POLYN'UCLEOTIDE SYNTHESIS IN Nl'CLEOLUS AND CI I KOMOSdM i;s I yQ 



transferred to non-radioactive food, only those which ate vigorously and passed 

 all of the red colored food through the intestinal tract within i or i hours were 

 used tor fixation at the later times. At each time interval about lo larvae were 

 quickly frozen in isopentane, which had been cooled to near its freezing point 

 with liciuid nitrogen, and (.lehydrated in absolute ethanol at — 45°("(22). They 

 were then fixed by heating tor i hour at 60° in 75 per cent ethanol in water. 

 These larvae were then dehydrated, embedded, and in 5-micron sections prepared 

 for autoradiography as previously described (17). 



The cells of the salivary gland are growing rapidly during most of the third 

 instar. In table i are shown data on the increase in volume for nucleolus, nuclei 

 and cytoplasm for the cells of the group of larvae used for the experiment de- 

 scribed here. Since nucleoli and nuclei are nearly spherical, their volumes were 

 calculated from direct measurements of the diameters. However, the cells have 

 very irregular shapes and their volumes had to be estimated by camera lucida 

 drawings of serial sections. The successive tracings of each cell were cut out and 

 weighed to get relative volumes. These were converted to absolute measurements 

 by comparing with nuclear volumes which were determined directly from the 

 measured diameters and by means of the tracings. 



During the 24 hours of the experiment the nucleolus almost doubled its vol- 

 ume and RNA content, since the concentration remained constant. Nuclear vol- 

 ume increased by a factor of 3 during the same period. The cytoplasm increased 

 by a factor of 3 in 14 hours. The nuclear-cytoplasmic ratio did not change appre- 

 ciably during the first 14 hours of the experiment, but the cytoplasm probably 

 grew faster in later stages. During the period of the experiment the whole cell 

 probably was growing as fast as embryonic cells during exponential growth. The 

 cell volume and its RNA content doubled in less than 10 hours. 



Data and Interpretation. From the data in table 2 and figure 5, the fast rate of 

 incorporation regularly seen in the nucleolar RNA is immediately apparent. The 

 relatively more rapid rise in its specific activity compared to the experiments with 

 P'*" (4) indicates a smaller pool size for the adenine and derivatives as compared 

 with phosphates. The smaller pool is a fortunate circumstance, for it allows 

 quicker response to changes in food that reveal different metabolic types of RNA. 

 Since water- and alcohol-soluble substances are always extracted from the tissues 

 during preparation for autoradiography, direct determination of soluble pre- 

 cursors is not feasible. 



When the larvae were removed from the radioactive food at the eiui of 2 hours 

 the labeling of nucleolar RNA already showed evidence of an approach to an 

 ecjuilibrium while the label in both the cytoplasmic and chromosomal (chromatin 

 or nuclear) RNA's, after an initial lag, was still increasing at approximately a 

 linear rate. The change in specific activity of nucleolar RNA is rapid; it tiropped 

 in 5 hours to less than one-half its peak value. During this whole period of 5 



