Epstein-Barr virus latent membrane protein 2A promotes invasion of nasopharyngeal carcinoma cells through ERK/Fra-1-mediated induction of matrix metalloproteinase 9.

Epstein-Barr virus (EBV)-associated nasopharyngeal carcinoma (NPC) is highly metastatic, and this malignant feature may be promoted by an EBV oncoprotein, latent membrane protein 2A (LMP2A). Acting as a signal regulator, LMP2A can enhance invasiveness and motility of epithelial cells. Downstream from the LMP2A-triggered signaling events, it is largely unknown what key effector proteins are induced and essentially promote cell invasion. In the present study, we found that in NPC cells, LMP2A upregulated matrix metalloproteinase 9 (MMP9), a metastasis-associated protease. LMP2A increased MMP9 expression at both the mRNA and protein levels. It also activated the MMP9 promoter, in which two AP-1 elements were required for the promoter activation. Among AP-1 transcription factors, Fra-1 was induced by LMP2A and is essential for LMP2A-triggered MMP9 expression. Induction of Fra-1 was dependent on the LMP2A-activated ERK1/2 pathway, and induction of the ERK1/2-Fra-1-MMP9 axis required PY motifs in the amino-terminal domain of LMP2A. Notably, LMP2A-promoted invasion of NPC cells was blocked when MMP9 expression, Fra-1 induction, or ERK1/2 activation was inhibited. In addition, we found an association of LMP2A with MMP9 expression in NPC tumor biopsy specimens, where Fra-1 was a major mediation factor. This study reveals an underlying mechanism of LMP2A-induced cell invasion, from signal transduction to upregulation of a critical protease. Considering that MMP9 can also be upregulated by another EBV oncoprotein, LMP1, this protease may be a pivotal effector at which the EBV-induced, invasion-promoting mechanisms converge, serving as an attractive therapeutic target for NPC treatment.

Association of NRAMP 1 gene polymorphism with susceptibility to tuberculosis in Taiwanese aboriginals.

BACKGROUND/PURPOSE: The human homologue of mice natural-resistance-associated macrophage protein 1 (Nramp 1) gene, NRAMP 1, has been reported to play a role in susceptibility to tuberculosis in humans. The aboriginal population in Taiwan has a five-fold higher prevalence of tuberculosis than people of Han ethnicity. Whether genetic factors such as NRAMP 1 polymorphism play a role in the prevalence of tuberculosis in Taiwanese aboriginals should be clarified. METHODS: NRAMP 1 polymorphism was studied using a case-control design of patients with tuberculosis, including subjects of Han (Hans) and aboriginal ethnicity in Hualien, eastern Taiwan. The polymorphisms of NRAMP 1 at loci INT4, D543N, 77-385C/T, 3-UTR (CAAA) deletion and 5-(CA)n microsatellite markers were assessed by polymerase chain reaction on tissue DNA isolated from 105 aborigines and 110 Hans with tuberculosis. Comparable numbers of ethnically-matched controls were studied simultaneously. RESULTS: Two NRAMP 1 polymorphisms, INT4 and 5-(CA)n, were significantly associated with susceptibility to tuberculosis in aboriginals (p = 0.0070 and p = 0.0031, respectively). However, no association was detected at the five loci of NRAMP 1 polymorphisms among Hans (p > 0.08). CONCLUSION: Genetic variation in NRAMP 1 may affect susceptibility to and increase risk for tuberculosis in Taiwanese aboriginals. Although environmental factors play an important role in tuberculosis infection, genetic factors such as NRAMP 1 polymorphism may also contribute to the high prevalence of tuberculosis in Taiwanese aboriginals.

Presence of DAZL transcript and protein in mature human spermatozoa.

OBJECTIVE: To identify the DAZL transcript and protein location in human spermatozoa. DESIGN: In vitro experiment. SETTING: University-based reproductive genetics laboratory. PATIENT(S): A fertile volunteer. INTERVENTION(S): Reverse transcription-polymerase chain reaction (RT-PCR), Western blot, and immunostaining for DAZL. MAIN OUTCOME MEASURE(S): Expression of DAZL in human spermatozoa. RESULT(S): The DAZL-specific primers yield a 128 bp product in ejaculate. A protein of approximately 33.5 kDa was detected by Western blot analysis. Immunofluorescence staining showed strong homogeneous staining in the midpiece of spermatozoa and weak staining in the principal piece. A speckled-type distribution was found in the head region. CONCLUSION(S): The DAZL transcript and protein are present in human spermatozoa. The roles of DAZL protein in sperm motility and in the sperm-oocyte interaction await further investigation.

Human papilloma virus detection in neoplastic and non-neoplastic nasopharyngeal tissues in Taiwan.

