Transformation of chicken embryo fibroblasts and tumor induction by the middle T antigen of polyomavirus carried in an avian retroviral vector.

Kornbluth, Sally; Cross, Frederick R.; Harbison, Mary Alice; Hanafusa, Hidesaburô

doi:10.1128/mcb.6.5.1545

Cited by 64 publications

(52 citation statements)

References 45 publications

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“…Infection of CEF with a retrovirus interferes with infection by a second virus unless the two viruses have different subgroup specificities (14). For the double-infection experiments, we therefore produced virus stocks of subgroup A or B by transfecting with the appropriate version of pSR-REP (27). Soft agar colony formation was performed as described previously (33).…”

Section: Methodsmentioning

confidence: 99%

Deletions in the SH2 domain of p60v-src prevent association with the detergent-insoluble cellular matrix.

Fukui¹,

O'Brien²,

Hanafusa³

1991

Mol. Cell. Biol.

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p60v-src has been shown to associate with a detergent-insoluble cellular matrix containing cytoskeletal proteins, but p60csrc does not bind to this matrix. We analyzed the association of mutant src proteins with the matrix and found that mutants which lack an amino-terminal portion (residues 149 to 169) of the SH2 domain cannot bind to the matrix. Neither the SH3 region nor other portions of the SH2 region were required for association. We also tested protein kinase-defective mutants and chimeras of p6O-src and p60csrc. We found a strong correlation between the kinase activity of p6Osrc and its association with the detergent-insoluble matrix. Double infection of kinase-defective and kinase-active mutants did not result in matrix binding of the kinase-defective src proteins. We also found that Tyr-416, the major site of autophosphorylation in p60vsrc, was not required for matrix association.Rous sarcoma virus (RSV) encodes the oncogene v-src. The product of v-src, p60vsrc, is a tyrosine-specific protein kinase (19). p60vsrc is myristoylated on a glycine residue at the amino terminus and localizes in the membrane fraction (19). A number of cellular proteins are phosphorylated on tyrosine upon transformation with p60vsrc. Most of these are also in the membrane fraction (13), suggesting that specific localization of p60v-src is important for cell transformation.Burr et al. (3) have demonstrated association of p60v-src with a detergent-resistant subcellular structure that consists mostly of cytoskeleton. It has been previously reported that most transforming variants of p60vsrc associate with this structure but that nontransforming variants, including the cellular proto-oncogene p60csrc, do not (12,30). However, it is not known which sequences of p60v-src are necessary for its association with this structure.Two regions of sequence conservation have been identified in the amino termini of cytoplasmic tyrosine kinases, including p6Osrc (32, 36). These regions are SH2 (src homology 2, residues 137 through 241 in src; see Fig. lA) and SH3 (src homology 3, residues 84 through 114). SH2 sequences are also present in phospholipase C--y (39), GAP (ras GTPase-activating protein) (43), and the crk oncogene product (32). Mutations of the SH2 regions of v-src and v-fps produce transformation-defective or temperature-sensitive tyrosine kinase proteins (2,5,25,26,36,44). Point mutations in the SH2 region of the proto-oncogene p60c-src have been shown to elevate kinase activity and transforming ability (17,33). Certain SH2 mutations give host-dependent phenotypes (7,8,42), indicating that the SH2 region may interact with host cell proteins. Recently, SH2 sequences have been shown to bind to phosphotyrosine-containing proteins (31).SH3 sequences are present in most proteins containing SH2, as well as in a-spectrin (28), Acanthamoeba myosin-IB (34), and the actin binding protein of Saccharomyces cerevisiae, ABPlp (9). The latter three proteins are all associated with the membrane cytoskeleton, suggesting that the SH3 sequence mediat...

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Section: Methodsmentioning

confidence: 99%

Deletions in the SH2 domain of p60v-src prevent association with the detergent-insoluble cellular matrix.

Fukui¹,

O'Brien²,

Hanafusa³

1991

Mol. Cell. Biol.

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“…In most cases, the foreign DNA sequences were inserted in place of the coding region for src, which is dispensable for virus replication (7,11,15). Other vectors, based on the replication-competent avian spleen necrosis virus, have been constructed and enable the introduction of foreign sequences downstream from the coding sequences for env (8).…”

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confidence: 99%

Construction and properties of replication-competent murine retroviral vectors encoding methotrexate resistance.

1989

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A series of replication-competent Moloney murine leukemia virus vectors was constructed in which each vector contained a mutant dihydrofolate reductase (DHFR) cDNA insert in the U3 region of the viral long terminal repeat. Two of the resulting viruses, MLV (murine leukemia virus) DHFR*-5 and MLV DHFR*-7, were able to stably transfer methotrexate resistance to infected fibroblast cells upon multiple rounds of virus replication and in the absence of drug selection. Cell lines producing recombinant virus with high titers were established, which indicated that the insert did not grossly interfere with viral replication functions. These vectors should be useful for introducing and expressing foreign genes in vivo in tissues and whole animals in which virus spread is needed for efficient infection.

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“…Many studies have shown that PyMT can transform epithelial and endothelial cells leading to hemangiosarcomas, mammary tumors, and other neoplasms of epithelial origin (17,20,22,38). However, leukemia or lymphoma are not a reported consequence of PyMT expression in animal models.…”

Section: Discussionmentioning

confidence: 99%

“…Polyoma middle T (PyMT) is a membrane-associated oncoprotein that can transform endothelial cells and induce hemangiomas in chickens (20) and in mice (21,22). Using SCL-TVA mice and recombinant RCAS/PyMT viruses, we were able to efficiently deliver PyMT to vascular endothelial progenitors and found that the infected mice rapidly succumbed to a fatal hemorrhagic disease due to hemangioma formation.…”

mentioning

confidence: 99%

RCAS/SCL-TVA Animal Model Allows Targeted Delivery of Polyoma Middle T Oncogene to Vascular Endothelial Progenitors In vivo and Results in Hemangioma Development

Sausville

Molinolo²,

Cheng³

et al. 2008

Clinical Cancer Research

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Purpose: To recapitulate the generation of cancer stem cells in the context of an intact animal using a retroviral vector capable of in vivo delivery of oncogenes to primitive endothelial and hematopoietic stem cells. Experimental Design: Targeting of these progenitors was achieved using transgenic mice in which the avian TVA retroviral receptor was placed under the control of the stem cell leukemia (scl/tal-1) gene promoter and SCL +19 enhancer.

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Transformation of chicken embryo fibroblasts and tumor induction by the middle T antigen of polyomavirus carried in an avian retroviral vector.

Cited by 64 publications

References 45 publications

Deletions in the SH2 domain of p60v-src prevent association with the detergent-insoluble cellular matrix.

Deletions in the SH2 domain of p60v-src prevent association with the detergent-insoluble cellular matrix.

Construction and properties of replication-competent murine retroviral vectors encoding methotrexate resistance.

RCAS/SCL-TVA Animal Model Allows Targeted Delivery of Polyoma Middle T Oncogene to Vascular Endothelial Progenitors In vivo and Results in Hemangioma Development

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