90 CONTROL MECHANISMS IN CELLULAR PROCESSES 



Autoradiography showed that this incorporation occurred mainly 

 into the nuclear RNA. Amoebae could be cut into nucleate and 

 anucleate halves and onlv the former incorporated P''- into RNA. 

 When a nucleus containing radioactive RNA was transplanted into 

 a non-radioactive nucleated or enucleated amoeba, the radioactivity 

 was transferred into the cytoplasm, where it could still be recognized 

 as RNA. The conclusion was clear: RNA is made in the nucleus and 

 transferred to the cvtoplasm. In later work, however, Plant and 

 Rustad (1957, 1959) observed RNA formation in enucleated halves 

 also, concluding that the nucleus is not the only site of RNA forma- 

 tion. Prescott (1959) tried to explain this later finding by a per- 

 sistence of some live bacteria in the food vacuoles of experimental 

 amoebae. By using an axenic culture of Acanthamoeba, Prescott 

 could ( 1960a ) demonstrate that there is no uptake of C^^ cytidine 

 by the enucleated halves, even over a 24-hour period. 



Plant's contention of RNA formation in the absence of nuclei 

 seemed to be supported by experiments in Acetabularia. Brachet 

 et al. ( 1955 ) reported an increase in RNA content in anucleate frag- 

 ments of this alga, as measured by the tracer dilution method. 

 Richter (1957), measuring RNA production with the UV light-ab- 

 sorption method, could find no such increase. Finally, Naora et al. 

 ( 1959 ) , in Brachet's laboratory, by using both methods, could show 

 tliat there was no increase in the cvtoplasmic RNA after enucleation, 

 but rather a small decrease. However, anucleate plants were still 

 able to incorporate C^^ precursors into RNA. The meaning of this 

 incorporation is not clear; it could be due to a real turnover of cyto- 

 plasmic RNA or only to exchange reactions. This points to the 

 pitfalls of tracer work. It is not possible to distinguish between net 

 svnthesis and exchange reactions in the incorporation of radioactive 

 precursors. Furthermore, in autoradiography, it is not easy to follow 

 the incorporation into a specific substance, since other incorporating 

 substances may not be readily eliminated. 



In Neurospora, it has been possible to demonstrate that, in the 

 actively growing mycelium, the major part, if not all, of the RNA is 

 synthesized in the nuclear fraction (Zalokar, 1959a, 1960b). Hyphae 

 were fed H'^ uridine for different intervals of time and then centri- 

 fuged to separate cell constituents inside still living hyphae. Cen- 

 trifuged hyphae were fixed, extracted with cold trichloracetic acid 

 and autoradiographs made. Centrifugation sedimented all ribo- 

 somes (Zalokar, 1960c, 1961) into a layer, separated from a layer 



