FIBRILLAR SYSTEMS IN THE MITOTIC APPARATUS 167 



proteins in the structural organization of the mitotic apparatus is 

 more strongly indicated by the in vivo studies of Mazia (1958) and 

 Mazia and Zimmerman (1958). They observed that proper concen- 

 trations of mercaptoethanol (HSCH.CH^OH), a simple, penetrat- 

 ing SH-compound of low toxicity, applied to sea-urchin and sand- 

 dollar eggs any time before metaphase, blocked the first cleavage 

 division. If it was applied during and subsequent to metaphase, no 

 interference with the first cleavage division was detected. The in 

 vivo experiments using mercaptoethanol argue strongly for the in- 

 volvement of protein-SH groups during the buildup of the mitotic 

 apparatus prior to cleavage, but do not give much insight into the 

 actual mechanism involved. The in vitro observations on the solu- 

 bilitv characteristics of mitotic apparatus isolated from ethanol- 

 preserved material by the digitonin method (Mazia, 1958) favor 

 the interpretation that some intermolecular disulfide bonds may be 

 responsible for maintaining the spatial orientation of the "precursor" 

 molecules in the mitotic apparatus. 



The speculation that the reversible conversion of sulfhydryl 

 groups to intermolecular disulfide linkages performs an essential role 

 in the construction and function of the division figure finds support 

 from other investigators. Some cytochemical studies for sulfhydryl 

 groups in sea-urchin eggs during the first cleavage division were 

 made by Kawamura and Dan ( 1958 ) . Using Bennett's reagent, they 

 observed that the cytoplasm and nucleus of the unfertilized egg did 

 not stain, but 3 minutes after fertilization, they detected an in- 

 creased general stainability. At metaphase, the entire mitotic figure 

 of one species of sea urchin was intensely and uniformly stained, 

 contrasting sharply with the less strongly stained cytoplasm. In 

 early anaphase, the traction fibers were very intense, while the 

 interzonal region and centrospheres took up much less of the stain. 

 Little change in the astral rays was evident. Immediately after 

 cleavage, the staining character of the cytoplasm of the blastomeres 

 was essentially the same as that observed for the unfertilized egg; 

 that is, it was unstained. Shimamura et al. ( 1957 ) have stained the 

 mitotic apparatus for both sulfhydryl and disulfide groups, in paral- 

 lel, simultaneous experiments. Their photographs clearly reveal 

 that the regions stained for sulfhydryl groups usually complement 

 the areas stained for disulfide groups at given stages. This was most 

 evident in the anaphase and early telophase figures, in which the 

 asters stained only lightly for sulfhydryl groups but very intensely 



