Population structure and genetic diversity of Tamarix chinensis as revealed with microsatellite markers in two estuarine flats.

Background: Tamarix chinensis Lour. is a 3-6-meter-tall small tree with high salt- and alkali- tolerance and aggressive invasiveness, mainly distributed in the eastern part of China in warm-temperate and subtropical climate zones, yet there is little information available regarding genetic diversity and population structure. Methods: A total of 204 individuals of nine T. chinensis populations were investigated for genetic diversity and population structure using a set of 12 highly polymorphic microsatellite markers. Results: The total number of alleles detected was 162, the average number of effective allele was 4.607, the average polymorphism information content (PIC) value of the 12 loci was 0.685, and the mean observed heterozygosity (Ho) and the mean expected heterozygosity (He) was 0.653 and 0.711, respectively. Analysis of molecular variance (AMOVA) showed a 5.32% genetic variation among T. chinensis populations. Despite a low population differentiation, Bayesian clustering analysis, discriminant analysis of principal components (DAPC) and the unweighted pair group method with arithmetic mean (UPGMA) clearly identified three genetic clusters correlated to the populations' geographic origin: the northern populations including those from Yellow River Delta, the Fangshan (FS) population from Beijing, the Changyi (CY) population from Bohai Bay, the Huanjiabu (HHJ) population from Hangzhou Bay, and the remaining two populations from Hangzhou Bay. There was a significant relationship between the genetic distance and geographical distance of the paired populations. Gene flow (Nm) was 4.254 estimated from FST. Conclusion: T. chinensis possessed high genetic diversity comparable to tree species, and although the population differentiation is shallow, our results classified the sampled populations according to sampling localities, suggesting the different origins of the study populations.

The Diverse Analysis Identifies Mutated KRAS Associated With Radioresistance in Non-Small Cell Lung Cancer.

Background: To analyze the relationship between V-Ki-ras2 Kirsten rat sarcoma viral oncogene homolog (KRAS) status and radioresistance in non-small cell lung cancer (NSCLC), we identified potential genotypic differences and pathways involved. Methods: We retrospectively analyzed epidermal growth factor receptor (EGFR) and KRAS status in patients undergoing definitive radiotherapy for NSCLC between 2004 and 2018. Cox proportional hazard models were used to evaluate local progression-free survival (LPFS). Using clonogenic survival and measurement of γH2AX foci, we analyzed the difference in radiosensitivity between NSCLC cell lines with different KRAS status. The Cancer Genome Atlas (TCGA) analysis was used to explore the potential pathways involved. Results: The results showed that of the 286 patients identified, 68 (24%) had local tumor progression (mean ± standard deviation (SD), 27 ± 17.4 months); of these patients, KRAS mutations were found in 14 (23%), and KRAS status was associated with LPFS. After adjusting for concurrent chemotherapy, gross tumor volume, and mutation status in multivariate analysis, KRAS mutation was associated with shorter LPFS (hazard ratio: 1.961; 95% confidence interval: 1.03 - 2.17; P = 0.032). KRAS mutation showed higher radioresistance in vitro. TCGA data showed that the ERK1/2 pathway, phosphatidylinositol I3 kinase (PI3K)/mTOR, p38 MAPK pathway, cell cycle checkpoint signaling, DNA damage, repair pathways, and EGFR/PKC/AKT pathway were differentially expressed in patients with KRAS mutations or cell lines compared with their expression in the wild-type group. Conclusions: Diverse analyses identified that KRAS mutation was associated with radioresistance in NSCLC. KRAS mutation status may be helpful as a biomarker of radioresistance and a potential target to increase radiosensitivity.

Erratum: Oncogenic roles of carbonic anhydrase IX in human nasopharyngeal carcinoma.

[This corrects the article on p. 2942 in vol. 7, PMID: 25031713.].

Assessment of endophytic bacterial diversity in rose by high-throughput sequencing analysis.

