198 1. MALONATE 



Egg cleavage is generally rather sensitive to malonate. Although Arbacia 

 eggs divide normally in 1 mM malonate (Krahl and Clowes, 1940), Dend- 

 raster eggs are inhibited quite well at this concentration (Pease, 1941). The 

 development of bilaterality in Dendraster, seen with many inhibitors, does 

 not occur with malonate even at 10 mM, demonstrating a true differential 

 effect on cleavage. Division of Arbacia and Chaetopterus eggs is inhibited 

 completely by malonate at 70 mM, although 40 mM is essentially without 

 action, and this is completely reversed by fumarate, indicating a block of 

 the cycle (Brust and Barnett, 1952; Barnett, 1953). This is not a sodium 

 effect since NaCl does not inhibit. Such high concentrations of malonate 

 are not unreasonable in work in sea water and the inhibitions here may be 

 quite specific. Egg cleavage seems to depend on the ATP generated in the 

 cycle, because concentrations of malonate that completely inhibit the cleav- 

 age of Chaetopterus and Lytechinus eggs cause an immediate drop in the 

 high-energy phosphate to the unfertilized levels, and inorganic phosphate 

 is actually lost from the cells (Barnett and Downey, 1955). 



The effects of malonate on the development of Arbacia eggs were 

 thoroughly investigated by Rulon (1948), who demonstrated abnormal dif- 

 ferentiation with low concentrations of malonate. If fertilized eggs in the 

 1-2-cell stage are placed in 1.2 mM malonate, a slight retardation of cleav- 

 age is observed, and at 13 hr (when the controls are swimming bilateral 

 gastrulae) there is no evidence of gastrulation, development having pro- 

 gressed only to spherical blastulae with no ventral flattening. After 24 hr 

 (when the controls are plutei), abnormal gastrulae with thickened apical 

 ends and long active cilia are seen. At 48 hr some had developed into ab- 

 normal plutei with ciliated apical knobs rather than normal arms. When 

 malonate is removed after 13 hr, quite normal plutei are formed, showing 

 that the effect is readily reversible. It is interesting that eggs exposed to 

 malonate for varying times, then washed free of malonate and fertilized, 

 show abnormal development, demonstrating that malonate can so disturb 

 egg metabolism that the effects are made evident later after the malonate 

 is no longer present. Exposure of unfertilized eggs to 1.44 mM malonate 

 for 12 hr, for example, leads to only 30 40% cleavage with irregular cleav- 

 age furrows and cells of unequal sizes, development not progressing 

 beyond irregular blastulae. Rulon postulated a gradient of succinate de- 

 hydrogenase throughout the cells and embryos, paralleling the physiological 

 activity gradient, but it is not necessary to assume this to explain effects on 

 differentiation. In connection with our work on parapyruvate (Montgomery 

 and Bamberger, 1955), we examined the development of Strongylocentrotus 

 purpuratus in 25 mM malonate. Up to 24 hr no discernible differences from 

 the controls are seen, but at 44 hr a slight inhibition of development can be 

 detected, with less formation of the primary mesoderm. Incoordination of 

 ciliary activity is also evident, most of the blastulae simply rotating rather 



