Abrogation of Fv-1b restriction with murine leukemia viruses inactivated by heat or by gamma irradiation

Abstract

Fv-lb restriction in BALB/3T3 cells is temporarily abrogated following infection with N-tropic murine leukemia virus. The mechanism of this phenomenon was investigated by comparing the inactivation rates for viral infectivity and for the ability of the same virus to abrogate Fv-1 restriction. Inactivation of the abrogating ability of N-tropic murine leukemia virus following graduated doses of gamma radiation proceeded at half the rate of that for viral infectivity. This result indicates that viral RNA must function in abrogating Fv-lb restriction but that only a portion of the viral genome is required. The inactivation kinetics of Ntropic murine leukemia virus were also determined following incubation of virus at 43°C. Abrogating ability of N-tropic murine leukemia virus was found to be about six times as stable under these conditions as was viral infectivity. Interestingly, virion-associated reverse transcriptase activity was inactivated at the same rate as was viral infectivity, indicating that this enzyme may not need to function during abrogation. Virus heated at 43°C was used to study the kinetics of the abrogation phenomenon itself. Abrogation was shown to be transient, requiring 6 to 9 h after virus infection to become maximally effective and beginning to disappear after about 18 h. The data reported here confirm the idea that abrogation of Fv-1 restriction can be separated experimentally from virus replication, and they raise the possibility that a separate biochemical pathway exists for incoming viral RNA in Fv-1 restrictive cells. Certain murine leukemia viruses (MuLV's) have reported that Fv-i restriction is not assoare inhibited in their ability to replicate in ciated with a two-hit requirement for productive certain mouse cells by a naturally occurring re-infection of nonpermissive celLs (18, 31). striction system dependent upon the presence of To understand and to characterize Fv-1 rethe Fv-i mouse gene (16, 24). Studies of Fv-i striction more fully, we have initiated a series of restriction thus far have shown that MuLV's are experiments designed to examine the nature of probably inactivated after they adsorb to and the two-hit requirement for MuLV replication penetrate nonpermissive cells (17,21), but before in nonpermissive cells. This work has been DNA transcribed from incoming viral RNA is greatly facilitated by the use of defective murine integrated into the host cell genome (19, 32). sarcoma virus (MSV) pseudotypes which have The exact mechanism of Fv-i restriction, howbeen rendered phenotypically sensitive to Fv-1 ever, has yet to be elucidated. restriction following mixed infection with Nor Fv-1 restriction characteristically alters the B-tropic MuLV during the "rescue" process (1, titration patterns of N-and B-tropic MuLV's. 7). Although these virus stocks infect corresponding In a previous report we used an N-tropic pseupermissive cells with one-hit kinetics, they give dotype of Moloney MSV, rescued from transtwo-hit or multi-hit dose-response curves in Fv-formed S+L-cells by N-tropic MuL...

Mechanism of restriction of murine leukemia viruses varies between different strains of Fv‐1n mice

“…The basic infectious center technique and the procedures for sequential infections in time-course experiments have been described before (Duran-Troise et ui., 1977). However, recent work has shown that N-tropic MuLV which has been inactivated at 43" C is quite efficient at abrogating Fv-lb restriction in BALB/3T3 cells (Bassin et al, 1978). We have therefore measured Fv-1 restriction in these experiments by a different technique from \ *\ 132 YE--, Mvcv *\ 0 that employed earlier (Duran-Troise et al, 1977): in place of fully infectious MuLV, we have used heat-inactivated MuLV as the " abrogating " virus, while fully infectious MuLV has replaced the corresponding MSV pseudotype as an " indicator " of cellular permissiveness.…”

“…A study of the abrogation of Fv-l restriction in various cell strains enables us to probe the mechanism of the cellular restriction process itself, since abrogation results from specific Fv-1 -mediated intracellular events following viral penetration of the non-permissive cells (Hudng et a/., 1973; Krontiris et a/., 1973). Since N-tropic virus heated at 43" C retains its ability to abrogate Fv-lb re-striction (Bassin et al, 1978), it serves as a convenient reagent with which to study the mechanism of Fv-1 restriction in various cells. We thsreforz tested the ability of heated virus preparations to abrogate Fv-1" restriction in DBA/2 czlls.…”

“…Anothcr characteristic of abrogation in BALB/3T3 cells is its time-dependence (Niwa et al, 1976;Duran-Troise et al. 1977;Bassin et a/., 1978Bassin et a/., , 1979. It takes about 6 h for MuLV to abrogate Fv-lb restriction after infection of BALB/3T3 cells, and the effect lasts for about 18 h after which the cells again exhibit Fv-1 restriction.…”

“…Although the mechanism of abrogation in BALB/ 3T3 cells is unknown at present, our studies show that abrogation of Fv-lb restriction in a cell requires infection with a single virus particle. Neither viral reverse transcriptase nor viral DNA synthesis is required for abrogation, and it has been postulated that the RNA of a restricted virus may function as a message for the synthesis of a viral protein which is capable of inactivating the Fv-1 gene product (Bassin et al, 1978). The phenomenon of abrogation appears to be a unique feature of Fv-1 Received: July 18, 1979. restriction, representing a mechanism whereby two identical virus particles can complement one another functionally (Bassin et al, 1978).…”

“…Neither viral reverse transcriptase nor viral DNA synthesis is required for abrogation, and it has been postulated that the RNA of a restricted virus may function as a message for the synthesis of a viral protein which is capable of inactivating the Fv-1 gene product (Bassin et al, 1978). The phenomenon of abrogation appears to be a unique feature of Fv-1 Received: July 18, 1979. restriction, representing a mechanism whereby two identical virus particles can complement one another functionally (Bassin et al, 1978). Even though the mechanism of Fv-1 restriction and its abrogation in BALB/3T3 cells is not fully understood, comparative studies designed to determine whether or not the mechanism of restriction in Fv-1" cells is the same as it is in Fv-lb cells, can be carried out at this time and are reported below.…”