Phycomyces

This monographic review on a fungus is not addressed to mycologists. None of the authors has been trained or has otherwise acquired a general proficiency in mycology. They are motivated by a common interest in the performances of signal handling exhibited by the sense organs of all organisms and by the desire to attack these as yet totally obscure aspects of molecular biology by the study of a microorganism with certain desirable properties. The sporangiophore of the fungus Phycomyces is a gigantic, single-celled, erect, cylindrical, aerial hypha. It is sensitive to at least four distinct stimuli: light, gravity, stretch, and some unknown stimulus by which it avoids solid objects. These stimuli control a common output, the growth rate, producing either temporal changes in growth rate or tropic responses. We are interested in the output because it gives us information about the reception of the various signals. In the absence of external stimuli, the growth rate is controlled by internal signals keeping the network of biochemical processes in balance. The external stimuli interact with the internal signals. We wish to inquire into the early steps of this interaction. For light, for instance, the cell must have a receptor pigment as the first mediator. What kind of a molecule is this pigment? Which organelle contains it? What chemical reaction happens after a light quantum has been absorbed? And how is the information introduced by this primary photochemical event amplified in a controlled manner and processed in the next step? How do a few quanta or a few molecules trigger macroscopic responses? Will we find ourselves confronted with devices wholly distinct from anything now known in biology

Functional Analysis of the Phycomyces carRA Gene Encoding the Enzymes Phytoene Synthase and Lycopene Cyclase

Phycomyces carRA gene encodes a protein with two domains. Domain R is characterized by red carR mutants that accumulate lycopene. Domain A is characterized by white carA mutants that do not accumulate significant amounts of carotenoids. The carRA-encoded protein was identified as the lycopene cyclase and phytoene synthase enzyme by sequence homology with other proteins. However, no direct data showing the function of this protein have been reported so far. Different Mucor circinelloides mutants altered at the phytoene synthase, the lycopene cyclase or both activities were transformed with the Phycomyces carRA gene. Fully transcribed carRA mRNA molecules were detected by Northern assays in the transformants and the correct processing of the carRA messenger was verified by RT-PCR. These results showed that Phycomyces carRA gene was correctly expressed in Mucor. Carotenoids analysis in these transformants showed the presence of ß-carotene, absent in the untransformed strains, providing functional evidence that the Phycomyces carRA gene complements the M. circinelloides mutations. Co-transformation of the carRA cDNA in E. coli with different combinations of the carotenoid structural genes from Erwinia uredovora was also performed. Newly formed carotenoids were accumulated showing that the Phycomyces CarRA protein does contain lycopene cyclase and phytoene synthase activities. The heterologous expression of the carRA gene and the functional complementation of the mentioned activities are not very efficient in E. coli. However, the simultaneous presence of both carRA and carB gene products from Phycomyces increases the efficiency of these enzymes, presumably due to an interaction mechanism

A Carotenogenic Enzyme Aggregate in Phycomyces: Evidence from Quantitive Complementation

Wild-type Phycomyces blakesleeanus accumulates β-carotene, while the mutant strain C2 is unable to synthesize carotenoids, and the mutant strain C9 accumulates lycopene. Heterokaryons containing a proportion, p, of C2 nuclei and (l — p) of C9 nuclei accumulate lycopene, γ-carotene, and β-carotene in the relative amounts (l — p), p(l — p), and p(2), respectively, for different values of p. It is shown that these results are expected from the operation of a carotenogenic enzyme aggregate that works as an assembly line and contains two copies of a cyclase, which is defective in strain C9, as well as other enzymes