Recombinant retroviral DNA yielding high expression of hepatitis B surface antigen.

Stratowa, C; Doehmer, Johannes; Wang, Y.; Hofschneider, Peter Hans

doi:10.1002/j.1460-2075.1982.tb01357.x

Cited by 30 publications

(16 citation statements)

References 33 publications

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“…After 7 days an amount of 80 ng of HBsAg per ml was found (27). This is about 20 times lower than the yield of 1.7 ,ug/ml obtained from Y1 cells after 7 days.…”

Section: Resultsmentioning

confidence: 58%

See 1 more Smart Citation

Enhanced production of hepatitis B surface antigen in NIH 3T3 mouse fibroblasts by using extrachromosomally replicating bovine papillomavirus vector.

Wang¹,

Stratowa²,

Schaefer-Ridder³

et al. 1983

Mol. Cell. Biol.

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We have constructed a recombinant pBR322 plasmid composed of a subgenomic transforming fragment of bovine papillomavirus DNA and the hepatitis B surface antigen gene from cloned hepatitis B virus DNA and used it for transfection of NIH 3T3 mouse fibroblasts. The transformed cells retain the plasmids in extrachromosomal form with a copy number of about 50 to 100 per cell. Expression of the hepatitis B surface antigen gene linked to bovine papillomavirus DNA is independent of its orientation relative to the bovine papillomavirus vector. Cell lines continuously secreting high amounts of hepatitis B surface antigen into the medium could be established. The antigen is released into the culture medium as 22-nm particles, having the same physical properties and constituent polypeptides as those found in the serum of hepatitis B virusinfected patients.

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“…After 7 days an amount of 80 ng of HBsAg per ml was found (27). This is about 20 times lower than the yield of 1.7 ,ug/ml obtained from Y1 cells after 7 days.…”

Section: Resultsmentioning

confidence: 58%

“…We have recently used the integrated proviral form of Moloney mouse sarcoma virus to introduce an HBV fragment containing the HBsAg gene and its regulation signals into mouse fibroblasts (27).…”

Section: Received 1 December 1982/accepted 7 March 1983mentioning

confidence: 99%

Enhanced production of hepatitis B surface antigen in NIH 3T3 mouse fibroblasts by using extrachromosomally replicating bovine papillomavirus vector.

Wang¹,

Stratowa²,

Schaefer-Ridder³

et al. 1983

Mol. Cell. Biol.

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“…Details of the templates used in this work have been presented by Standring et al (34) and are summarized below. Nucleotide numbering of the HBV genome is given relative to the EcoRI site as the zero point (0/3,221) in accordance with the generally accepted system (11) and differs from that presented previously (35), in which the EcoRI site occurs at nucleotide 1,404. The plasmid containing the adenovirus major late promoter (pSmaF) was obtained from R. Roeder.…”

mentioning

confidence: 99%

Transcription of hepatitis B virus by RNA polymerase II.

Rall¹,

Standring²,

Laub³

et al. 1983

Mol. Cell. Biol.

119

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We employed an in vitro cell-free transcription system to locate RNA polymerase II promoters on the hepatitis B virus genome. The strongest promoter precedes the surface antigen (HBsAg) gene, which is comprised of a long (500 base pairs) presurface region as well as the mature HBsAg coding sequence. The origin of this transcript was localized by using truncated templates and S1 endonuclease.mapping. The activity of the promoter was confirmed in transfection experiments in which the complete HBsAg gene was introduced into monkey kidney cells via a simian virus 40 expression vector. A second RNA polymerase II promoter preceding the HBcAg gene was also active in the cell-free system. The presence of multiple promoters in the hepatitis B virus genome suggests that the relative levels of viral-specific proteins detected in liver and serum may reflect differential or regulated promoter efficiency.Hepatitis B virus (HBV) is a partially singlestranded DNA virus whose host range is restricted to humans and chimpanzees (reviewed in reference 37). The complete nucleotide sequences of the cloned HBV (11,27,38,40) and the related woodchuck hepatitis virus (10) define the major viral protein coding regions. The organization of genetic information is compact: four reading frames (A, B, C, and S) are found in the viral long strand and one small open reading frame (D) is found in the short strand. The viral surface (S) and core (C) protein coding sequences have been identified and incorporated into both procaryotic (9, 27) and eucaryotic (8, 19a, 25, 32, 35) expression vectors, generating products that react with antibodies directed against these antigens (HBsAg and HBcAg). The predicted sequences suggest that the surface antigen is a transmembrane protein, whereas on the basis of the protamine-like COOH terminal region (40), the core antigen is a DNAbinding protein. The S gene has a long 5' leader sequence and the C gene has a shorter 5' leader upstream from the mature coding regions, suggesting that both proteins could be synthesized as precursors. Of the remaining two long-strand open reading frames, the larger (A, -80% of the viral genome) presumably encodes the viral DNA polymerase. This gene overlaps part of the C gene, all of the S gene, and part of the B gene but would be translated with a different register. No specific protein has yet been associated with the smaller reading frame (B).This sequence information has not resulted in a detailed characterization of HBV promoters of RNA transcripts, mainly due to the lack of a reproducible tissue culture system for viral propagation. The definition of HBV promoters is important for (i) determining how the viral genetic information is expressed, (ii) assessing a possible oncogenic role of integrated HBV sequences, and (iii) studying replication, which has recently been proposed to involve reverse transcription of a full-length viral RNA transcript (36). The HBV DNA sequence provides some information on potential transcription signals such as TATA boxes and AATAAA-directed p...

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“…The amount of HBsAg produced by BKV vectors is comparable to other eukaryotic vectors systems, e.g. bovine papillomavirus (BPV), murine sarcoma virus and simian virus 40 (SV40) (Stratowa et al, 1982;Denniston et al, 1984;Will et al, 1984). The orientation of the HBsAg gene relative to the BKV sequences, the type of HBsAg gene promoter, the HBV subtype as well as the tissue origin of the human cell clones did not determine major differences in copy number or level of expression of the HBsAg gene using different BKV plasmid constructs.…”

Section: Discussionmentioning

confidence: 83%

Expression of Hepatitis B Surface Antigen in Human Cells by a Recombinant BK Virus DNA Vector

Caputo

Barbanti-Brodano

Reschiglian

et al. 1988

Journal of General Virology

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Recombinant retroviral DNA yielding high expression of hepatitis B surface antigen.

Cited by 30 publications

References 33 publications

Enhanced production of hepatitis B surface antigen in NIH 3T3 mouse fibroblasts by using extrachromosomally replicating bovine papillomavirus vector.

Enhanced production of hepatitis B surface antigen in NIH 3T3 mouse fibroblasts by using extrachromosomally replicating bovine papillomavirus vector.

Transcription of hepatitis B virus by RNA polymerase II.

Expression of Hepatitis B Surface Antigen in Human Cells by a Recombinant BK Virus DNA Vector

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