BACKGROUND: Human papilloma virus (HPV) has been implicated in the carcinogenesis and prognosis of certain head and neck cancers. Whether it also has a role in the pathogenesis of nasopharyngeal carcinoma (NPC) in Taiwan is unclear. METHODS: Detection and genotyping of HPVs were performed in 43 primary NPCs (one WHO-I and 42 WHO-II/III) and 40 nasopharyngeal controls using PCR-based HPV genotyping arrays. Localisation of high-risk HPV and Epstein-Barr virus genomes was performed in another 46 primary NPCs (five WHO-I and 41 WHO-II/III) and seven paired metastatic WHO-II/III NPCs using in situ hybridisation. RESULTS: In the HPV genotyping cohort, oncogenic HPVs were detected equally in WHO-II/III NPCs (31%, 13/42) and nasopharyngeal controls (35%, 14/40). Tumour high-risk HPV status did not correlate with the prognosis of patients with NPC. In the high-risk HPV in situ hybridisation cohort, 14 (88%) of the 16 oncogenic HPV-positive WHO-II/III NPCs showed a unique cytoplasmic/perinuclear staining pattern, which is distinct from the typical dot/punctate nuclear staining pattern indicating HPV genome integration. In addition, oncogenic HPVs were not always retained in NPC cells during the process of metastasis. CONCLUSIONS: This study does not support an association between oncogenic HPV and the carcinogenesis or prognosis of WHO-II/III NPCs in Taiwan.

Endogenous latent membrane protein 1 in Epstein-Barr virus-infected nasopharyngeal carcinoma cells attracts T lymphocytes through upregulation of multiple chemokines.

Tumor-infiltrating T lymphocytes are considered to facilitate development of Epstein-Barr virus (EBV)-associated nasopharyngeal carcinoma (NPC), but how EBV in NPC tumor cells directs T cell infiltration remains unclear. Here we compare EBV-infected NPC cells with and without spontaneous expression of viral latent membrane protein 1 (LMP1) and find that culture supernatants of LMP1-positive NPC cells exert enhanced chemoattraction to primary T cells. Knockdown of endogenous LMP1 in the cells suppresses the chemotactic activity. Endogenous LMP1 in NPC cells upregulates multiple chemokines, among which MIP-1alpha, MIP-1beta and IL-8 contribute to T cell chemotaxis. We further reveal that LMP1-induced production of MIP-1alpha and MIP-1beta in NPC cells requires not only two carboxyl-terminal activation regions of LMP1 but also their downstream NF-kappaB and JNK pathways. This study corroborates that endogenous LMP1 in EBV-infected NPC cells induces multiple chemokines to promote T cell recruitment and perhaps other pathogenic events in NPC.

Endoplasmic reticulum stress triggers XBP-1-mediated up-regulation of an EBV oncoprotein in nasopharyngeal carcinoma.

Endoplasmic reticulum (ER) stress-activated unfolded protein response (UPR) plays multiple roles in cancer development, but its specific roles for virus-associated cancers have not been fully understood. It is still unknown whether ER stress/UPR occurs in and contributes to nasopharyngeal carcinoma (NPC), an epithelial malignancy closely associated with EBV. Here, we report that UPR proteins are frequently detected in NPC biopsies. In addition, we reveal that, in EBV-infected NPC cells, ER stress inducers up-regulate a potent EBV oncoprotein latent membrane protein 1 (LMP1), and the ER stress-induced LMP1 enhances production of interleukin-8. ER stress triggers LMP1 expression at a transcriptional level, activating a distal LMP1 promoter TR-L1. TR-L1 contains an ER stress-responsive element, which is targeted by an UPR protein XBP-1. Ectopic expression of XBP-1 induces LMP1 expression, and knockdown of XBP-1 blocks ER stress-triggered up-regulation of LMP1 in NPC cells. Furthermore, XBP-1 significantly correlates with LMP1 expression in NPC tumor biopsies. Therefore, this study not only provides a novel clue linking ER stress/UPR to EBV-associated NPC but also suggests that ER stress/UPR can promote virus-associated cancer in a unique way by driving expression of a viral oncogene.

Induction of the early growth response 1 gene by Epstein-Barr virus lytic transactivator Zta.

Early growth response 1 (Egr-1) is a cellular transcription factor involved in diverse biologic functions. Egr-1 has been associated with Epstein-Barr virus (EBV) infection, but it is still unknown whether any EBV protein regulates Egr-1 expression. In this study, we first showed that EBV reactivation is involved in upregulation of Egr-1 and that Egr-1 can be induced by Zta, an EBV lytic transactivator. Zta not only binds to the Egr-1 promoter but also activates the ERK signaling pathway to trigger binding of Elk-1 to the Egr-1 promoter. In addition, knockdown of Egr-1 significantly reduces the spontaneous expression of Zta and Rta in EBV-infected 293 cells, suggesting that a positive-feedback network involving Egr-1 is required for EBV reactivation. This study also implies that Zta has the potential to affect expression of certain genes through Egr-1.