(Fig, 3). You cannot detect that prestalk cells 

 have differentiated at this stage. In the late 

 aggregate prestalk cells begin to differentiate 

 (Fig. 4). These prestalk cells are characterized 

 by the fact that they tend to lose their cyto- 

 plasmic antigens. Consequently, they do not 

 stain with high intensity. At the same time you 

 see spots of intense staining in the prespore 

 cells which mark the synthesis of prespore 

 antigen. Consequently, all the cells in this area 

 form prespore cells and the anterior cells which 

 stain the least become prestalk cells. 



Figure 5 shows a migrating pseudoplas- 

 modium of D. mucoroides. This has been stained, 

 however, with the normal conjugated antiserum. 

 Little or no staining was found with normal con- 

 jugated serum. The preparation itself tends to 

 transmit light in such a way that it appears to 

 be bright, but the fluorescent staining is rela- 

 tively low. 



LOVETT: Is that region in the center 

 the stalk? 



GREGG: Yes. Figure 6 is another D. muco- 

 roides slug stained with the antiserum. You can 

 see that stalk formation is occurring; the stalk 

 runs down through the center of the slime mold. 

 The prestalk cells in the anterior area are fully 

 differentiated now, resulting in the formation of 

 a proportional slug. The prespore antigen in- 

 creases in the prespore cells throughout the 

 entire area. This results in a sharp delineation 

 between the prespore cells and the prestalk 

 cells. Thus, by this time these two types of cells 

 have developed with the prestalk cells always 

 in the anterior or leading end of the slug. The 

 question arises, how does this polarity develop? 

 Takeuchi has suggested that the bright-staining 

 and dark-staining cells that he found - and that 

 I have seen - in the aggregating streams sort 

 out during aggregation. Simultaneously, the 

 dark-staining cells lose even more of their 

 staining and eventually end up in the anterior 

 tip, thus composing the prestalk area. Conse- 

 quently, the brightly- staining cells form the 

 prespore area. 



MASSARO: What is the magnification here? 



GREGG: That's about 120X. 



DEERING: What's that film along the edge 

 of the slug in some figures? Is it something that 

 peeled off? 



GREGG: It's a slime track or slime sheath 

 that's produced along the edges of the slime 

 mold. 



Now, it's possible that bright-staining and 

 dark-staining cells sort out to form these two 

 areas. However, the slug has developed propor- 



tionally, and in order to account for this we 

 would have to assume that the prestalk and the 

 prespore cells differentiated during the aggre- 

 gation stage and aggregated in numbers suitable 

 to form this proportionality in a cell mass of a 

 certain size. It's a little difficult to conceive of 

 this occurring. It would seem more obvious that 

 proportionality results after the cells come 

 together. However, since these cells can revers- 

 ibly differentiate, it's possible that they differ- 

 entiate in either direction, depending upon the 

 necessity, in order for the proportionality to be 

 established. Before continuing, however, in this 

 discussion let's look at the situation in another 

 system, D. discoideum. 



TS'O: Excuse me for asking a question on 

 the biology of this organism. Can you take a 

 single cell and generate a mass like this or do 

 you have to always start with lots of cells? 



GREGG: Yes, it is possible. Either a single 

 mature spore cell or a single amoeba will pro- 

 duce innumerable colonies. 



In D. discoideum we have also been bright- 

 and dark-staining cells in the aggregating 

 stream. However, in the early aggregate we 

 again see no evidence that the prestalk cells 

 have differentiated (Fig. 7). This upper margin 

 is what we call an edge effect, which you get in 

 certain fresh preparations. This artifact does 

 not represent the differentiation of prestalk 

 cells. 



Figure 8 shows a late aggregate of D. dis- 

 coideum. This is the orientation of a late aggre- 

 gate on an agar plate. They stand up just prior 

 to flopping over and migrating about on the agar. 

 Even at this relatively late stage one usually 

 cannot see a differentiation of prestalk cells. 

 On occasion there is a small tip end of prestalk 

 cells which have differentiated, but otherwise 

 the cell mass appears to be uniformly stained. 

 It's obvious that the form and the polarity of the 

 cell mass is independent of the differentiation 

 of the prestalk cells. Thus, prestalk cells need 

 not differentiate in order to produce this par- 

 ticular shape. Consequently, this suggests that 

 subtle differences exist in the cell mass, prior 

 to prestalk and prespore cell differentiation. 

 Now, I suggest that one of these subtle differ- 

 ences is that of acrasin production which is at 

 its greatest intensity in the anterior tip. Bonner 

 (4) has shown this in D. discoideum. Perhaps 

 such differences as this result in the differen- 

 tiation of the cell according to the point at which 

 it happens to be located. 



Immediately after the late aggregate it is 

 obvious that prestalk differentiation has oc- 



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