differentiation are shown in Fig. 2. Firstly, tlie 

 epithelial cells have basophilic staining prop- 

 erties whereas the fiber cell has acidophilic 

 staining properties. This change in staining 

 properties may have been brought about by the 

 synthesis of r-crystallins. These proteins are 

 slightly basic having isoelectric points ranging 

 from pH 7.5 - 9.0. In view of their basic prop- 

 erties the r-crystallins may bind to the nucleic 

 acids of the nucleus and cytoplasm thus effect- 

 ing the observed alteration in staining 

 properties. It should also be pointed out that 

 the isoelectric points of the a-crystallins is 

 5.2 and of the ^-crystallins ranges from 6.0 to 

 7.0. Both these groups of proteins as well as 

 the rich population of ribosomes would con- 

 tribute to the basophilic staining properties of 

 this cell. 



In addition to the change in staining prop- 

 erties there is also a conversion from a rough 

 endoplasmic reticulum in the epithelial cell to 

 a smooth endoplasmic reticulum in the fiber 

 cell. The breakdown and subsequent decrease 

 in the ribosomes may be directly related to 

 the appearance of the endoplasmic reticulum. 

 At present I cannot present any information 

 on the mechanism of this ribosomal breakdown. 

 The fact, however, that there is also an overall 

 decrease in the rate of protein synthesis in the 

 fiber cells may be a consequence of the ribosomal 

 breakdown. 



The increase in the size of the nucleus and 

 nucleoli and the increase in the ribosomal 

 population at the time of cellular elongation 

 might indicate the initiation of an overall syn- 

 thesis of materials required for the morpho- 

 logical changes of the cell. Although we know 

 nothing of the function of r-crystallins, it has 

 been shown that the synthesis of this major 

 group of proteins is initiated during cellular 

 elongation. In addition to the increase in protein 

 synthesis there must also be an increase in the 

 synthesis of nucleic acids, both ribosomal and 

 m-RNA. The synthesis of these two classes of 

 RNA may account, therefore, for the morpho- 

 logical changes in the nuclei and nucleoli. 



In the fiber cell it has been observed that 

 there is a gradual decrease in the size of the 

 nucleus and as the cell gets older the nucleus 

 disappears. Through the use of ■^H-thymidine 

 we have shown, as would be expected in a 

 replicative cell, that the epithelial cells contain 

 metabolically active DNA whereas the fiber 

 cells no longer have the capacity to incorporate 

 precursors into its DNA. Furthermore, the data 

 from our MAK columns have shown that there 



is a significant loss of DNA in the fiber cell. 

 Both of these observations are in complete 

 agreement with the cytological observations 

 on the fate of the nucleus in lens fiber cell 

 differentiation. 



The loss of nuclear activity brings up the 

 question of the synthesis of m-RNA for the 

 continuation of protein synthesis. Upon inacti- 

 vation of the nucleus, the synthesis of m-RNA 

 stops and the cell would require some mecha- 

 nism for the conservation of existing m-RNA 

 for the continuation of protein synthesis. The 

 stabilization of m-RNA in these fiber cells 

 has been shown; the mechanism of stabiliza- 

 tion, which has not been worked out, should 

 prove to be an important one for understanding 

 the molecular aspects of terminal cellular 

 differentiation. 



I have presented just a limited spectrum 

 of the macromolecular interactions which occur 

 during the terminal stages of lens cell differ- 

 entiation. There are many cell types which 

 undergo similar morphological alterations dur- 

 ing their terminal stages of differentiation. 

 These cells are also involved in the synthesis 

 of highly specific proteins such as hemoglobin, 

 myosin and keratin and there are indications 

 that the same molecular alterations described 

 for the lens may also occur in these cells. 

 Thus, a specific series of macromolecular 

 interactions such as those described above 

 may be a basis for the biochemical definition 

 of the terminal stages of cellular differentia- 

 tion. The differentiation of the reticulocyte, for 

 example, involves inactivation of the nucleus 

 and stabilization of m-RNA. It remains to be 

 seen whether there also occurs a ribosomal 

 breakdown and the accumulation of a breakdown 

 product such as I have described here. Further- 

 more, the elucidation of the mechanisms of 

 reactions involving nuclear inactivation, the 

 stabilization of m-RNA and the breakdown of 

 the ribosomes may be the basis of the mecha- 

 nisms of terminal cellular differentiation. This 

 is important because most cells exhibiting the 

 property of synthesizing highly tissue specific 

 proteins, enter terminal stages of differentia- 

 tion and exhibit molecular properties similar 

 to those described above. 



The lens cell has reached its highest form 

 of cellular differentiation when it has formed 

 the fiber cell, and as it approaches this stage 

 it develops very specific metabolic activities. 

 With respect to the mechanism of lens fiber 

 cell formation, therefore, one would ask how 

 much of this metabolic activity is dependent 

 upon the morphological changes and whether 



63 



