366 



( II \l- I IR 2<S 



\irus is know ii (Figure 28—2); much less is 

 known about the sequence of ribotides in 

 TMV. One can v lsualize TMV as a polymer 

 of 3' mononucleotides ending in a nucleoside 

 — shown to be adenosine.' 



Snake venom phosphodiesterase (see p. 

 2S2 ) splits off 5'-nucleotides one at a time 

 starting at the free 3'-OH end — called the 

 .V or nucleoside end — of TMV, revealing 

 that the terminal base sequence is — UACUA 

 (or possibly — UAUCA). The sedimentary 

 behavior of TMV RNA from which one to 

 three nucleotides have been enzymatically re- 

 moved does not change, implying that the 

 enzyme primarily acts at the end of the RNA 

 molecule as an exonuclease. The intrinsic 

 integrity of the molecule is further demon- 

 strated by the infectivity of such terminally- 

 deleted TMV and the production of progeny 

 TMV. When the progeny TMV are in turn 

 treated with the same diesterase, they are 

 found to release first A, then U, and then C. 

 Thus, when RNA, from which several ter- 

 minal nucleotides have presumably been re- 

 moved, replicates, the original nucleotide se- 

 quence seems to be restored in the progeny. 7 

 It is possible that the sequence occurs in the 

 progeny by chance; it is also possible that the 

 free 5'-OH end — called the 5' or nucleotide 

 end — can normally base-pair for some dis- 

 tance with the 3' end of TMV. If diesterase 

 does remove the terminal — CUA, the short- 

 ened 3' end could be repaired by making the 

 complement of a UAG — sequence (which 



8 From work of T. Sugiyama and H. Fraenkel- 



Conrat (1961). 



7 See B. Singer and H. Fraenkel-Conrat (1963). 



we would presume starts the 5' end ). On the 

 other hand, it seems more likely that the in- 

 fectivity of these degraded RNAs, which is 

 about 109? o\' normal, is due to residual 

 undegraded RNAs, which, of course, infect 

 perfectly normally. The degradation with 

 snake venom diesterase is very likely not 

 synchronous, so that some molecules can 

 have several nucleotides removed before 

 others lose any. 



Results obtained with TMV and other 

 RNA-viruses show that complementary 

 RNA strand formation occurs during repli- 

 cation of RNA genetic material. RNA- 

 dependent RNA polymerase, RNA synthe- 

 tase, or RNA replicase, an enzyme which 

 utilizes riboside triphosphates and takes di- 

 rections from RNA to make complementary 

 RNA, has been isolated and purified. s One 

 RNA synthetase uses the single-stranded 

 RNA of the <f>MS2 — called the "plus" strand 

 — as a template to synthesize the comple- 

 mentary "minus 1 ' RNA strand in vivo. The 

 double-stranded product — called the replica- 

 tive form — is used in vitro as a natural tem- 

 plate by the same or another RNA synthetase 

 to synthesize "plus" strands." In this con- 

 nection it should be noted that the infective 

 forms of a wound virus obtained from sweet 

 clover and a reovirus associated with the 

 respiratory and enteric tracts of animals, in- 

 cluding man, have double-stranded RNA as 

 their genetic material. 10 



s By I. Haruna, K. Nozu, Y. Ohtaka, and S. Spie- 

 gelman (1963), and by C. Weissmann, L. Simon, 

 and S. Ochoa (1963). See D. Baltimore (1964). 

 °See C. Weissmann et al. ( 1964). 

 10 See P. J. Gomatos and I. Tamm (1963). 



SUMMARY AND CONCLUSIONS 



RNA is the sole carrier of genetic properties in certain viruses. Some animal RNA 

 viruses can undergo genetic recombination. Mature RNA viruses carry either single- 

 stranded or double-stranded RNA. Viral RNA replication involves RNA synthetase 

 and the formation of complementary RNA chains. 



