve as a novel and attractive molecular target for cancer treatment.

B-Myb Mediates Proliferation and Migration of Non-Small-Cell Lung Cancer via Suppressing IGFBP3

B-Myb has been shown to play an important oncogenic role in several types of human cancers, including non-small-cell lung cancer (NSCLC). We previously found that B-Myb is aberrantly upregulated in NSCLC, and overexpression of B-Myb can significantly promote NSCLC cell growth and motility. In the present study, we have further investigated the therapeutic potential of B-Myb in NSCLC. Kaplan–Meier and Cox proportional hazards analysis indicated that high expression of B-Myb is significantly associated with poor prognosis in NSCLC patients. A loss-of-function study demonstrated that depletion of B-Myb resulted in significant inhibition of cell growth and delayed cell cycle progression in NSCLC cells. Notably, B-Myb depletion also decreased NSCLC cell migration and invasion ability as well as colony-forming ability. Moreover, an in vivo study demonstrated that B-Myb depletion caused significant inhibition of tumor growth in a NSCLC xenograft nude mouse model. A molecular mechanistic study by RNA-seq analysis revealed that B-Myb depletion led to deregulation of various downstream genes, including insulin-like growth factor binding protein 3 (IGFBP3). Overexpression of IGFBP3 suppressed the B-Myb-induced proliferation and migration, whereas knockdown of IGFBP3 significantly rescued the inhibited cell proliferation and motility caused by B-Myb siRNA (small interfering RNA). Expression and luciferase reporter assays revealed that B-Myb could directly suppress the expression of IGFBP3. Taken together, our results suggest that B-Myb functions as a tumor-promoting gene via suppressing IGFBP3 and could serve as a novel therapeutic target in NSCLC.

DJ-1 promotes colorectal cancer progression through activating PLAGL2/Wnt/BMP4 axis

Metastasis remains a big barrier for the clinical treatment of colorectal cancer (CRC). Our previous proteomics analysis identified DJ-1 as a potential metastasis biomarker of CRC. In this study, we found that DJ-1 was upregulated in CRC. The levels of DJ-1 were closely correlated with the depths of invasion and predicted patient outcome. Enforced expression of DJ-1 could enhance CRC proliferation and metastasis in vitro and in vivo by stimulating Wnt-β-catenin signaling. Specifically, DJ-1-induced β-catenin nuclear translocation stimulated TCF transcription activity, which promoted BMP4 expression for CRC cell migration and invasion, and elevated CCND1 expression for CRC cell proliferation, respectively. Furthermore, DJ-1-induced Wnt signaling activation was dependent on PLAGL2 expression. In conclusion, our study demonstrates that DJ-1 can promote CRC metastasis by activating PLAGL2–Wnt–BMP4 axis, suggesting novel therapeutic opportunities for postoperative adjuvant therapy in CRC patients.

DEPDC1 is required for cell cycle progression and motility in nasopharyngeal carcinoma

