Jinyi Liu

X-linked Inhibitor of Apoptosis Protein (XIAP) Mediates Cancer Cell Motility via Rho GDP Dissociation Inhibitor (RhoGDI)-dependent Regulation of the Cytoskeleton

X-linked inhibitor of apoptosis protein (XIAP) overexpression has been found to be associated with malignant cancer progression and aggression in individuals with many types of cancers. However, the molecular basis of XIAP in the regulation of cancer cell biological behavior remains largely unknown. In this study, we found that a deficiency of XIAP expression in human cancer cells by either knock-out or knockdown leads to a marked reduction in β-actin polymerization and cytoskeleton formation. Consistently, cell migration and invasion were also decreased in XIAP-deficient cells compared with parental wild-type cells. Subsequent studies demonstrated that the regulation of cell motility by XIAP depends on its interaction with the Rho GDP dissociation inhibitor (RhoGDI) via the XIAP RING domain. Furthermore, XIAP was found to negatively regulate RhoGDI SUMOylation, which might affect its activity in controlling cell motility. Collectively, our studies provide novel insights into the molecular mechanisms by which XIAP regulates cancer invasion and offer a further theoretical basis for setting XIAP as a potential prognostic marker and specific target for treatment of cancers with metastatic properties.

Association between urinary polycyclic aromatic hydrocarbon metabolites and sperm DNA damage: a population study in Chongqing, China.

Background Polycyclic aromatic hydrocarbons (PAHs), a class of the most ubiquitous environmental contaminants, may reduce male reproductive functions, but the data from human population studies are very limited. Objectives We designed this study to determine whether environmental exposure to PAHs contributes to the alteration in semen quality, sperm DNA damage, and apoptosis in the general male human population. Methods We measured urinary levels of four PAH metabolites and assessed semen quality, sperm apoptotic markers with Annexin V assay, and sperm DNA damage with comet assay in 232 men from Chongqing, China. Results We found that increased urinary 2-hydroxynaphthalene (2-OHNa) levels were associated with increased comet parameters, including the percentage of DNA in the tail (tail%) [β coefficient = 13.26% per log unit 2-OHNa (micrograms per gram creatinine); 95% confidence interval (CI), 7.97–18.55]; tail length (12.25; 95% CI, 0.01–24.52), and tail distribution (7.55; 95% CI, 1.28–18.83). The urinary level of 1-hydroxypyrene was associated only with increased tail% (5.32; 95% CI, 0.47–10.17). Additionally, the increased levels of four urinary PAH metabolites were significantly associated with decreased vital Annexin V negative sperm counts. However, there was no significant association between urinary PAH metabolite levels and human semen parameters or morphology of the sperm samples. Conclusions Our data indicate that the environmental level of PAH exposure is associated with increased sperm DNA damage but not with semen quality. These findings suggest that exposure to PAHs may disrupt sperm DNA and thereby interfere with human male fertility.

Urinary phthalate metabolites and male reproductive function parameters in Chongqing general population, China.

This study was designed to investigate the phthalates exposure levels in general population in Chongqing City of China, and to determine the possible associations between phthalate exposure and male reproductive function parameters. We recruited 232 general men through Chongqing Family Planning Research Institute and Reproductive Center of Chongqing. In a single spot urine sample from each man, phthalate metabolites, including mono-butyl phthalate (MBP), mono-ethyl phthalate (MEP), mono-(2-ethylhexyl) phthalate (MEHP), mono-benzyl phthalate (MBzP), phthalic acid (PA), and total PA were analyzed using solid phase extraction and coupled with high-performance liquid chromatography and detection by tandem mass spectrometry. Semen parameters were dichotomized based on World Health Organization reference values. Sperm DNA damage were analyzed using the alkaline single-cell gel electrophoresis assay. Reproductive hormones were determined in serum by the radioimmunoassay kit. We observed a weak association between urinary MBP concentration and sperm concentration in Chongqing general population. MBP levels above the median were 1.97 times (95% confidence interval [CI] 0.97-4.04) more likely to have sperm concentration below the reference value. There were no other associations between phthalate metabolites and reproductive function parameters after adjusted for potential risk factors. Our study suggested that general population in Chongqing area of China exposure to the environmental level of phthalate have weak or without adverse effects on the reproduction.

