| Summary: | 碩士 === 中山醫學大學 === 醫學分子毒理學研究所 === 93 === The genome of hepatitis delta virus ( HDV ) is composed of a circular single-stranded RNA molecule of 1.7 Kb. The gene of hepatitis delta antigen ( HDAg ) consists of two related proteins. The small form is a 24 KDa ( 195 amino acids ) of S-HDAg, which is essential for replication of HDV RNA genome. The large form is L-HDAg, which is a 27 KDa ( 214 amino acids ), that is essential for particle assembly of HDV RNA. It has been indicated that HDV cDNA contain endogenous promoters both in genomic and antigenomic strands. The position of the antigenomic HDV cDNA promoter-like sequence was located to a 29-nucleotide region, whereas the promoter activity of the genomic HDV cDNA was still unclear. We already knew that antigenomic HDV cDNA has an endogenous promoter activity and is located in TR-P1 fragment, the activity of this promoter is not restricted by its orientation. Another promoter is located in TR-I 1 fragment. In this study, we attempt to identify the essentral promoter sequence of these two HDV cDNA fragments. First, HDV cDNA fragments were inserted into PGL-3-basic vector, and using luciferase as a reporter gene to indentify the HDV cDNA promoter activity. We found that TR-I 5 activity was higher than TR-I 1.The deletion and point mutation analysis in TR-P 1 provided further information about TR-P 1 promoter activity. Finally, we cotransfected L-HDAg or S-HDAg to study HDAg effects on HDV cDNA promoter activity. We observed that L-HDAg and S-HDAg can alter the specific HDV cDNA promoter activity.
Overall, the HDV cDNA sequences containing promoter activity were found and the novel promoter sequence may be used for studying HDV RNA replication or transcription mechanism in the future.
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