The endophytic bacterial diversity of rose was analyzed by high-throughput sequencing of 16S rDNA and functional prediction of the bacterial community. The number of bacterial sequence reads obtained from 18 rose samples ranged from 63,951 to 114,833, and reads were allocated to 1982 OTUs based on sequences of the V3-V4 region. The highest Shannon Index was found in Luogang rose (1.93), while the lowest was found in Grasse rose (0.35). The bacterial sequence reads were grouped into three different phyla: Firmicutes, Proteobacteria, and Actinobacteria. At the genus level, Bacillus and Staphylococcus had the highest abundance across all 18 samples; Bacillus was particularly abundant in Daguo rose (99.09%), Rosa damascena (99.65%), and Fenghua rose (99.58%). Unclassified OTUs were also found in all samples. PICRUSt gene prediction revealed that each endophyte sample contained multiple KEGG functional modules related to human metabolism and health. A high abundance of functional genes were involved in (1) Amino Acid Metabolism, (2) Carbohydrate Metabolism, (3) Cellular Processes and Signaling, (4) Energy Metabolism, and (5) Membrane Transport, indicating that the endophytic community comprised a wide variety of microorganisms and genes that could be used for further studies. The rose endophytic bacterial community is rich in diversity; community composition varies among roses and contains functional information related to human health.

EFEMP1 inhibits migration of hepatocellular carcinoma by regulating MMP2 and MMP9 via ERK1/2 activity.

The role of epidermal growth factor-containing fibulin-like extracellular matrix protein 1 (EFEMP1) inhibiting migration in hepatocellular carcinoma (HCC) remains unknown. Expression of EFEMP1 in HCC cell lines were quantified by western blotting and real-time PCR. The role of EFEMP1 in HCC cell migration was explored in vitro via siRNA and adding purified EFEMP1 protein. The associated molecule expression was detected by western blotting after downregulation of EFEMP1 and also tested by immunohistochemistry. Eight pairs of HCC non-HCC liver samples and 215 HCC samples were subjected to immunohistochemistry. EFEMP1 was highly expressed in 7,721 and HepG2 HCC cell lines while HuH7 HCC cell line expressed the lowest level of EFEMP1 compared with the others. Downregulating EFEMP1 by siRNA markedly increased the migration ability of HCC cells while adding purified EFEMP1 protein inhibited HCC cell migration. Downregulation of EFEMP1 increased the expression of ERK1/2, MMP2 and MMP9. Furthermore, U0126 (a highly selective and potent inhibitor of pERK1/2) could abrogate the migration ability enhanced by siRNA. Accordingly, MMP2 and MMP9 were inversely expressed with EFEMP1 expression by immunohistochemistry. EFEMP1 downregulated in HCC tissues, and lower EFEMP1 expression was significantly associated with HCC patients with ascites (P=0.050), vascular invasion (P=0.044), poorer differentiation (P=0.002) and higher clinical stage (P=0.003).

KCTD12 Regulates Colorectal Cancer Cell Stemness through the ERK Pathway.

Targeting cancer stem cells (CSCs) in colorectal cancer (CRC) remains a difficult problem, as the regulation of CSCs in CRC is poorly understood. Here we demonstrated that KCTD12, potassium channel tetramerization domain containing 12, is down-regulated in the CSC-like cells of CRC. The silencing of endogenous KCTD12 and the overexpression of ectopic KCTD12 dramatically enhances and represses CRC cell stemness, respectively, as assessed in vitro and in vivo using a colony formation assay, a spheroid formation assay and a xenograft tumor model. Mechanistically, KCTD12 suppresses CRC cell stemness markers, such as CD44, CD133 and CD29, by inhibiting the ERK pathway, as the ERK1/2 inhibitor U0126 abolishes the increase in expression of CRC cell stemness markers induced by the down-regulation of KCTD12. Indeed, a decreased level of KCTD12 is detected in CRC tissues compared with their adjacent normal tissues and is an independent prognostic factor for poor overall and disease free survival in patients with CRC (p = 0.007). Taken together, this report reveals that KCTD12 is a novel regulator of CRC cell stemness and may serve as a novel prognostic marker and therapeutic target for patients with CRC.

