Amplification of ACK1 promotes gastric tumorigenesis via ECD-dependent p53 ubiquitination degradation.

Amplification or over-expression of an activated Cdc42-associated kinase 1 (ACK1) gene is common in breast, lung and ovarian cancers. However, little is known about the role of ACK1 in gastric tumorigenesis. Here, we found that DNA copy numbers of the ACK1 gene and its mRNA expression levels were significantly increased in gastric cancer (GC) compared to normal gastric tissues. Additionally, silencing ACK1 inhibited GC cell proliferation and colony formation, induced G2/M arrest and cellular apoptosis in vitro, and suppressed tumor growth in vivo. Gene Ontology annotation revealed that 147 differential proteins regulated by ACK1 knockdown were closely related with cellular survival. A cell cycle regulator, ecdysoneless homolog (ECD), was found to be significantly down-regulated by ACK1 knockdown. Silencing of ECD inhibited colony formation and induced G2/M arrest and cell apoptosis, which is similar to the effects of ACK1 knockdown. Silencing of ECD did not further enhance the effects of ACK1 knockdown on G2/M arrest and apoptosis, while silencing of ECD blocked the enhancement of colony formation by ACK1 over-expression. Over-expression of ACK or ECD promoted the ubiquitination of tumor suppressor p53 protein and decreased p53 levels, while silencing of ACK1 or ECD decreased the p53 ubiquitination level and increased p53 levels. Silencing of ECD attenuated the ubiquitination enhancement of p53 induced by ACK1 over-expression. Collectively, we demonstrate that amplification of ACK1 promotes gastric tumorigenesis by inducing an ECD-dependent ubiquitination degradation of p53.

Down-regulation of TRPS1 stimulates epithelial-mesenchymal transition and metastasis through repression of FOXA1.

The tricho-rhino-phalangeal syndrome 1 gene (TRPS1), which was initially found to be associated with tricho-rhino-phalangeal syndrome, is critical for the development and differentiation of bone, hair follicles and kidney. However, its role in cancer progression is largely unknown. In this study, we demonstrated that down-regulation of TRPS1 correlated with distant metastasis, tumour recurrence and poor survival rate in cancer patients. TRPS1 was frequently down-regulated in high-metastatic cancer cell lines from the breast, colon and nasopharynx. Silencing of TRPS1 stimulated epithelial-mesenchymal transition (EMT), migration and invasion in vitro and metastasis in vivo, while TRPS1 over-expression exhibited the opposite effects. Using quantitative proteomics, FOXA1, a negative regulator of epithelial-mesenchymal transition (EMT), was shown to be down-regulated by TRPS1 knockdown. Ectopic expression of FOXA1 blocked the enhancement of EMT, migration and invasion induced by TRPS1 silencing. Mechanistically, TRPS1, acting as a transcription activator, directly induced FOXA1 transcription by binding to the FOXA1 promoter. We further showed that down-regulation of TRPS1 was induced by miR-373 binding to the 3' UTR of TRPS1. Over-expression of TRPS1, but not TRPS1 3' UTR, blocked the enhancement of migration and invasion induced by miR-373. Taken together, we consider that down-regulation of TRPS1 by miR-373, acting as a transcriptional activator, promotes EMT and metastasis by repressing FOXA1 transcription, expanding upon its previously reported role as a transcription repressor. Copyright © 2016 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Genistein-induced mitotic arrest of gastric cancer cells by downregulating KIF20A, a proteomics study.

Genistein exerts its anticarcinogenic effects by inducing G2/M arrest and apoptosis of cancer cells. However, the precise molecular mechanism of action of genistein has not been completely elucidated. In this study, we used quantitative proteomics to identify the genistein-induced protein alterations in gastric cancer cells and investigate the molecular mechanism responsible for the anti-cancer actions of genistein. Total 86 proteins were identified to be regulated by genistein, most of which were clustered into the regulation of cell division and G2/M transition, consistent with the anti-cancer effect of genistein. Many proteins including kinesin family proteins, TPX2, CDCA8, and CIT were identified for the first time to be regulated by genistein. Interestingly, five kinesin family proteins including KIF11, KIF20A, KIF22, KIF23, and CENPF were found to be simultaneously downregulated by genistein. Significantly decreased KIF20A was selected for further functional studies. The silencing of KIF20A inhibited cell viability and induced G2/M arrest, similar to the effects of genistein treatment in gastric cancer. And the silencing of KIF20A also increased cancer cell sensitivity to genistein inhibition, whereas overexpression of KIF20A markedly attenuated genistein-induced cell viability inhibition and G2/M arrest. These observations suggested that KIF20A played an important role in anti-cancer actions of genistein, and thus may be a potential molecular target for drug intervention of gastric cancer.

