Transcriptase activity and genome composition of defective influenza virus

Seven complementation-recombination groups of temperature-sensitive (ts) influenza WSN virus mutants have been previously isolated. Recently two of these groups (IV and VI) were shown to possess defects in the neuraminidase and the hemagglutinin gene, respectively, and two groups (I and III) were reported to have defects in the P3 and P1 proteins which are required for complementary RNA synthesis. In this communication we report on the defects in the remaining three mutant groups. Wild-type (ts+) recombinants derived from ts mutants and different non-ts influenza viruses were analyzed on RNA polyacrylamide gels. This technique permitted the identification of the P2 protein, the nucleoprotein, and the M protein as the defective gene products in mutant groups II, V, and VII, respectively. Based on the physiological behavior of mutants in groups II and V, it appears that P2 protein and nucleoprotein are required for virion RNA synthesis during influenza virus replication.

Section: Resultsmentioning

confidence: 99%

Identification of the defective genes in three mutant groups of influenza virus

Ritchey¹,

Palese²

1977

“…The loss of large RNA species from von Magnus particles has been observed consistently (1,3,7,18,22), but exactly what gene segment(s) was lost has not been determined unambiguously. The finding in our previous study (17) and this study and the study made by Janda et al (10) showed that the loss of polymerase genes varied from one von Magnus preparation to another, just as the species of extra RNAs contained in them were variable.…”

Section: Discussionmentioning

confidence: 99%

Extra RNAs of von Magnus particles of influenza virus cause reduction of particular polymerase genes

Ueda¹,

Nakajima²,

Sugiura³

1980

Extra RNAs, or RNA species other than eight gene segments, in von Magnus particles of the influenza virus WSN strain were studied by polyacrylamide gel electrophoresis and oligonucleotide mapping. From the original virus stock, various cloned stocks were obtained, each giving rise to a characteristic set of extra RNAs. One cloned virus stock contained a large number of von Magnus particles. The RNA pattern was characterized by two prominent extra RNAs (X1 and X2) and a decrease in the content of two polymerase genes, P1 and P2. Segregation of the two extra RNAs was carried out by coinfection of cells with a von Magnus particle and infectious virions. The results showed that the presence of one of the extra RNAs (X2) was associated with a reduction in the amount of the P1 gene and that the presence of the other extra RNA (X1) was associated with a reduction in the amount of the P2 gene. Oligonucleotide mapping showed that both extra RNAs, X1 and X2, were derived from the P1 gene. The results suggested that an extra RNA did not necessarily cause the reduction of the progenitor polymerase gene, but might cause the reduction of another polymerase gene.

“…NAYAK ET AL. reports indicated that DI viruses lack entirely or contain only reduced amounts of the largest segment of viral RNA (4, 6, 23). However, this view that DI influenza viruses lack a specific RNA segment has been disputed by a recent observation which suggests that influenza DI viruses do not selectively lose a specific RNA segment (2). There are a number of major difficulties in studying DI influenza viruses.…”

mentioning

confidence: 99%

Homologous interference mediated by defective interfering influenza virus derived from a temperature-sensitive mutant of influenza virus

Nayak¹,

Tobita²,

Janda³

et al. 1978

A temperature-sensitive group II mutant of influenza virus, ts-52, with a presumed defect in viral RNA synthesis, readily produced von Magnus-type defective interfering virus (DI virus) when passed serially (four times) at high multiplicity in MDBK cells. The defective virus (ts-52 DI virus) had a high hemagglutinin and a low infectivity titer, and strongly interfered with the replication of standard infectious viruses (both ts-52 and wild-type ts+) in co-infected cells. Progeny virus particles produced by co-infection of DI virus and infectious virus were also defective and also had low infectivity, high hemagglutinating activity, and a strong interfering property. Infectious viruses ts+ and ts-52 were indistinguishable from ts-52 DI viruses by sucrose velocity or density gradient analysis. Additionally, these viruses all possessed similar morphology. However, when the RNA of DI viruses was analyzed by use of polyacrylamide gels containing 6 M urea, there was a reduction in the amount of large RNA species (Vl to V4), and a number of new smaller RNA species (Dl to D6) with molecular weights ranging from 2.9 x 105 to 1.05 x 105 appeared. Since these smaller RNA species (Dl to D6) were absent in some clones of infectious viruses, but were consistently associated with DI viruses and increased during undiluted passages and during co-infection of ts-52 with DI virus, they appeared to be a characteristic of DI viruses. Additionally, the UV target size of interfering activity and infectivity of DI virus indicated that interfering activity was 40 times more resistant to UV irradiation than was infectivity, further implicating small RNA molecules in interference. Our data suggest that the loss of infectivity observed among DI viruses may be due to nonspecific loss of a viral RNA segment(s), and the interfering property of DI viruses may be due to interfering RNA segments (DI RNA, Dl to D6). ts-52 DI virus interfered with the replication of standard virus (ts+) at both permissive (34°C) and nonpermissive temperatures. The infectivity of the progeny virus was reduced to 0.2% for ts+ and 0.05% for ts-52 virus without a reduction in hemagglutinin titer. Interference was dependent on the concentration of DI virus. A particle ratio of 1 between DI virus (0.001 PFU/cell) and infectious virus (1.0 PFU/cell) produced a maximal amount of interference. Infectious virius yield was reduced 99.9% without any reduction of the yield of DI viruses. Interference was also dependent on the time of addition of DI virus. Interference was most effective within the first 3 h of infection by infectious virus, indicating interference with an early function during viral replication. Although defective interfering virus particles properties of DI influenza viruses. The intracel-(DI virus or von Magnus virus) possessing nor-lular events that lead to the formation of these mal or nearly normal hemagglutinating activity DI particles remain unknown. Furthermore, the but decreased infectivity were first shown in mechanism by which DI influenza viruses ca...