TEL2 suppresses metastasis by down-regulating SERPINE1 in nasopharyngeal carcinoma.

Metastasis is the major cause of treatment failure in patients with nasopharyngeal carcinoma (NPC). However, the molecular mechanisms of NPC metastasis are poorly understood. Here, using our customized gene microarray containing all of the known human transcription factors and the current markers for epithelial-mesenchymal transition, we report that TEL2 was down-regulated in highly metastatic NPC cells and the metastatic tissues in lymph node. Mechanistically, TEL2 inhibits the cell migration and invasion in vitro and metastasis in vivo by directly suppressing the SERPINE1 promoter in NPC. Consistently, an inverse correlation was observed between the protein levels of TEL2 and SERPINE1 using clinical NPC samples. Collectively, we have provided the first evidence that TEL2 plays a key role in NPC metastasis by directly down-regulating SERPINE1, and that this novel axis of TEL2 / SERPINE1 may be valuable to develop new strategies for treating NPC patients with metastasis.

Down-regulation of prostate stem cell antigen (PSCA) by Slug promotes metastasis in nasopharyngeal carcinoma.

Distant metastasis and local recurrence are still the major causes for failure of treatment in patients with nasopharyngeal carcinoma (NPC), making it urgent to further elicit the molecular mechanisms of NPC metastasis. Using a gene microarray including transcription factors and known markers for cancer stem cells, prostate stem cell antigen (PSCA) was found to be significantly down-regulated in metastatic NPC in lymph node, compared to its primary tumour, and in NPC cell lines with high metastatic ability compared to those with low metastatic ability. NPC patients with low PSCA expression had a consistently poor metastasis-free survival (p = 0.003). Knockdown and overexpression of PSCA respectively enhanced and impaired the migration and invasion in vitro and the lung metastasis in vivo of NPC cells. Mechanistically, the enhancement of NPC metastasis by knocking down PSCA probably involved epithelial-mesenchymal transition (EMT), by up-regulating N-cadherin and ZEB1/2 and by activating RhoA. The down-regulation of PSCA in NPC cells resulted directly from the binding of Slug to the PSCA promoter. PSCA may be a potential diagnostic marker and therapeutic target for patients with NPC.

Aspirin Suppresses the Growth and Metastasis of Osteosarcoma through the NF-κB Pathway.

PURPOSE: Aspirin has recently been reported to reduce both the incidence and the risk of metastasis in colon cancer. However, there is no evidence at the cellular levels or in the animal models for such an effect of aspirin on cancer metastasis. EXPERIMENTAL DESIGN: MTT assay, colony formation assay, and apoptosis assay were employed to analyze the effects of aspirin on the osteosarcoma cell viability in vitro. The NF-κB activity was measured by the NF-κB p65 luciferase reporter. Western blotting was used to analyze the proteins in cells. The migration and invasion abilities of osteosarcoma cells in vitro were measured by the Transwell assay. Xenograft-bearing mice were used to assess the roles of aspirin in both tumor growth and metastasis of osteosarcoma in vivo (n = 5-8 mice/group). An unpaired Student t test or ANOVA with the Bonferroni post hoc test were used for the statistical comparisons. RESULTS: Aspirin reduced cell viability in a dose- and time-dependent manner in osteosarcoma cell lines, and aspirin synergistically sensitized osteosarcoma cells to cisplatin (DDP) in vitro and in vivo (P < 0.001). Moreover, aspirin markedly repressed the migration and invasion of osteosarcoma cells in vitro (P < 0.001), and dramatically diminished the occurrence of osteosarcoma xenograft metastases to the lungs in vivo (P < 0.001). Mechanistically, aspirin diminishes osteosarcoma migration, invasion, and metastasis through the NF-κB pathway. CONCLUSIONS: Aspirin suppresses both the growth and metastasis of osteosarcoma through the NF-κB pathway at the cellular level and in the animal models.

