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We have used mild detergent to analyze the core of Moloney Murine Leukemia Virus (MoMLV) and core-like complexes in infected cells. The immature core consists of the Gag polyprotein (PrGag) and viral RNA (vRNA). It is known to be detergent-resistant, in contrast to the mature Gag core. The core matures by cleavage of PrGag into MA (matrix), p12, CA (capsid) and NC (nucleocapsid) protein. We found that mature Gag proteins were bound to the PrGag cores. The degree of binding differed widely. No (<0.1%) p12 bound, low amount of CA (3-5%), and higher amount of MA (13-20%) bound. Varying NC was bound (5-15%). NC could be released by RNase A in agreement with its binding to viral RNA. The TM (transmembrane) protein was also examined. A low amount of TM was bound to the PrGag core (approximately 5%), whereas a very high amount (65%) of the PreTM (TM with the cytoplasmic R peptide tail) bound. The binding in the PrGag core appears to occur by direct protein-protein interactions as only minute amounts of lipids including raft lipids were observed after detergent treatment., (Copyright © 2013 Elsevier B.V. All rights reserved.)

We previously examined Moloney murine leukemia virus-induced fusion-from-within (FFWI) and fusion-from-without (FFWO) of SC-1 mouse cells. FFWI and FFWO can be distinguished by their stimulation by ionophores and polycations, respectively. FFWI is caused by infected cells. Normally, fusion between an infected cell and uninfected cells (heterofusions) is described, but we have surprisingly found that the infected cells also caused homofusion between uninfected cells in their vicinity (named neighbor homofusions). It was shown that neighbor homofusions were not induced by free virus particles (by FFWO). Transfectants expressing envelope proteins only induced heterofusions, indicating that virus production is necessary for the formation of neighbor homofusions. Both plasma membrane fragments and easily removable material from the surface of infected cells were able to induce fusion with the same stimulation pattern as FFWI and neighbor homofusion. These materials, especially the latter, have properties in common with virions, and it is discussed whether immature virions are involved in the formation of the neighbor homofusions.

Fusion-from-without (FFWO) and fusion-from-within (FFWI) for Moloney MLV in SC-1 cells were selectively stimulated by the polycation polybrene and the ionophore amphotericin B, which can be used to discriminate between the two fusion types. FFWI was stimulated by a number of different K+ ionophores. The stimulation occurred within few hours, did not require protein synthesis, and depended on the uninfected cell type, which suggests that FFWI is stimulated by a permutation of the K+ gradient of the uninfected cell. Plasma membrane vesicles from infected cells could also stimulate fusion with the same stimulation pattern as FFWI.

Virus-mediated cell-cell fusion with Moloney MLV and SC-1 cells was characterized. The level of fusion was highly dependent on the cell line used for propagation of the virus. Efficient fusion appeared to be very sensitive to negative charges on the cell surface and surroundings. Addition of polycations, removal of serum, and treatment with neuraminidase or hyaluronidase all stimulated fusion. Conversely, fusion was inhibited by fibronectin. Kinetic results and the time of action of inhibitors indicated that virus particles (or virus material) on the cell surface lead directly to fusion. The fusion then proceeded rapidly and required actin movement as shown by cytochalasin inhibition.