An investigation of the effect of antisense RNA gene on bovine leukaemia virus reproduction in cell culture

Murovska, Modra; Chernobayeva, L. G.; Miroshnichenko, Olga I.; Tomsons, V. P.; Konicheva, V. V.; Иванова, С В; Tikhonenko, T I

doi:10.1016/0378-1135(92)90063-y

Cited by 6 publications

(3 citation statements)

References 10 publications

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“…Demonstration that this ribozyme or antisense (14,28,29) inhibits viral expression in persistently infected cells suggests possibilities for BLV control. BLV is a blood-borne disease primarily transmitted mechanically by vectors and fomites (30).…”

Section: Discussionmentioning

confidence: 99%

Ribozyme cleaves rex/tax mRNA and inhibits bovine leukemia virus expression.

Cantor

McElwain

Birkebak

et al. 1993

Proc. Natl. Acad. Sci. U.S.A.

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Bovine leukemia virus (BLV) encodes at least two regulatory proteins, Rex and Tax. Tax, the transactivating protein, stimulates the long terminal repeat to promote viral transcription and may be involved in tumorigenesis. Rex is involved in the transition from early expression of regulatory proteins to later expression of viral structural proteins. We have targeted ribozymes against the mRNA encoding Rex and Tax. The ribozymes consist of the hammerhead catalytic motif flanked by antisense sequences that hybridize with the complementary rex/tax mRNA. To evaluate cleavage in a cell-free system, we transcribed portions of rex/tax mRNA and incubated them with synthetic RNA ribozymes. A ribozyme was identified that cleaves >80% of the target RNA. Synthetic DNA encoding this ribozyme was cloned into the expression vector pRc/RSV and transfected into BLV-infected bat lung cells. Intracellular cleavage of rex/tax mRNA was confirmed by reverse transcriptase PCR. In cells expressing the ribozyme, viral expression was markedly inhibited. Expression of the BLV core protein p24 was inhibited by 61%, and reverse transcriptase activity in supernatant was inhibited by 92%. Ribozyme inhibition of BLV expression suggests that cattle expressing these sequences may be able to control BLV replication.Bovine leukemia virus (BLV), a retrovirus structurally similar to human T-cell leukemia viruses I and II (HTLV-I and -II), causes persistent lymphocytosis and B-lymphocyte lymphoma in cattle and sheep (1). After initial infection, BLV expresses a doubly spliced transcript encoding the regulatory proteins Rex and Tax (2). Tax trans-activates the viral long terminal repeat and also cellular promoters, including c-fos and somatostatin (3). Cotransfection experiments have shown that Tax is necessary for viral expression in vitro (4). Rex regulates the transition from early expression of the doubly spliced transcript encoding regulatory proteins to the later expression of singly spliced or unspliced transcripts that express the env gene or the gag and pol genes (5). Recently, the 3' region of HTLV-I and BLV has been shown to encode RNA with alternative splice patterns that may express other regulatory proteins (6-8).Because of the critical role of regulatory proteins such as Tax and Rex in the HTLV/BLV group of viruses, we hypothesized that the rex/tax mRNA would be a rational target for ribozyme-mediated inhibition. The hammerhead motif, first identified in plant RNA pathogens (9, 10), cleaves the phosphodiester bond downstream of a GUC triplet (9,10). By flanking the hammerhead motif with antisense sequences, Haseloff and Gerlach (11) demonstrated cleavage of specific target RNAs. In this study, we demonstrate a ribozyme that cleaves rex/tax mRNA and, when transfected into BLV-infected cells, markedly inhibits viral expression.The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact...

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Section: Discussionmentioning

confidence: 99%

Ribozyme cleaves rex/tax mRNA and inhibits bovine leukemia virus expression.

Cantor

McElwain

Birkebak

et al. 1993

Proc. Natl. Acad. Sci. U.S.A.

