scholarly journals In vitroassembly of the Rous Sarcoma Virus capsid protein into hexamer tubes under physiological conditions

2017 ◽  
Author(s):  
Soumeya A. Jaballah ◽  
Graham D. Bailey ◽  
Ambroise Desfosses ◽  
Jaekyung Hyun ◽  
Alok K. Mitra ◽  
...  
Keyword(s):  
Capsid Protein ◽  
Rous Sarcoma Virus ◽  
Tube Formation ◽  
Maturation Process ◽  
Rous Sarcoma ◽  
C Terminus ◽  
Sarcoma Virus ◽  
Tube Assembly ◽  
A Minor

ABSTRACTDuring a proteolytically-driven maturation process, the ortho-retroviral capsid protein (CA) assembles to form the convex shell that surrounds the viral genome. In some orthoretroviruses, including Rous Sarcoma Virus (RSV), CA carries a short and hydrophobic spacer peptide (SP) at its C-terminus early in the maturation process, which is progressively removed as maturation proceeds. In this work, we show that RSV CA assemblesin vitroat physiological temperatures, forming hexamer tubes that effectively model the mature capsid surface. Tube assembly is strongly influenced by electrostatic effects, and is a nucleated process that remains thermodynamically favored at lower temperatures, but is effectively arrested by the large Gibbs energy barrier associated with nucleation. RSV CA tubes are multi-layered, being formed by nested and concentric tubes of capsid hexamers. However the spacer peptide acts as a layering determinant during tube assembly. If only a minor fraction of CA-SP is present, multi-layered tube formation is blocked, and single-layered tubes predominate. This likely prevents formation of biologically aberrant multi-layered capsids in the virion. The generation of single-layered hexamer tubes facilitated 3D helical image reconstruction from cryo-electron microscopy data, revealing the basic tube architecture.

scholarly journals In vitro assembly of the Rous Sarcoma Virus capsid protein into hexamer tubes at physiological temperature

2017 ◽  
Vol 7 (1) ◽  
Author(s):  
Soumeya A. Jaballah ◽  
Graham D. Bailey ◽  
Ambroise Desfosses ◽  
Jaekyung Hyun ◽  
Alok K. Mitra ◽  
...  
Keyword(s):  
Capsid Protein ◽  
Rous Sarcoma Virus ◽  
Virus Capsid ◽  
Physiological Temperature ◽  
Rous Sarcoma ◽  
In Vitro Assembly ◽  
Sarcoma Virus ◽  
Virus Capsid Protein

Functional domains of the pp60v-src protein as revealed by analysis of temperature-sensitive Rous sarcoma virus mutants

1984 ◽  
Vol 4 (8) ◽  
pp. 1508-1514
Author(s):  
A W Stoker ◽  
P J Enrietto ◽  
J A Wyke
Keyword(s):  
Rous Sarcoma Virus ◽  
Kinase Activity ◽  
Temperature Sensitive ◽  
Tyrosine Kinase Activity ◽  
Rous Sarcoma ◽  
Heat Labile ◽  
Src Gene ◽  
Sarcoma Virus

Four temperature-sensitive (ts) Rous sarcoma virus src gene mutants with lesions in different parts of the gene represent three classes of alteration in pp60src. These classes are composed of mutants with (i) heat-labile protein kinase activities both in vitro and in vivo (tsLA27 and tsLA29), (ii) heat-labile kinases in vivo but not in vitro (tsLA33), and (iii) neither in vivo nor in vitro heat-labile kinases (tsLA32). The latter class indicates the existence of structural or functional pp60src domains that are required for transformation but do not grossly affect tyrosine kinase activity.