The analysis of bi-level evolutionary graphs.

Evolutionary graphs (EGs), in which evolutionary dynamic is arranged on a graph, were initially proposed by Lieberman et al. [Lieberman, E., Hauert, C., Nowak, M.A., 2005. Evolutionary dynamics on graphs. Nature 433, 312-316] in the biological field and many biological phenomena are successfully explained. EGs on two levels (or bi-level EGs), based on some biological phenomena, are considered in this paper. The bi-level EGs are compared with the one-rooted EGs in two cases. One has the identical numbers of the followers, the other with the same numbers of total individuals. Then, some properties of the bi-level EGs are obtained. It is showed that bi-level EGs are more stable, and the bi-level EGs with just two leaders are the most stable, if they have identical followers respectively. The bi-level EGs theory can successfully explain the phenomena of symbiosis in biology.

[Effect of HLCDG1 gene transfection on growth of lung carcinoma cells].

BACKGROUND & OBJECTIVE: HLCDG1, which locates in chromosome 5q33 (between D5S436 and D5S470),is a novel gene that our laboratory has cloned recently. The expression of HLCDG1 gene was significantly down-regulated or deleted in the primary lung carcinoma. This study was designed to observe if HLCDG1 has the potential to suppress growth of lung carcinoma cells. METHODS: The recombinant plasmid, pcDNA3.1(+)/HLCDG1, was constructed and subsequently transfected into A549 cells through liposome transfection. The A549 cells stably expressing HLCDG1 gene were established by G418 selection. RT-PCR was used to demonstrate the expression of HLCDG1 gene. Furthermore, the cell proliferation assay, the soft agar assay, and the tumorigenesis assay were used to analyze the malignant phenotype of the HLCDG1-transfected cells. RESULTS: The HLCDG1-transfected cells exhibited the expression of HLCDG1 mRNA by RT-PCR. The population double time (PDT) of HLCDG1-transfected group, vector-transfected group, and nontransfected group were 70.0 hours, 43.3 hours, and 39.5 hours, respectively; the difference between HLCDG1-transfected group and the other two groups was significant (P< 0.05). The colony formation rates of HLCDG1-transfected group, the vector-transfected group, and nontransfected group were 8.5%, 29.0%, and 35.0%, respectively. The rate of HLCDG1-transfected cells was markedly lower than those of the other two groups (P< 0.05). Moreover, these clones were injected into athymic nude mice. After 43 days, they were killed, and their tumors were isolated. These tumors weighed 0.120g, 0.612g, and 0.924g, respectively. CONCLUSION: The expression of HLCDG1 in A549 cells may have the potential to suppress tumor cell growth and the tumorigenesis of A549 cells transplanted in nude mice. These results suggested that HLCDG1 gene might be a good candidate of tumor suppressor gene correlated with lung carcinoma.

[Cloning and expression analysis of lung carcinoma related gene HLCDG1].

BACKGROUND & OBJECTIVE: High frequent loss of hetero- zygosity (LOH) of 3p, 5q, 6q, 9p, 10q, 11p, 13q, 17p, and 19p in lung carcinoma was detected by comparative genomic hybridization (CGH) and genomic-wide scan for analysis of genetic alteration with microsatellite allelotying. It was indicated that there might be some other unknown tumor susceptible genes or suppressor genes likely to be involved in lung carcinoma development and progression. The aim of this study was to clone the full-length cDNA of LXDD1,an expressed sequence tag(EST) isolated by mRNA differential display, which is significantly down-regulated in lung carcinoma and represents a novel gene. METHODS: Differential expression of LXDD1 in lung carcinoma was confirmed by Northern blot analysis, the expression of the LXDD1 in human normal tissues and the size of the transcription of the LXDD1-representative gene were also determined using MTN (Multiple Tissues Northern Blots). The putative full-length cDNA of the EST-representative gene was cloned and analyzed by bioinformatics. In addition, differential reverse transcription polymerase chain reaction (RT-PCR) was used to detect the expression of the novel gene in various cancer cell lines, primary lung carcinomas, and matched normal lung tissues. RESULTS: The full length cDNA with no homology to any reported genes in the database of GenBank was successfully cloned and named HLCDG1 (Human lung carcinoma deleted gene 1, GenBank accession number AF447582). A transmembrane protein with 166 amino acids was deduced to come from the open reading frame of the 3113 bp full-length cDNA, HLCDG1 gene was confirmed to be located at chromosome band 5q33 by alignment of electric polymerase chain reaction (e-PCR). CONCLUSION: HLCDG1 is a novel gene down-regulated in lung carcinoma, which may be involved in the development of lung carcinoma.