Oncogenic roles of carbonic anhydrase IX in human nasopharyngeal carcinoma.

Carbonic anhydrase IX (CA IX), a hypoxia-inducible protein in tumors, has been shown to be valuable for the prognosis of nasopharyngeal carcinoma (NPC). However, the function and mechanism of CA IX has been not explored in NPC. Here, we found that CA IX was detected at higher levels in NPC cells and tissues than their corresponding partners. Furthermore, the cell growth, migration and invasion in vitro were altered with shRNA or overexpression of CA IX in NPC cells. More importantly, the metastatic ability of NPC cells stably expressing CA IX was significantly enhanced using the hepatic metastasis model of nude mice in vivo. Finally, the mTOR pathway was indicated to be involved in such effects of CA IX on NPC. This is the first evidence that CA IX may promote the NPC metastasis to potentially be a therapeutic target for NPC, and that the inhibitory molecules of CA IX and/or the mTOR pathway alone or combination with both may be worth to have a clinical trial for the patients with NPC.

Targeting the anaphase-promoting complex/cyclosome (APC/C)- bromodomain containing 7 (BRD7) pathway for human osteosarcoma.

Osteosarcoma is the most common primary malignant bone tumor in childhood and adolescence and has a propensity for local invasion and early lung metastasis. However, the current therapies often result in chemoresistance, and a therapeutic target is not available in the clinic for osteosarcoma. Here, we report that BRD7 forms a complex with the anaphase-promoting complex/cyclosome (APC/C) and is degraded by APC/C(cdh1) and APC/C(cdc20) during the cell cycle. Moreover, BRD7 is a tumor suppressor in osteosarcoma, and the BRD7 mutant resistant to degradation by APC/C is more efficient than the wild-type protein at suppressing proliferation, colony formation, and tumor growth of osteosarcoma in vitro and in vivo. The combination of proTAME, an inhibitor of APC/C, with chemotherapeutic drugs efficiently targets osteosarcoma in vitro. Furthermore, there is a strong inverse correlation of protein levels between BRD7 and Cdh1 or Cdc20, and lower BRD7 expression is an indicator for poor prognosis in patients with osteosarcoma. Collectively, our results indicate that targeting the APC/C-BRD7 pathway may be a novel strategy for treating osteosarcoma.

Glycogen synthase kinase-3ß, NF-κB signaling, and tumorigenesis of human osteosarcoma.

BACKGROUND: Glycogen synthase kinase-3ß (GSK-3ß), a serine/threonine protein kinase, may function as a tumor suppressor or an oncogene, depending on the tumor type. We sought to determine the biological function of GSK-3ß in osteosarcoma, a rare pediatric cancer for which the identification of new therapeutic targets is urgent. METHODS: We used cell viability assays, colony formation assays, and apoptosis assays to analyze the effects of altered GSK-3ß expression in U2OS, MG63, SAOS2, U2OS/MTX300, and ZOS osteosarcoma cell lines. Nude mice (n = 5-8 mice per group) were injected with U2OS/MTX300, and ZOS cells to assess the role of GSK-3ß in osteosarcoma growth in vivo and to evaluate the effects of inhibitors and/or anticancer drugs on tumor growth. We used an antibody array, polymerase chain reaction, western blotting, and a luciferase reporter assay to establish the effect of GSK-3ß inhibition on the nuclear factor-κB (NF-κB) pathway. Immunochemistry was performed on primary tumor specimens from osteosarcoma patients (n = 74) to determine the relationship of GSK-3ß activity with overall survival. RESULTS: Osteosarcoma cells with low levels of inactive p-Ser9-GSK-3ß formed colonies in vitro and tumors in vivo more readily than cells with higher levels and cells in which GSK-3ß had been silenced formed fewer colonies and smaller tumors than parental cells. Silencing or pharmacological inhibition of GSK-3ß resulted in apoptosis of osteosarcoma cells. Inhibition of GSK-3ß resulted in inhibition of the NF-κB pathway and reduction of NF-κB-mediated transcription. Combination treatments with GSK-3ß inhibitors, NF-κB inhibitors, and chemotherapy drugs increased the effectiveness of chemotherapy drugs in vitro and in vivo. Patients whose osteosarcoma specimens had hyperactive GSK-3ß, and nuclear NF-κB had a shorter median overall survival time (49.2 months) compared with patients whose tumors had inactive GSK-3ß and NF-κB (109.2 months). CONCLUSION: GSK-3ß activity may promote osteosarcoma tumor growth, and therapeutic targeting of the GSK-3ß and/or NF-κB pathways may be an effective way to enhance the therapeutic activity of anticancer drugs against osteosarcoma.