DEP domain containing 1 (DEPDC1) is a newly identified cancer-related and cell cycle related gene and has been demonstrated as a novel therapeutic target for bladder cancer. However, the functional involvement and therapeutic potential of DEPDC1 in nasopharyngeal carcinoma (NPC) remains unclear. Our results showed that DEPDC1 was overexpressed at both mRNA and protein levels in NPC tissues compared with normal or non-tumor tissues. The siRNA-mediated DEPDC1 depletion resulted in significant inhibition of proliferation and delay in cell cycle progression in both NPC cell lines, CNE-1 and HNE-1. Detailed analysis with indirect immunofluorescence assays revealed that DEPDC1 depletion caused significant mitotic arrest accompanied with mitotic defects such as multipolar spindles and multiple nuclei followed by apoptotic cell death. Notably, DEPDC1 depletion also reduces migration and invasion ability in both cell lines. Consistent with its regulatory role in NF-κB pathway, knockdown of DEPDC1 caused significant upregulation of A20 and downregulation of mutiple NF-κB downstream target genes implicated in proliferation and tumorigenesis (c-Myc, BCL2, CCND1, CCNB1 and CCNB2), and metastasis (MMP2, MMP9, ICAM1, vimentin, Twist1). Moreover, in vivo study demonstrated that DEPDC1 knockdown also caused significant inhibition of tumor growth in the NPC xenograft nude mouse model. Taken together, our present study demonstrated that DEPDC1 is essentially required for the accelerated cell cycle progression and motility in NPC cells, and strongly suggested that DEPDC1 may serve as a novel therapeutic target in NPC.DEP domain containing 1 (DEPDC1) is a newly identified cancer-related and cell cycle related gene and has been demonstrated as a novel therapeutic target for bladder cancer. However, the functional involvement and therapeutic potential of DEPDC1 in nasopharyngeal carcinoma (NPC) remains unclear. Our results showed that DEPDC1 was overexpressed at both mRNA and protein levels in NPC tissues compared with normal or non-tumor tissues. The siRNA-mediated DEPDC1 depletion resulted in significant inhibition of proliferation and delay in cell cycle progression in both NPC cell lines, CNE-1 and HNE-1. Detailed analysis with indirect immunofluorescence assays revealed that DEPDC1 depletion caused significant mitotic arrest accompanied with mitotic defects such as multipolar spindles and multiple nuclei followed by apoptotic cell death. Notably, DEPDC1 depletion also reduces migration and invasion ability in both cell lines. Consistent with its regulatory role in NF-κB pathway, knockdown of DEPDC1 caused significant upregulation of A20 and downregulation of mutiple NF-κB downstream target genes implicated in proliferation and tumorigenesis (c-Myc, BCL2, CCND1, CCNB1 and CCNB2), and metastasis (MMP2, MMP9, ICAM1, vimentin, Twist1). Moreover, in vivo study demonstrated that DEPDC1 knockdown also caused significant inhibition of tumor growth in the NPC xenograft nude mouse model. Taken together, our present study demonstrated that DEPDC1 is essentially required for the accelerated cell cycle progression and motility in NPC cells, and strongly suggested that DEPDC1 may serve as a novel therapeutic target in NPC.

The PRR11-SKA2 Bidirectional Transcription Unit Is Negatively Regulated by p53 through NF-Y in Lung Cancer Cells

We previously identified proline-rich protein 11 (PRR11) as a novel cancer-related gene that is implicated in the regulation of cell cycle and tumorigenesis. Our recent study demonstrated that PRR11 and its adjacent gene, kinetochore associated 2 (SKA2), constitute a classic head-to-head gene pair that is coordinately regulated by nuclear factor Y (NF-Y). In the present study, we further show that the PRR11-SKA2 bidirectional transcription unit is an indirect target of the tumor suppressor p53. A luciferase reporter assay revealed that overexpression of wild type p53, but not mutant p53, significantly represses the basal activity and NF-Y mediated transactivation of the PRR11-SKA2 bidirectional promoter. Deletion and mutation analysis of the PRR11-SKA2 promoter revealed that p53-mediated PRR11-SKA2 repression is dependent on the presence of functional NF-Y binding sites. Furthermore, a co-immunoprecipitation assay revealed that p53 associates with NF-Y in lung cancer cells, and a chromatin immunoprecipitation assay showed that p53 represses PRR11-SKA2 transcription by reducing the binding amount of NF-Y in the PRR11-SKA2 promoter region. Consistently, the ability of p53 to downregulate PRR11-SKA2 transcription was significantly attenuated upon siRNA-mediated depletion of nuclear factor Y subunit beta (NF-YB). Notably, lung cancer patients with lower expression of either PRR11 or SKA2 along with wild type p53 exhibited the best overall survival compared with others with p53 mutation and/or higher expression of either PRR11 or SKA2. Taken together, our results demonstrate that p53 negatively regulates the expression of the PRR11-SKA2 bidirectional transcription unit through NF-Y, suggesting that the inability to repress the PRR11-SKA2 bidirectional transcription unit after loss of p53 might contribute to tumorigenesis.