In vivo transfection of bovine leukemia provirus into sheep

Willems, Luc; Portetelle, Daniel; Kerkhofs, Pierre; Gao, Chen; Burny, Arsène; Mammerickx, Marc; Kettmann, Richard

doi:10.1016/0042-6822(92)90604-n

Cited by 78 publications

(100 citation statements)

References 8 publications

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“…This virus is not infectious in vivo when injected into sheep. In contrast, when a functional tax gene is provided, the recombinant provirus becomes infectious and replicates at wild-type levels in vivo (Willems et al, 1993). Since the proviruses recombinant for the Tax phosphorylation sites 106 and 293 propagate at wildtype levels in vivo, it appears that essential genes such as Tax and Rex are not aected by these mutations.…”

Section: Discussionmentioning

confidence: 95%

“…This observation provides a strong argument for the absence of phosphorylation requirements to ensure transactivation. Indeed, we have previously shown that the infectious potential of BLV proviruses requires a functional Tax protein (Willems et al, 1993). In fact, a provirus (clone 395) exhibits a tenfold reduction in transactivation activity due to the presence of two mutations in the tax gene (two glutamine codons are substituted with lysine residues).…”

Section: Discussionmentioning

confidence: 99%

“…Indeed, the transactivation of the LTR promoter is not required for Tax to transform these cells (Willems et al, 1992;Yamaoka et al, 1996). Finally, Tax is an essential gene for viral replication since BLV recombinant viruses that lack transactivation potential are not infectious in vivo (Willems et al, 1993). It thus appears that the Tax protein plays a key role in the oncogenic potential and in the replication of the virus.…”

Section: Introductionmentioning

confidence: 99%

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Phosphorylation of bovine leukemia virus Tax protein is required for in vitro transformation but not for transactivation

Willems

Grimonpont

Kerkhofs³

et al. 1998

Oncogene

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The Tax proteins of the oncovirinae viruses are phosphorylated transcriptional activators that exhibit oncogenic potential. The role of phosphorylation in their functional activities remains unknown. As a model for the Human T-cell leukemia virus type I (HTLV-I), Bovine Leukemia Virus (BLV) permits the characterization of viral replication and leukemogenesis in vivo. Here, we show that the BLV Tax protein is phosphorylated on serine residues 106 and 293 both in insect and in mammalian cells. These sites can also be eciently phosphorylated by the cdc2 and MAP kinases in vitro. Mutation of these residues does not aect the capacity of the Tax protein to function as a transactivator. Indeed, the Tax proteins mutated at one or both serines increase LTR-directed viral transcription at levels similar to those obtained with wild-type Tax in cell culture. Moreover, inhibition of Tax phosphorylation by W7, a calmodulin antagonist, does not alter its transactivation activity. Thus, phosphorylation on serines 106 and 293 is not required for transactivation by Tax. However, simultaneous substitution of both serines into alanine residues destroys the capacity of Tax to cooperate with the Haras oncogene to transform primary rat embryo ®bro-blasts and induce tumors in nude mice. When the serines were replaced with aspartic acid residues, the oncogenic potential of Tax was maintained indicating that the negative charge rather than the phosphate group itself was required for Tax oncogenicity. Finally, to assess the role of the serine residues in vivo, recombinant viruses which express the Tax mutants were constructed and injected into sheep. It appeared that the mutated proviruses replicate at levels similar to the wild-type virus in vivo. We conclude that Tax phosphorylation is dispensable for transactivation and viral replication in vivo but is required for its oncogenic potential in vitro.