Correlation of Skp2 overexpression to prognosis of patients with nasopharyngeal carcinoma from South China.

S-phase kinase-associated protein 2 (Skp2), which plays a role in cell cycle regulation, is commonly overexpressed in a variety of human cancers and associated with poor prognosis. However, its role in nasopharyngeal carcinoma (NPC) is not well understood. In this study, we examined the clinical significance of Skp2, with a particular emphasis on overall survival (OS) and disease-free survival (DFS), in NPC cases in South China, where NPC is an epidemic. Additionally, we explored the function of Skp2 in maintaining a cancer stem cell-like phenotype in NPC cell lines. Skp2 expression was assessed for 127 NPC patients using tissue microarrays and immunohistochemistry and analyzed together with clinicopathologic features, OS, and DFS. Skp2 expression was detectable, or positive, in 75.6% of patients. Although there was no correlation between Skp2 and any clinicopathologic factor, Skp2 expression significantly portended inferior OS (P = 0.013) and DFS (P = 0.012). In the multivariate model, Skp2 expression remained significantly predictive of poor OS [P = 0.009, risk ratio (RR) = 4.06] and DFS (P = 0.008, RR = 3.56), and this was also true for clinical stage (P = 0.012 and RR=3.201 for OS; P = 0.002 and RR=1.94 for DFS) and sex (P = 0.016 and RR=0.31 for OS; P = 0.006 and RR = 0.27 for DFS). After Skp2 knockdown, a colony formation assay was used to evaluate the self-renewal property of stem-like cells in the NPC cell lines CNE-1 and CNE-2. The colony formation efficiency in CNE-1 and CNE-2 cells was decreased. In Skp2-transfected CNE-1 and CNE-2 cells, side population (SP) proportion was increased as detected by flow cytometry. Skp2 is an independent prognostic marker for OS and DFS in NPC. Skp2 may play a role in maintaining the cancer stem cell-like phenotype of NPC cell lines.

The dominance of China 1 in the spectrum of Epstein-Barr virus strains from Cantonese patients with nasopharyngeal carcinoma.

Nasopharyngeal carcinoma is a disease with a remarkable geographic and ethnic distribution, and has a high incidence in southern China. Infection with Epstein-Barr virus (EBV) is an important contributing factor. The profile of EBV strains in Cantonese patients from Guangdong, the nasopharyngeal carcinoma endemic region in southern China, is described on the sequence variations in latent membrane protein 1 carboxyl-terminus. The results show that China 1 was the dominant EBV strain detected in both the tumor biopsies and samples of throat washings, whereas multiple strains, including China 1, China 2, B95-8, and Med, were detected in blood samples. In addition, a new strain named China 4 was found in blood samples. These findings suggest that the host population is susceptible to the predominant China 1 strain in the nasopharyngeal carcinoma endemic region of China, but its relationship with the host remains to be characterized further.

[Functions of V-val subtype of Epstein-Barr nuclear antigen 1].

