VIII. PROTEIX SYNTHESIS AND GENE ACTION 383 



actually making protein, was indicated by finding such RXA bound to 

 ribosomes. RXA having the characteristics of messenger RXA has been 

 reported in ribosomes of phage infected cells by Nomura et al. (1960), 

 Gros et al. (1961a); and Brenner et al. (1961). A similar ribosome- 

 associated RNA was demonstrated in normal (not phage-infectedj E. 

 coli by pulse-labeling (Gros et al., 1961a). Evidence for messenger RNA 

 in Pseudomonas aeruginosa, E. coli. and Bacillus megaterium was 

 reported by Hayashi and Spiegelman (1961). There authors used "step- 

 down" cultures, e.g., transferred from a rich to a limited medium, to 

 inhibit ribosomal RNA synthesis and thus permit a more clear-cut 

 labeling of the rapidly synthesized RNA fraction. This fraction was 

 characterized by hybridization to specific DNA (Hall and Spiegelman, 

 1961). ^inall3^ the elegant experiments of Brenner et al. (1961) have 

 shown that no new ribosomes were synthesized after phage infection, 

 and that the newly synthesized messenger RNA, as well as newly 

 synthesized protein, were associated with old ribosomes (present before 

 phage infection). 



The distribution of messenger RNA. in either normal or infected cells, 

 depends on the magnesium ion concentration. Binding to ribosomes 

 occurs in 0.01 M magnesium ion and most of the highly labeled RNA 

 can be dissociated in 10^ M magnesium ion. In addition to this type of 

 binding noted by several authors, Gros et al. (1961a) found a part of 

 the messenger RNA bound to 70S ribosomes in a more stable linkage. 

 Thus, two stages in messenger RNA fvmction may be, first, a dissociable 

 binding to ribosomes, followed by a more stable linkage. The results of 

 Tissieres et al. (1960) suggest that ribosomes containing the messenger 

 bound in stable linkage are active in protein synthesis. The results of 

 Brenner et al. (19611 are in agreement with this conclusion. 



Perhaps the earliest complete "expression" of a messenger RNA 

 concept was that of Jacob and Alonod (1961). The concept was based 

 partly on studies mentioned above, but also on studies of induced enzjaue 

 formation. While this latter subject is outside the scope of this review, 

 it is not possible to completely avoid this topic in any discussion of 

 messenger RNA. The experiments of Riley et al. (1960) illustrate the 

 type of evidence which led to the messenger RNA concept. The structural 

 gene (z-") for ^S-galactosidase was introduced into a recipient cell by 

 mating, and within a few minutes the maximal rate of enzyme synthesis 

 of the zygote was attained and continued at a uniform rate. If DNA 

 itself is not the template, then a rapid transfer of information to the 

 enzyme-forming site has taken place. This is similar to the entry of 

 phage DNA into the bacterial cell noted above. However, in addition, the 

 kinetics indicated that a constant number of active templates were pres- 



