CELLULAR DIFFERENTIATION IN THE SLIME MOLD 227 



Nevertheless, conditions can be arranged in such a way that aggre- 

 gation need not require the presence of an I-cell. Thus a given number 

 of m) xamoebae, dispensed on washed agar to which had been added a 

 cell-free extract obtained from cells already in the act of aggregation, 

 formed more than ten times the number of centers ordinarily seen in 

 the absence of the extract (Ennis and Sussman, 1958b). Yet a corre- 

 sponding increase in the number of I-cells was not detected. The same 

 result was obtained when wild-type myxamoebae were placed on one 

 side of a thin agar membrane and, on the other, a huge number of 

 cells of an aggregateless mutant strain, themselves unable to aggregate 

 spontaneously. Again a multiplicity of centers was formed by the wild 

 type without a corresponding increase in the number of I-cells. 



Most interesting was the finding that R-cells, although unable to 

 initiate centers among their developmental contemporaries (except 

 under the special conditions described above), could still induce their 

 developmental juniors to aggregate ( Sussman and Ennis, 1959 ) . Popu- 

 lations of R-cells, devoid of I-cells, were incubated on agar for 12 

 hours, a time by which aggregation would ordinarily have commenced. 

 Single R-cells were abstracted and micro-manipulated to test popula- 

 tions newly laid down upon the agar {i.e., 12 hours junior to the cells 

 to be tested). After an additional 12 hours' incubation, about one of 

 four of these test populations was found to have been induced to ag- 

 gregate by the addition of older R-cells. Yet the samples from which 

 the older R-cells were taken still displayed no signs of aggregation! 



The fact that experimental conditions can be arranged so that 

 myxamoebae other than I-cells initiate centers of aggregation raises the 

 question of how important the latter may be under "natural conditions," 

 i.e., in the normal ecological niches occupied by D. discoideum. Unfor- 

 tunately, very little of ecological significance is known about the cellu- 

 lar slime molds, save that they live in the soil, are associated with bac- 

 teria, and can be found in the forests of Virginia, the heaths of Eng- 

 land, the golf courses of Wilmette, Illinois, and on the dung of white 

 rabbits from Atlanta, Georgia ( Raper and Thorn, 1932; Sussman, 1956; 

 Cohen, 1953). Only one micropedological photograph, taken by acci- 

 dent, attests to the fact that slime molds actually aggregate and fruit in 

 nature (albeit, one could not conceive that they would not). Conse- 

 quently it is impossible at present to assess this point. 



Nevertheless, these results have suggested that the difference be- 

 tween the two cell types with respect to initiative capacity is not a qual- 

 itative but only a quantitative one. Perhaps the superior capacity of the 

 I-cells is simply a reflection of their considerably greater size and 

 possibly higher rate of general metabolic activity ( as judged by motil- 

 ity, pseudopodial protrusions, vacuolar contraction ) . Experiments have 

 demonstrated that the initiative signal can be conducted over consider- 



