PROTEIN UPTAKE IN AMOEBAE 615 



1^. In 48 hr. many of the ferritin particles within the vesicles lose their 

 discrete character and form dense amorphous masses, as though their 

 protein coats were removed. 



These features are illustrated in Figs. 6, 7, and 8, which show successive 

 stages encountered in vesicles containing ferritin, and in Figs. 9, 10, and 11, 

 of comparable stages in vesicles containing methyl ferritin. 



Conclusion 



These studies on ferritin and methylated ferritin demonstrate that the 

 initial reaction of proteins with the cell surface depends upon net charge 

 effects, that the binding step may be temporarily uncoupled from the 

 subsequent stages of pinocytosis, and that neither free ferritin nor bound 

 methvl ferritin escapes in recognizable form from the pinocytosis vesicle. 

 Such observations, when considered with the experimental evidence 

 alreadv available from other studies, lead to the following view of pino- 

 cytosis in amoebae : 



Pinocytosis is a co-ordinated sequence of three main processes or 

 stages. The initial binding is an ion exchange reaction, which is capable of 

 some selectivity and of concentrating positively charged substances from 

 the environment. The binding reaction, under normal conditions, sets off 

 the active process of vesicle formation. In this stage, both the membrane 

 proper and the coat substance, with some free fluid as well, are carried into 

 the cell as vesicles are pinched off. This process is metabolicallv linked, as 

 the first is not, and appears to depend on cytoplasmic contractilitv in a 

 way which as yet has not been studied adequately. 



The third stage comprises a complex series of morphological and 

 chemical events within the cell. The lipoprotein membrane, the muco- 

 polysaccharide carrier substance, and the ingested substances are all 

 modified, each in a different way. The evidence, though by no means 

 complete, suggests that the changes include the digestion of protein and the 

 breaking up or partial digestion of the mucopolysaccharide carrier. This 

 implies the accumulation of hydrolytic enzymes within the vesicle, and 

 supports the idea, for which there is as well morphological evidence, that 

 the pinocytosis vesicle is fundamentally the same as the normal food 

 vacuole in which the amoeba digests his prey [16]. 



From what is known of the fate of smaller molecules, such as [^^C]- 

 glucose [17] and ribonuclease [18], it seems that such substances do pass 

 readily from the primary vesicle into the ground cytoplasm, or into the 

 microvesicles which are formed in great numbers from the primary vesicle. 

 Since neither free ferritin nor bound methyl ferritin escapes the vesicle, 

 such exchanges cannot be the result of a gross breakdown of membrane 