BACKGROUND & OBJECTIVE: Epstein-Barr viral nuclear antigen 1 (EBNA1) plays a crucial role in the latency of Epstein-Barr virus (EBV). A close relation of V-val subtype of EBNA1 with nasopharyngeal carcinoma (NPC) was suggested by its preference to infect NPC cells. This study was to investigate the functional difference between prototype and V-val EBNA1 in epithelial cell line HEK293. METHODS: The coding sequences of prototype and V-val EBNA1 were amplified by polymerase chain reaction and cloned into pGFP vector, then transfected into HEK293 cells respectively. The biological consequences of EBNA1 gene expression were examined. The transcriptional activation ability was compared between prototype and V-val subtype of EBNA1 using luciferase reporter system containing family of repeats (FR) sequence of EBV. RESULTS: Prototype and V-val EBNA1 showed no effect on cell proliferation, while the cloning efficiency of prototype EBNA1-expressing cells was obviously lower than that of V-val EBNA1-expressing cells. No tumor formed in nude mice after injection of prototype or V-val EBNA1-trasfected HEK293 cells. However, the luciferase activity was significantly higher in V-val EBNA1-expressing HEK293 cells than in prototype EBNA1-expressing HEK293 cells in transient transfection assay. CONCLUSION: Prototype and V-val EBNA1 have no direct transforming activity on cells, whereas the transactivation activity of V-val EBNA1 in FR-containing plasmid is significantly higher than that of prototype EBNA1.

[Correlation of Epstein-Barr virus A73 gene polymorphisms to susceptibility to nasopharyngeal carcinoma].

BACKGROUND & OBJECTIVE: A73 gene, one of the primary members in Epstein-Barr virus (EBV) BamHI-A rightward transcripts (BARTs) family, is related to the development of nasopharyngeal carcinoma (NPC). Its mRNA level is elevated in NPC tissues, suggesting that A73 participates in the carcinogenesis of NPC. This study was to explore the polymorphism loci of A73 gene and find its correlation to susceptibility to NPC. METHODS: Nested polymerase chain reaction (PCR) and direct sequencing were used to identify A73 gene polymorphisms in 23 specimens of NPC and 19 throat washing (TW) samples from non-NPC patients. The same methods were also used on additional 54 unitized NPC specimens [carcinoma tissue, matched TW sample and peripheral blood (PB) sample from the same patient], and 48 TW samples and 51 PB samples from healthy Cantonese to detect a polymorphism with high frequency found in previous step. Its distribution among different samples was compared. All sequences were compared with the sequence of B95.8 cells. RESULTS: Three polymorphisms A157154C, G159188C, and G159209C were identified in A73 gene. Among which, A157154C showed high frequency. The frequency of A157154C was significant higher in PB samples from NPC patients than in those from healthy Cantonese (96.29% vs. 64.70%, P<0.001). Among 54 unitized specimens, 13 cases showed discordances in different tissue origins. CONCLUSION: A157154C in A73 gene might be correlated to the occurrence and development of NPC.

Haplotype of gene Nedd4 binding protein 2 associated with sporadic nasopharyngeal carcinoma in the Southern Chinese population.

BACKGROUND: Bcl-3 as an oncoprotein is overexpressed in nasopharyngeal carcinoma (NPC). Nedd4 binding protein 2 (N4BP2), which is located in the NPC susceptibility locus, is a Bcl-3 binding protein. This study is aimed to explore the association between N4BP2 genetic polymorphism and the risk of NPC. METHODS: We performed a hospital-based case-control study, including 531 sporadic NPC and 480 cancer-free control subjects from southern China. PCR-sequencing was carried out on Exons, promoter region and nearby introns of the N4BP2 gene. The expression pattern of N4BP2 and Bcl-3 was also analyzed. RESULTS: We observed a statistically significant difference in haplotype blocks ATTA and GTTG between cases and controls. In addition, three novel SNPs were identified, two of which were in exons (loc123-e3l-snp2, position 39868005, A/G, Met171Val; RS17511668-SNP2, position 39926432, G/A, Glu118Lys), and one was in the intron6 (RS794001-SNP1, position 39944127, T/G). Moreover, N4BP2 was at higher levels in a majority of tumor tissues examined, relative to paired normal tissues. CONCLUSION: These data suggest that haplotype blocks ATTA and GTTG of N4BP2 is correlation with the risk of sporadic nasopharyngeal carcinoma in the Southern Chinese population and N4BP2 has a potential role in the development of NPC.

