and I think they rush into fruiting so fast that 

 they do not have time to form prespore cells 

 proportionally in all instances. 



TILL: Will the spores that you get from 

 these transections give a normal organism? 



GREGG: Oh, I'm sure they would. 



MASSARO: To go back for a minute. What 

 did you mean by uniform distribution? Do you 

 mean X number of cells surrounded by Y number 

 of undifferentiated ones? 



GROSS: Yes. If there's another type of cell 

 that is neither prestalk nor prespore, they have 

 to be somewhere in the slug. Now, when you cut, 

 you can cut any part of it, and in principle, you 

 can get back the whole thing. 



MASSARO: You could have the third type 

 of cell anywhere. 



GROSS : That' s the point; they are anywhere. 

 They're a population of finite size. Now, as you 

 reduce the sizes of pieces you cut, the fraction 

 of the uncommitted cells relative to those cells 

 that have already differentiated is going to change 

 depending on where you cut. If you cut in the 

 anterior end, you're going to have a large num- 

 ber of prestalk cells and a very small number 

 of prespore cells; and you still have a small 

 number of undifferentiated cells. There's no 

 replication, so you've got a large number of 

 prestalk cells that can't go anywhere and no 

 prespore cells; now, the small number of un- 

 committed cells must, in that instance, all 

 differentiate to form prespore cells. Suppose 

 you don't have enough. It seems to me that as 

 the piece gets smaller, like 7 cells, you're not 

 going to have enough of those relatively uncom- 

 mitted cells. 



MASSARO: Well, maybe these cells have 

 only a certain degree of noncommittedness. 

 Maybe we're looking at the noncommitted cells 

 too harshly and saying we have a cell here which 



is definitely noncommitted. Maybe certain pre- 

 stalk cells are less committed than other pre- 

 stalk cells. 



GROSS: This is an argument that extends 

 far beyond the slime molds. It's one that has 

 plagued embryologists for many years. 



KAHN: Jim, did you look at Polysphon- 

 dylium at all? 



GREGG: No, I didn't. 



KAHN: Well, this might be worthy of men- 

 tion along these lines. If you do the same sorts 

 of things that Gregg has done with fluorescent 

 technique with various histochemical techniques, 

 you do get a differential staining between the 

 presumptive stalk and the presumptive spore 

 areas. This is true in Dictyostelium discoideum, 

 also. 



GREGG: The presumptive stalk region is a 

 very small area. 



KAHN: I was going to get to that. The in- 

 teresting thing about Polys phondy Hum is that 

 you don't see these differences until very late; 

 so, in effect, the whole mass is uncommitted 

 until the very last moment. 



GREGG: You can differentiate between the 

 types of cells in a number of ways: PAS stain- 

 ing, vital stains, antibodies. 



PERSON: Is there a vital stain that can 

 differentiate between the two types of cells so 

 that you could keep an individual cell alive and 

 look at it? 



GREGG: Yes. Bonner's used stains such as 

 Nile blue sulfate, neutral red and Bismarck 

 brown. 



GRUN: Do they all produce a darker stain- 

 ing in the nonstalk area and a lighter staining 

 in the stalk area? 



GREGG: I believe the staining is more in- 

 tense in the anterior end with most of these 

 stains. 



ACKNOWLEDGEMENTS 



The meticulous histological preparations 

 which were made by Mrs. Doris Gennaro during 

 the course of this investigation are gratefully 

 acknowledged by the author. 



This investigation was supported in part by 

 a Public HealthService Career Programs Award 

 5-K3-HD-15, 780 from the National Institute of 

 Child Health and Human Development, Research 

 Grants E-1452 and GM-10138 from the National 

 Institutes of Health. 



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