Prediction of Promoter Motifs in Virophages / 病毒学报

Virophages have crucial roles in ecosystems and are the transport vectors of genetic materials. To shed light on regulation and control mechanisms in virophage--host systems as well as evolution between virophages and their hosts, the promoter motifs of virophages were predicted on the upstream regions of start codons using an analytical tool for prediction of promoter motifs: Multiple EM for Motif Elicitation. Seventeen potential promoter motifs were identified based on the E-value, location, number and length of promoters in genomes. Sputnik and zamilon motif 2 with AT-rich regions were distributed widely on genomes, suggesting that these motifs may be associated with regulation of the expression of various genes. Motifs containing the TCTA box were predicted to be late promoter motif in mavirus; motifs containing the ATCT box were the potential late promoter motif in the Ace Lake mavirus . AT-rich regions were identified on motif 2 in the Organic Lake virophage, motif 3 in Yellowstone Lake virophage (YSLV)1 and 2, motif 1 in YSLV3, and motif 1 and 2 in YSLV4, respectively. AT-rich regions were distributed widely on the genomes of virophages. All of these motifs may be promoter motifs of virophages. Our results provide insights into further exploration of temporal expression of genes in virophages as well as associations between virophages and giant viruses.

Relationship Between Adeno-Associated Virus and High Risk Human Papilloma Viruses in Cervical Biopsies Using Microdissection Technique / 대한산부인과학회잡지

OBJECTIVE: Adeno associated virus (AAV) is a human DNA virus and is included in the Parvovirus family. AAV has been detected in cervical tissues as well as cervical cancer cell lines. Previous studies showed that AAV infection has some negative effects on HPV infection and that the cervical cancer cell growth is inhibited by AAV infection. The aim of this study is to determine the prevalence of AAV 2 infection and its possible roles for influencing HPV 16 and 18 infection in Korean women by analyzing adjacent normal, CIN, and invasive cervical cancer tissue samples. METHODS: CIN I (20), CIN II (24), CIN III (25), invasive cervical cancer (23) tissues were investigated by microdissection and PCR analyses using primers of HPV 16, 18 and AAV 2 as well as beta- globin as an internal control. RESULTS: AAV 2 was detected in 57 out of 92 cervical lesion biopsies. Among these, mild dysplasia, moderate dysplasia, severe dysplasia and invasive cancer showed 55% (11/20), 95.8% (23/24), 52% (13/25) and 52.2% (12/23), respectively. However, HPV 16 was detected in 14 out of 92 cervical lesion biopsies. Among these, mild dysplasia, moderate dysplasia, severe dysplasia and invasive cancer showed 0% (0/20), 8.3% (2/24), 24% (6/25) and 26.1% (6/23), respectively. HPV 18 was detected in 3 out of 92 cervical lesion biopsies. Among these, mild dysplasia, moderate dysplasia, severe dysplasia and invasive cancer showed 0% (0/20), 4.2% (1/24), 8% (2/25) and 0% (0/23), respectively. In contrast, In 92 perilesional normal biopsies, AAV 2, HPV 16 and HPV 18 were detected to be 57.6% (53/92), 3.3% (3/92) and 0% (0/92), respectively. CONCLUSION: AAV 2 was detected in CIN and invasive cervical cancer biopsies by microdissection and PCR analyses in Korean women. It is difficult to confirm any significant roles of AAV 2 infection for developing cervical cancer. However, we observe that there is some correlation between AAV 2 and HPV infection in the carcinogenesis of cervical cancer. Further research remains to be done to further elucidate AAV 2 infection and its role for HPV infection and cervical cancer.

Constructtion of the Recombinant pAAVCMVp53 for Cervical Cancer Gene therapy / 대한산부인과학회잡지

OBJECTIVE: Human papillomavirus (HPV) has been identified in the majority of invasive cervical cancer patient and has been found to contribute in a significant way to the genesis of human cervical cancer. HPV has two transforming genes that encode the oncoproteins E6 and E7, E6 can form complexes with p53 and promote p53 degradation, E7 inhibit retinoblastoma protein (RB). The p53 protein is as a phosphoprotein which co-immunoprecipitated with the SV40 T-Antigen. The wild type p53 protein is capable of suppressing the tumorigenic phenotype and regulating cell cycle. Adeno-associated virus(AAV) is a linear single stranded DNA parvovirus which is dependent upon cotransfection by a second unrelated virus to undergo productive infection. It has been well documented that AAV DNA integrates into cellular DNA as one to several tandem copies joined to cellular DNA through the termini. In order to introduce wild type p53 through AAV virus into a cervical cancer patient for gene therapy, we had constructed recombinant p53 adeno associated virus plasmid (pAAVCMVp53). METHODS: pAAVCMVp53 was created new AAV-vector system, pRc/CMVp53 including p53 cDNA and AAV-derivative vector, pASPA-AAV-CMV-polyA were made to HindIII/blunt fragments. Eluated 1.8 kb fragment of wild type p53 cDNA was ligated to pAAV-CMV-polyA, 4.9 kb fragment deprived hASPA cDNA. RESULT: Recombinant AAVCMVp53 was constructed by using pRc/CMVp53 andpASPA-AAV-CMV-polyA. This pAAVCMVp53 was confirmed by various restriction enzyme-digestions and Southern-blotting. This new vector system will be studied on expression, stability in cervical cancer cell lines and animals. CONCLUSION: This system will be one of the useful vector system for cervical cancer gene therapy.