VIII. PROTEIN SYNTHESIS AND GENE ACTION 369 



found in all three chromatographic fractions. Bloemendal et al. (1961) 

 observed a similar transfer of labeled RNA to detergent-treated liver 

 ribosomes. When the soluble RNA was prelabeled in the terminal posi- 

 tion with C"-adenine and incubated with ribosomes, the C^*-adenine was 

 found in internal positions in the ribosomal RNA (63% of the radio- 

 activity was isolated as adenosine monophosphate after alkaline hydrol- 

 ysis). It appears, then, that the transfer which occurred in this system 

 resulted in the covalent linkage of solul)le RNA with microsomal RNA. 

 It is not clear, however, that this reaction is related in any way to the 

 transfer of amino acids from amino acyl-RNA. In fact, the addition of 

 soluble enzymes and an amino acid mixture failed to stimulate the RNA 

 transfer (Bloemendal et al., 1961). 



A transfer of labeled S-RNA to ribosomes has been reported by 

 Hoagland and Comly (1960). In this case, P''=-labeled, soluble RNA was 

 coupled with C'^-leucinc so that the transfer of both the RNA and the 

 amino acid could be studied. It was shown not only that the P^--labeled 

 RNA would enter the sedimentable fraction, but also that it could be 

 displaced by unlabeled soluble RNA. A measure of specificity was 

 indicated by the inability of unlabeled microsomal RNA to displace the 

 labeled soluble RNA from the microsomes. In agreement with the experi- 

 ments of Bloemendal et al. (1961), when the C^'^-leucine was removed 

 from the P^--RNA, no inhibition of RNA transfer was observed. 



These various experiments do demonstrate that an association between 

 soluble RNA and the ribonucleoprotein particles may be very readily 

 achieved. The slight stimulations observed upon the addition of ATP 

 or GTP may well indicate simply that this is not an energy-dependent 

 step. 



Since it is altogether likely that a physical association can occur 

 between soluble RNA and the microsomes, it might be expected that 

 when C"-amino acid-labeled amino acyl-RNA is incubated briefly with 

 microsomes, some portion of the amino acyl-RNA will sediment with 

 the microsomal fraction. Zamecnik (1960) has in fact observed such a 

 transfer to the RNA of the sedimentable fraction in a rat liver system. 

 The problem of the first intermediate in the ribosome has been studied 

 in the reticulocyte system starting with C^^-leucyl-RNA (G. Favelukes 

 and R. Schweet, unpublished data) to avoid complications of contam- 

 ination with free C'^-amino acid. Incorporation into ribosomes in alkali- 

 labile linkage was used as a measure of amino acyl-RNA binding. As 

 shown in Table II, a GTP- and enzyme-dependent linkage was found. 

 However, the significance of these alkali-labile counts as an inter- 

 mediate in jirotein synthesis is dubious, since these counts could not 

 be "chased" into alkali-stable (protein) linkage. It seems likely, there- 



