JEAN BRACHET 



amoebae lend some support to the idea that the nucleus might play an important 

 part in the production of coenzymes and perhaps regulate in this way the activity of 

 the cytoplasmic enzymes. 



We have seen earlier that non-nucleated fragments of Amoeba proteus retain a 

 normal oxygen consumption for as long as ten days; however, a very striking and 

 rapid change, first described by Mazia and Hirshfield (1950), and confirmed after- 

 wards in this laboratory (Brachet, 1952), occurs when the penetration into the cell 

 of radioactive phosphate is followed : the 32 P uptake is strongly reduced in the non- 

 nucleated halves, only a few hours after the removal of the nucleus. Since oxidations 

 and phosphate uptake are usually closely linked, we were led to believe (Brachet, 

 1 95 1, 1952a) that enucleation interrupts the normal coupling between oxidations 

 and phosphorylations, just as poisons like dinitrophenol, usnic acid, gramicidine, etc. 

 do ; if so, production by the nucleus of coenzymes necessary to couple oxidations 

 with phosphorylations seemed a plausible explanation for the experimental results. 



Later experiments (Brachet, 1952^) have shown, however, that such a simple 

 hypothesis can hardly be correct, since the adenosinetriphosphate (ATP) content 

 of nucleated and non-nucleated halves remains almost identical for a considerable 

 length of time (twelve days) : a drop in the ATP content of the non-nucleated halves 

 was of course to be expected if removal of the nucleus really uncoupled phosphory- 

 lations from oxidations. 



But very recently (Brachet, 1954) we have studied the ATP content of nucleated 

 and non-nucleated halves when they are placed in anoxic conditions, which strongly 

 affect the motility of the amoebae (Pan tin, 1930; Kitching, 1939); it was found 

 that, in the case of fragments from amoebae which had been sectioned for a few 

 hours only, there is a 70 per cent, drop in the ATP level of the non-nucleated halves 

 after 16 hours' anaerobiosis ; this drop is only 15-20 per cent, in the case of the nu- 

 cleated halves. When the fragments are 4-9 days old, the differences become less: 

 we obtain a 50 per cent, drop in the case of the nucleated halves, as compared with 

 70 per cent, again in the non-nucleated ones. 



These findings are probably correlated with the fact, already mentioned in this 

 paper, that utilization of glycogen is much poorer in the non-nucleated than in the 

 nucleated halves: during the initial 1-3 day period, the utilization of glycogen by 

 the nucleated halves is faster than it is later on ; in the non-nucleated fragments there 

 is a restricted utilization of glycogen at the beginning (1-3 days), and it completely 

 ceases thereafter. Preliminary cytochemical observations on the glycogen content 

 (Bauer's method) of both types of fragment in anaerobiosis have shown that nuclea- 

 ted halves from well-fed amoebae (0-1 day after bisection) lose a very large pro- 

 portion of their glycogen during the 16 hours' period of anaerobiosis; such striking 

 changes do not occur in the non-nucleated halves of freshly bisected amoebae. 

 Apparently, removal of the nucleus quickly and simultaneously impairs both 

 glycogen breakdown and ability to keep ATP phosphorylated, under anoxic con- 

 ditions. 



These results strongly suggest that the nucleus controls certain steps of glycolysis, a 

 conclusion which agrees with the observation of Stern and Mirsky (1952) that 

 wheat germ nuclei are richer in several glycolytic enzymes than the cytoplasm; 

 however, it was found (Brachet, 1954) that enolase, which according to Stern and 



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