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Section: Discussionmentioning

confidence: 95%

Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Phosphorylation of bovine leukemia virus Tax protein is required for in vitro transformation but not for transactivation

Willems

Grimonpont

Kerkhofs³

et al. 1998

Oncogene

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“…Three sheep (nos. 8, 105, and 293) were persistently infected with wild-type pBLVIX (29), whereas animals 104, 2658, 2667, and 2668 harbored provirus pBLVTax106 ϩ 293, pLTR-NF1, pLTR-NF2, and pLTR-Ebox, respectively (31,32). Importantly, all proviruses behaved as wild type during pathogenesis, despite the presence of some mutations.…”

Section: Methodsmentioning

confidence: 97%

Increased cell proliferation, but not reduced cell death, induces lymphocytosis in bovine leukemia virus-infected sheep

Debacq

Asquith

Kerkhofs

et al. 2002

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

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Lymphocyte homeostasis is the result of a critical balance between cell proliferation and death. Disruption of this subtle equilibrium can lead to the onset of leukemia, an increase in the number of lymphocytes being potentially due to both of these parameters. The relative importance of cell proliferation vs. apoptosis during pathogenesis induced by the primate T cell lymphotropic viruses and bovine leukemia virus (BLV) has been difficult to assess because of conflicting data from a range of in vitro and ex vivo experimental systems. Here, we aim to resolve this issue by measuring the rates of cell proliferation and death in the BLV-ovine system, an animal model of human T lymphotropic virus (HTLV-1). We use a method based on the i.v. injection of 5-bromodeoxyuridine into BLV-infected sheep. We show that B lymphocytes in BLV ؉ asymptomatic sheep proliferate significantly faster than in uninfected controls (average proliferation rate: 0.020 per day vs. 0.011 per day). In contrast, the rates of cell death were not significantly different between aleukemic BLV-infected and control sheep (average death rate 0.089 per day vs. 0.094 per day, respectively). We conclude that the increase in the number of B cells during BLVinduced lymphocytosis results from higher proliferation rates but is not due to a significant decrease in apoptosis, in contrast to data from in vitro (ex vivo) experiments. The imbalance created by the net increase in proliferation in the absence of compensating cell death reveals a complex mechanism of feedback regulation controlling homeostasis in the blood compartment.

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“…These viral miRNAs result from the transcription of five independent transcriptional units located in a particular BLV proviral region found to be dispensable for in vivo infectivity and considered functionally irrelevant thus far (2,15,16). While we were preparing this manuscript, Kincaid et al (17) reported that BLV was capable of encoding miRNAs in vitro.…”

mentioning

confidence: 99%

Deep sequencing reveals abundant noncanonical retroviral microRNAs in B-cell leukemia/lymphoma

Rosewick

Momont

Durkin

et al. 2013

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

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103

135

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Viral tumor models have significantly contributed to our understanding of oncogenic mechanisms. How transforming delta-retroviruses induce malignancy, however, remains poorly understood, especially as viral mRNA/protein are tightly silenced in tumors. Here, using deep sequencing of broad windows of small RNA sizes in the bovine leukemia virus ovine model of leukemia/lymphoma, we provide in vivo evidence of the production of noncanonical RNA polymerase III (Pol III)-transcribed viral microRNAs in leukemic B cells in the complete absence of Pol II 5′-LTR-driven transcriptional activity. Processed from a cluster of five independent self-sufficient transcriptional units located in a proviral region dispensable for in vivo infectivity, bovine leukemia virus microRNAs represent ∼40% of all microRNAs in both experimental and natural malignancy. They are subject to strong purifying selection and associate with Argonautes, consistent with a critical function in silencing of important cellular and/or viral targets. Bovine leukemia virus microRNAs are strongly expressed in preleukemic and malignant cells in which structural and regulatory gene expression is repressed, suggesting a key role in tumor onset and progression. Understanding how Pol III-dependent microRNAs subvert cellular and viral pathways will contribute to deciphering the intricate perturbations that underlie malignant transformation.noncoding RNA | oncogenesis | retrovirus silencing | HTLV-1

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In vivo transfection of bovine leukemia provirus into sheep

Cited by 78 publications

References 8 publications

Phosphorylation of bovine leukemia virus Tax protein is required for in vitro transformation but not for transactivation

Phosphorylation of bovine leukemia virus Tax protein is required for in vitro transformation but not for transactivation

Increased cell proliferation, but not reduced cell death, induces lymphocytosis in bovine leukemia virus-infected sheep

Deep sequencing reveals abundant noncanonical retroviral microRNAs in B-cell leukemia/lymphoma

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