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“…For certain diseases, such as FMD, vaccination is not allowed in many countries except under very specific circumstances, such as containment of an active outbreak. In such scenarios, antivirals might be used to provide immediate protection Borisenko et al (1992) Antisense RNA Inhibition of BLV replication by targeting the gene X Murovska et al (1992)…”

Section: Discussionmentioning

confidence: 99%

Potential applications for antiviral therapy and prophylaxis in bovine medicine

Newcomer

Walz

Givens

2014

Anim. Health. Res. Rev.

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Viral disease is one of the major causes of financial loss and animal suffering in today's cattle industry. Increases in global commerce and average herd size, urbanization, vertical integration within the industry and alterations in global climate patterns have allowed the spread of pathogenic viruses, or the introduction of new viral species, into regions previously free of such pathogens, creating the potential for widespread morbidity and mortality in naïve cattle populations. Despite this, no antiviral products are currently commercially licensed for use in bovine medicine, although significant progress has been made in the development of antivirals for use against bovine viral diarrhea virus (BVDV), foot and mouth disease virus (FMDV) and bovine herpesvirus (BHV). BVDV is extensively studied as a model virus for human antiviral studies. Consequently, many compounds with efficacy have been identified and a few have been successfully used to prevent infection in vivo although commercial development is still lacking. FMDV is also the subject of extensive antiviral testing due to the importance of outbreak containment for maintenance of export markets. Thirdly, BHV presents an attractive target for antiviral development due to its worldwide presence. Antiviral studies for other bovine viral pathogens are largely limited to preliminary studies. This review summarizes the current state of knowledge of antiviral compounds against several key bovine pathogens and the potential for commercial antiviral applications in the prevention and control of several selected bovine diseases.

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“…As shown by WILLEMS et al (1992), the N-terminal region of BLV-tax forming a putative zinc finger structure is important for LTR-directed transcription. Furthermore, recent experimental data concerning the relationship between tax and BLVantigen synthesis have been published, which show that the blocking of tax/rexm R N A with appropriate oligonucleotides leads to a significant decrease of BLVantigen synthesis in FLK/BLV-cells (MUROVSKA et al, 1992). In productive BLVinfected bat cells, the antigen synthesis was dramatically reduced as a result of cleaving the tax/rex-mRNA by a hammerhead ribozyme linked with a tax/rex-antisense RNA (CANTOR et al, 1993).…”

Section: Discussionmentioning

confidence: 99%

Increase of Antigen Production in BLV‐infected Cell Lines via Additional Expression of Tax

Wagner

Blankenstein

Bondzio

et al. 1995

Journal of Veterinary Medicine, Series B

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Summary The selection of animals infected with the bovine leukaemia virus (BLV) is performed by the immunological detection of antibodies against the virus, commonly using the antigen gp51. Furthermore, research is being carried out to develop protective vaccines against BLV that have gp51 as their main component. Taking both of these factors into account, it is clear that there will be an increasing requirement for the virus antigen gp51 for some time to come. The permanently BLV‐infected foetal lamb kidney cell line FLK/BLV (and its sublines) has been proved to be the most useful culture for the mass production of the virus antigen. Stable cell lines producing higher quantities of BLV antigen have not been established, either by subcloning of the FLK/BLV or by infection of other permanent cells with BLV. Here, a report is made on efforts to increase the expression of gp51 in BLV‐infected cells via the additional expression of homologous transactivating virus protein tax. Selectable tax expression vectors that integrate into the host cell genome were constructed using BL provirus DNA fragments. Highly productive FLK/BLV cells were transfected with these vectors. Following selection with G 418, gp51‐producing cell lines were established and tested for their productivity for several months. Some tax‐vector‐containing cell lines have produced 1.5‐2 times more gp51 than the highly productive parental control cell line FLK/BLV 44‐1.

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An investigation of the effect of antisense RNA gene on bovine leukaemia virus reproduction in cell culture

Cited by 6 publications

References 10 publications

Ribozyme cleaves rex/tax mRNA and inhibits bovine leukemia virus expression.

Ribozyme cleaves rex/tax mRNA and inhibits bovine leukemia virus expression.

Potential applications for antiviral therapy and prophylaxis in bovine medicine

Increase of Antigen Production in BLV‐infected Cell Lines via Additional Expression of Tax

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