RhoGDI SUMOylation at Lys-138 increases its binding activity to Rho GTPase and its inhibiting cancer cell motility.

Background: RhoGDI affects biological activities of small Rho GTPases, leading to regulation of actin polymerization and cell motility. Results: RhoGDI SUMOylation at Lys-138 was crucial for the function of RhoGDI in cell motility. Conclusion: The regulation of SUMOylation of RhoGDI by XIAP plays a key role in regulating cancer cell Rho GTPase activation. Significance: Our study reveals a molecular basis for XIAP mediation of cancer cell invasion and metastasis. The Rho GDP dissociation inhibitor (RhoGDI) can bind to small GTPases and keep them in a biologically inactive state in cytoplasm, through which it affects actin polymerization and cell motility. However, mechanisms underlying how RhoGDI regulates Rho GTPase complex formation/membrane extraction/GTPase dissociation remain largely unexplored. Our previous studies reported that X-linked inhibitor of apoptosis protein (XIAP) interacted with RhoGDI via its RING domain and negatively modulated RhoGDI SUMOylation and HCT116 cancer cell migration. Here, we identified that RhoGDI SUMOylation specifically occurred at Lys-138, which was inhibited by XIAP domain. We further demonstrated that RhoGDI SUMOylation at Lys-138 was crucial for inhibiting actin polymerization and cytoskeleton formation as well as cancer cell motility. Moreover, SUMO-RhoGDI had a much higher binding affinity to small Rho GTPase compared with the un-SUMOylated form of RhoGDI. Taken together, our study demonstrated a novel modification of RhoGDI, SUMOylation at Lys-138, which played a key role in regulating Rho GTPase activation in cancer cells. The physiological regulation of RhoGDI SUMOylation by the RING domain of XIAP may account for modulation of cancer cell invasion and metastasis by XIAP.

Association between mobile phone use and semen quality: a systemic review and meta-analysis

Possible hazardous health effects of radiofrequency electromagnetic radiations emitted from mobile phone on the reproductive system have raised public concern in recent years. This systemic review and meta‐analysis was prepared following standard procedures of the Cochrane Collaboration and the Preferred Reporting Items for Systematic Reviews and Meta‐Analyses statement and checklist. Relevant studies published up to May 2013 were identified from five major international and Chinese literature databases: Medline/PubMed, EMBASE, CNKI, the VIP database and the Cochrane Central Register of Controlled Trials in the Cochrane Library. Eighteen studies with 3947 men and 186 rats were included in the systemic review, of which 12 studies (four human studies, four in vitro studies and four animal studies) with 1533 men and 97 rats were used in the meta‐analyses. Systemic review showed that results of most of the human studies and in vitro laboratory studies indicated mobile phone use or radiofrequency exposure had negative effects on the various semen parameters studied. However, meta‐analysis indicated that mobile phone use had no adverse effects on semen parameters in human studies. In the in vitro studies, meta‐analysis indicated that radiofrequency radiation had detrimental effect on sperm motility and viability in vitro [pooled mean difference (MDs) (95% CI): −4.11 (−8.08, −0.13), −3.82 (−7.00, −0.65) for sperm motility and viability respectively]. As for animal studies, radiofrequency exposure had harmful effects on sperm concentration and motility [pooled MDs (95% CI): −8.75 (−17.37, −0.12), −17.72 (−32.79, −2.65) for sperm concentration and motility respectively]. Evidence from current studies suggests potential harmful effects of mobile phone use on semen parameters. A further multicentred and standardized study is needed to assess the risk of mobile phone use on the reproductive system.

