M. WESTERGAARD AND H. HIRSCH 



on P-minimal medium. After 2 weeks, however, when tyrosinase activity in mycelia 

 grown on P-minimal medium went down, the activity was very much increased on 

 the AA-substrate, and now the pigments turned black during the enzymatic test. 

 No protoperithecia developed on the AA-mycelium with 3,000 mg./l., nor was 

 melanin formed during actual growth. At a lower concentration (1,500 mg./l. ), 

 protoperithecia were formed after 2 weeks, but they could not be fertilized. 



Hirsch made further experiments to test the hypothesis that there is a connexion 

 between tyrosinase and differentiation. A number of tyrosinase inhibitors (^-amino- 

 benzoic acid, sodium-thioglycolic acid, cystein and phenylthiourea) were added to 

 P-minimal medium. They were all able to suppress protoperithecium formation more 

 or less completely, whereas other enzyme inhibitors without specific affinity towards 



TIME (DAYS) 



Figure 3. Tyrosinase activity and protoperithecium forma- 

 tion as a function of time in Neurospora crassa strain 

 W 2/49 A, grown on P-minimal medium at 25 C. 

 Curve 1 : tyrosinase activity of ground suspensions ; net in- 

 crease in optical density X 1 00 in 30 minutes. Curve 2 : 

 ditto; net increase in optical density X 100 in 60 minutes. 

 Curve 3: number of protoperithecia. (Hirsch, 1954.) 



the tyrosinase system (e.g. streptomycin and others) did not interfere with the differ- 

 entiation process. 



It might be mentioned at this point that the present differentiating system has 

 certain advantages over the growth-system studied by other workers: it is possible 

 to study the metabolism of mycelia in which differentiation is blocked. The same, 

 however, cannot be done with mycelia in which growth is blocked, because the 

 alternative to growth is lethality. Thus, from studying the metabolism of mycelia 

 in which differentiation was blocked by various environmental factors, it has been 

 possible to get information about the biochemistry of differentiation even without 

 bringing mutant strains into the picture. 



However, the relationship between tyrosinase activity, melanin formation and 

 development of female sex organs is far more complicated than might appear from 

 the data hitherto presented. In recent years tyrosinase activity in Neurospora has been 

 studied by Fox and Gray (1950), and especially by Horowitz and his co-workers 

 (Horowitz and Shen, 1952; Horowitz and Fling, 1953; see also Horowitz, 1951). 



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