Phenotypic change from transformed to normal induced by benzoquinonoid ansamycins accompanies inactivation of p60src in rat kidney cells infected with Rous sarcoma virus

1986 ◽  
Vol 6 (6) ◽  
pp. 2198-2206
Author(s):  
Y Uehara ◽  
M Hori ◽  
T Takeuchi ◽  
H Umezawa
Keyword(s):  
Immune Complex ◽  
Rous Sarcoma Virus ◽  
Morphological Changes ◽  
Rat Kidney ◽  
Phenotypic Change ◽  
Permissive Temperature ◽  
Rous Sarcoma ◽  
Cell Extracts ◽  
Sarcoma Virus

Three benzenoid ansamycin antibiotics (herbimycin, macbecin, and geldanamycin) were found to reduce the intracellular phosphorylation of p60src at a permissive temperature (33 degrees C) in a rat kidney cell line infected with a temperature-sensitive mutant of Rous sarcoma virus. This effect was accompanied by morphological changes from the transformed to the normal phenotype. The filamentous staining pattern of actin fibers was observed in the cells treated with these antibiotics at 33 degrees C. Removal of the antibiotics allowed the cells to revert to the transformed morphology. Ansamitocin, another benzenoid ansamycin, and naphthalenoid ansamycins such as streptovaricin and rifamycins did not show this effect. Pulse-labeling of the antibiotic-treated cultures with 32Pi showed a marked reduction of 32P radioactivity incorporated into p60src. A parallel experiment with [35S]methionine showed that synthesis of p60src was slightly inhibited. The immune complex prepared by mixing the herbimycin-treated cell extracts with antibody against p60src was inactive in vitro in phosphorylating the complex itself. On the contrary, the immune complex derived from untreated cells was active in vitro even in the presence of the antibiotics. These results suggest that benzoquinonoid ansamycins have no direct effect on src kinase but destroy its intracellular environment, resulting in an irreversible alteration of p60src and loss of catalytic activity.

Myristic acid is attached to the transforming protein of Rous sarcoma virus during or immediately after synthesis and is present in both soluble and membrane-bound forms of the protein

1984 ◽  
Vol 4 (12) ◽  
pp. 2697-2704
Author(s):  
J E Buss ◽  
M P Kamps ◽  
B M Sefton
Keyword(s):  
Plasma Membrane ◽  
Myristic Acid ◽  
Rous Sarcoma Virus ◽  
Minor Component ◽  
Rous Sarcoma ◽  
Cytoplasmic Face ◽  
Sarcoma Virus ◽  
Stable Part ◽  
Membrane Bound ◽  
A Minor

Myristic acid, a minor component of cellular fatty acids, has been shown previously to be covalently bound to most molecules of p60src, the transforming protein of Rous sarcoma virus. We have now determined at what time during the life cycle of p60src, and where within the cell, this lipid becomes attached to the protein. p60src was found to acquire myristic acid at only one time, during or immediately after its synthesis. p60src is known to be synthesized on free polysomes and appears at the cytoplasmic face of the plasma membrane after a lag of 10 min. The addition of myristic acid to p60src therefore precedes the binding of the protein to the plasma membrane. The lipid attached to p60src is a permanent, metabolically stable part of the protein; we found no evidence for turnover of the myristyl moiety. However, we did find myristate attached to various soluble forms of p60src and to a large number of cytosolic cellular proteins as well. This demonstrates that the attachment of myristic acid to a protein is not in itself sufficient to convert a soluble protein into a membrane-bound protein.

scholarly journals COMPARATIVE STUDIES IN ROUS SARCOMA WITH VIRUS, TUMOR CELLS, AND CHICK EMBRYO CELLS TRANSFORMED IN VITRO BY VIRUS

1962 ◽  
Vol 115 (1) ◽  
pp. 245-251 ◽  
Author(s):  
Robert M. Dougherty ◽  
Herbert R. Morgan
Keyword(s):  
Chick Embryo ◽  
Tumor Cells ◽  
Comparative Studies ◽  
Rous Sarcoma Virus ◽  
Rous Sarcoma ◽  
Transformed Fibroblasts ◽  
Embryo Cells ◽  
Sarcoma Virus

Chick embryo fibroblasts infected in vitro with Rous sarcoma virus have properties similar to tumor cells when injected into virus-immune chickens. When such virus-transformed fibroblasts are injected into normal chickens, they apparently participate in the production of tumors independent of their release of virus and are thus apparently malignant in vivo.

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