A Novel Polymorphism rs1329149 of CYP2E1 and a Known Polymorphism rs671 of ALDH2 of Alcohol Metabolizing Enzymes Are Associated with Colorectal Cancer in a Southwestern Chinese Population

Background: To screen for tagging single nucleotide polymorphisms (tagSNP) in the major alcohol metabolizing enzymes: ADH1B, ALDH2, and CYP2E1, and to evaluate the association between these tagSNPs and colorectal cancer (CRC) in a southwestern Chinese population. Methods: A hospital-based case-control study of 440 CRC patients and 800 cancer-free controls was conducted. Personal information was collected by a Semi-Quantitative Food Frequency Questionnaire. The tagSNPs were screened in the HapMap with Haploview by setting the minor allele frequency at 0.03 with the highest score of r2 form each block. Genotypes were identified by using the SNPLex System. Both crude and adjusted odds ratio (OR) and 95% confidence interval (CI) were used to evaluate the risk of each SNP. Results: Sixteen tagSNPs were selected, and 13 were successfully genotyped. A novel CYP2E1 locus rs1329149 and a known ALDH2 locus rs671 were found to be significantly associated with CRC risk. The adjusted OR was 1.86 (95% CI, 1.12-3.09) for the rs671 A/A genotype and 4.04 for the rs1329149 T/T genotype (95% CI, 2.44-6.70), compared with their common homozygous genotypes. Interaction was found between alcohol consumption and gene polymorphisms on CRC, the adjusted OR was 7.17 (95% CI, 2.01-25.53) for drinking habits combined with rs671 A/A or rs1329149 T/T genotype. Conclusion: The results of this study suggest that rs671 A/A and the first reported locus rs1329149 T/T genotypes increase the susceptibility to CRC, and gene-environmental interaction between the two loci and alcohol use existed for CRC in Southwestern Chinese. Larger studies are warranted to verify our findings. (Cancer Epidemiol Biomarkers Prev 2009;18(9):2522–7)

Anti-cancer effects of JKA97 are associated with its induction of cell apoptosis via a Bax-dependent and p53-independent pathway.

p53, one of the most commonly mutated genes in human cancers, is thought to be associated with cancer development. Hence, screening and identifying natural or synthetic compounds with anti-cancer activity via p53-independent pathway is one of the most challenging tasks for scientists in this field. Compound JKA97 (methoxy-1-styryl-9H-pyrid-[3,4-b]-indole) is a small molecule synthetic anti-cancer agent, with unknown mechanism(s). In this study we have demonstrated that the anti-cancer activity of JKA97 is associated with apoptotic induction via p53-independent mechanisms. We found that co-incubation of human colon cancer HCT116 cells with JKA97 inhibited HCT116 cell anchorage-independent growth in vitro and tumorigenicity in nude mice and also induced a cell apoptotic response, both in the cell culture model and in a tumorigenesis nude mouse model. Further studies showed that JKA97-induced apoptosis was dramatically impaired in Bax knock-out (Bax-/-) HCT116 cells, whereas the knock-out of p53 or PUMA did not show any inhibitory effects. The p53-independent apoptotic induction by JKA97 was confirmed in other colon cancer and hepatocarcinoma cell lines. In addition, our results showed an induction of Bax translocation and cytochrome c release from the mitochondria to the cytosol in HCT116 cells, demonstrating that the compound induces apoptosis through a Bax-initiated mitochondria-dependent pathway. These studies provide a molecular basis for the therapeutic application of JKA97 against human cancers with p53 mutations.

