262 



VINCENT G. ALLFREY 



We have shown that if thymus nuclei are isolated under isotonic con- 

 ditions in a medium containing 0-25 M sucrose and 0-003 ^ CaCl2, they 

 are able to retain their nucleic acids [5, 6], soluble enzymes [7], and low- 

 molecular weight compounds such as mononucleotides [8] and free 

 amino acids [9]. 



The isolated thymocyte nucleus also retains a surprising number of its 

 original synthetic capabilities. These include the synthesis of ATP by an 

 aerobic phosphorylating mechanism [8], the incorporation of thymidine 

 into deoxyribonucleic acid [10], and the utilization of a variety of purine 

 and pyrimidine precursors for ribonucleic acid synthesis [5, 6, 11, 12] and 

 partial turnover [11]. 



Nuclear activity in amino acid incorporation experiments is especially 

 striking [4, 5]. The latter process is illustrated by the uptake curves shown 



30 60 90 

 Time (min) 



120 



Fig. I. The time course of incorporation of [**C]-labelled amino acids into 

 the total proteins of isolated thymus nuclei during incubation /// vitro. 



in Fig. I ; these show the time course of labelling of the nuclear proteins 

 during incubations in the presence of either [i-^^C]-alanine, [i-^*C]- 

 glycine, or [2-^'*C]-lysine. 



There are two aspects of this incorporation process which mark it as 

 characteristically nuclear. First, amino acid uptake is sodium-ion depen- 

 dent. (The reasons for this are discussed below.) Secondly, amino acid 

 uptake does not take place if the nuclei are first treated with crystalline 

 pancreatic deoxyribonuclease. This evidence for the DNA-dependence of 

 nuclear amino acid incorporation is further supported by observations 

 that added DNA always restores uptake to DNAase-treated nuclei [5]. 

 (The reason for this is now clear, since recent experiments have shown 

 that nuclear ATP synthesis is DNA-dependent [13] ; removal of the DNA 

 stops nuclear phosphorylation, and without the necessary ATP, nuclear 

 protein synthesis cannot proceed [5, 8]. Restoring the DNA (or substitut- 