[Screening for mutations in the hotspot mutation regions of PIK3CA gene in nasopharyngeal carcinoma].

BACKGROUND & OBJECTIVE: Recent studies showed high frequency of phosphatidylinositol 3-kinase catalytic alpha polypeptide (PIK3CA) mutations in various human cancers; notably, these mutations frequently locate in the hotspot mutation regions of PIK3CA exon 9 and exon 20 with functional significance in tumorigenesis, invasion, and anti-apoptosis. This study was to screen for mutations in the hotspot mutation regions of PIK3CA in nasopharyngeal carcinoma (NPC), and explore the correlation of PIK3CA mutations to tumorigenesis of NPC. METHODS: PIK3CA exon 9 and exon 20 in 46 specimens of sporadic primary NPC tissues were screened by polymerase chain reaction (PCR)-clone sequencing; those in 46 samples of matched NPC peripheral blood and 3 NPC cell lines CNE1, CNE2, and SUNE1 were directly sequenced. RESULTS: Among the 46 specimens of NPC, 2 (4.3%) had point mutation in PIK3CA exon 9 [T1563G (521Asn-->Lys) and A1646G (549Asp-->Gly)], 18 had multiple mutations in PIK3CA exon 9 (A1634C-G1658C-del 1659T), which might be the homologous sequence of Cat Eye Syndrome region on 22q11.2; none had mutation in PIK3CA exon 20. Moreover, no mutation was detected in PIK3CA exon 9 and exon 20 in the 46 matched NPC peripheral blood samples and CNE1, CNE2, and SUNE1 cells. CONCLUSIONS: PIK3CA exon 9 and exon 20 rarely mutate in NPC. Clone sequencing is more sensitive than direct sequencing in screening for somatic mutation. A1634C-G1658C-del 1659T mutations in PIK3CA exon 9, detected by clone sequencing, are supposed to be the homologous sequence of Cat Eye Syndrome region on 22q11.2 instead of mutations in PIK3CA.

Functional advantage of NPC-related V-val subtype of Epstein-Barr virus nuclear antigen 1 compared with prototype in epithelial cell line.

Epstein-Barr virus (EBV) is closely associated with nasopharyngeal carcinoma (NPC) and the viral nuclear antigen 1 (EBNA1) plays a crucial role in viral latency. Three EBNA1 subtypes, P-ala, V-thr and V-val have been detected from healthy carriers in Guangzhou area. A close relation of V-val EBNA1 with NPC was suggested by its preference to infect NPC cells. We therefore investigated the functional difference among these three EBNA1 subtypes in human epithelial cell line. The three coding sequences of the EBNA1 subtypes were cloned into the pGFP-C2 vector, and transfected into 293 cells, respectively. Effect of EBNA1 expression on cell proliferation was examined. The maintenance activity and expression level of EBNA1-plasmid in 293 cells were evaluated by using GFP as a reporter. The expression of P-ala, V-thr or V-val EBNA1 had no effect on 293 cell growth, while the relative average intensity of fluorescence after 14-day selection in V-val-EBNA1/293 cells was statistically higher than P-ala-EBNA1/293 (P<0.05, t test). We suggest that V-val EBNA1 with the functional advantage compared with prototype shown in this study might contribute to the tumorigenesis of NPC by increasing the expression of itself or other viral or cellular genes.