CpG island hypermethylation of multiple tumor suppressor genes associated with loss of their protein expression during rat lung carcinogenesis induced by 3-methylcholanthrene and diethylnitrosamine

The epigenetic mechanisms underlying the tumorigenesis caused by polycyclic aromatic hydrocarbons and nitrosamine compounds such as 3-methylcholanthrene (MCA) and diethylnitrosamine (DEN) are currently unknown. We reported previously that dynamic changes in DNA methylation occurred during MCA/DEN-induced rat lung carcinogenesis. Here, we used the same animal model to further study the evolution of methylation alterations in tumor suppressor genes (TSGs) DAPK1, FHIT, RASSF1A, and SOCS-3. We found that none of these genes were methylated in either normal or hyperplasia tissue. However, as the severity of the cancer progressed through squamous metaplasia and dysplasia to carcinoma in situ (CIS) and infiltrating carcinoma, so methylation became more prevalent. Particularly dramatic increases in the level of methylation, the average number of methylated genes, and the incidence of concurrent methylation in three genes were observed in CIS and infiltrating carcinoma. Similar but less profound changes were seen in squamous metaplasia and dysplasia. Furthermore, methylation status was closely correlated to loss of protein expression for these genes, with protein levels markedly declining along the continuum of carcinogenesis. These results suggest that progressive CpG island hypermethylation leading to inactivation of TSGs might be a vital molecular mechanism in the pathogenesis of MCA/DEN-induced multistep rat lung carcinogenesis.

p27 suppresses arsenite-induced Hsp27/Hsp70 expression through inhibiting JNK2/c-Jun- and HSF-1-dependent pathways.

p27 is an atypical tumor suppressor that can regulate the activity of cyclin-dependent kinases and G0-to-S phase transitions. More recent studies reveal that p27 may also exhibit its tumor-suppressive function through regulating many other essential cellular events. However, the molecular mechanisms underlying these anticancer effects of p27 are largely unknown. In this study, we found that depletion of p27 expression by either gene knock-out or knockdown approaches resulted in up-regulation of both Hsp27 and Hsp70 expression at mRNA- and promoter-derived transcription as well as protein levels upon arsenite exposure, indicating that p27 provides a negative signal for regulating the expression of Hsp27 and Hsp70. Consistently, arsenite-induced activation of JNK2/c-Jun and HSF-1 pathways was also markedly elevated in p27 knock-out (p27−/−) and knockdown (p27 shRNA) cells. Moreover, interference with the expression or function of JNK2, c-Jun, and HSF-1, but not JNK1, led to dramatic inhibition of arsenite-induced Hsp27 and Hsp70 expression. Collectively, our results demonstrate that p27 suppresses Hsp27 and Hsp70 expression at the transcriptional level specifically through JNK2/c-Jun- and HSF-1-dependent pathways upon arsenite exposure, which provides additional important molecular mechanisms for the tumor-suppressive function of p27.

E3 ligase activity of XIAP RING domain is required for XIAP-mediated cancer cell migration, but not for its RhoGDI binding activity.

Although an increased expression level of XIAP is associated with cancer cell metastasis, the underlying molecular mechanisms remain largely unexplored. To verify the specific structural basis of XIAP for regulation of cancer cell migration, we introduced different XIAP domains into XIAP−/− HCT116 cells, and found that reconstitutive expression of full length HA-XIAP and HA-XIAP ΔBIR, both of which have intact RING domain, restored β-Actin expression, actin polymerization and cancer cell motility. Whereas introduction of HA-XIAP ΔRING or H467A mutant, which abolished its E3 ligase function, did not show obvious restoration, demonstrating that E3 ligase activity of XIAP RING domain played a crucial role of XIAP in regulation of cancer cell motility. Moreover, RING domain rather than BIR domain was required for interaction with RhoGDI independent on its E3 ligase activity. To sum up, our present studies found that role of XIAP in regulating cellular motility was uncoupled from its caspase-inhibitory properties, but related to physical interaction between RhoGDI and its RING domain. Although E3 ligase activity of RING domain contributed to cell migration, it was not involved in RhoGDI binding nor its ubiquitinational modification.