My Ph.D. thesis produced the first reliable information concerning the rate of the helix-coil transition for small DNA--0.1 to 3kb. Surprisingly, all but the smallest DNA initiated with internal loops. Temperature-jump and stopped flow kinetics were used. To do these studies, I was the first to isolate monodisperse (similar molecular weight) DNA in this size range.

I then went to Caltech to study the avoidance problem with the late Max Delbruck. Phycomyces senses and bends away from nearby objects. Trying to discover the mediation of this response was the most intellectually challenging scientific problem any of us ever faced. It also was the most fun. To this day, no one knows how this "primitive" organism does it.

At the University of Florida, I initiated several lines of biochemical and biochemical genetic investigations of Phycomyces. We studied the chemistry of the cell wall and the enzymology of chitinase and chitin synthetase to try to obtain a clue about the regulation of cell growth. Related studies were done on beta glucosaminidase, alkaline phosphatase and calmodulin. David Lapointe's dissertation showed that ornithine decarboxylase is important for growth and is blue-light regulated.

We also showed that cAMP and cGMP levels were quickly changed by blue light and that mutants not responding to light also lacked these changes. Cyclic nucleotide phosphodiesterase and adenylate cyclase were characterized and were shown to be altered by blue light. These latter investigations led to our recent work on G proteins.

Our focus in the molecular aspects of phototransduction in the model system Phycomyces blakesleeanus then shifted to G-proteins as putative mediators of the response. The 4 cm long fruiting body or sporangiophore of Phycomycescan detect the equivalent of a candle ten miles away. It adapts to lowering and raising light levels. It is the prototype for the blue light responses of plants and fungi.

    We have found evidence  for the presence of several plasma membrane  bound G-proteins: 1) GTPgammaS binding, 2) cholera toxin binding, and 3) Western blotting.   In addition, Northern analysis indicates several Galpha mRNA.  RT-PCR has been used to isolate and sequence  three independent gene fragments of  Galpha species.  They are similar to Galphas from other light senstive filamentous fungi and to transducin and gustucin. We are currently extending the sequencing and searching a cDNA library to isolate and sequence as many sporangiophore Galpha genes as feasible. We have available several mutants of Phycomyces unable to detect low light levels. A careful analysis of proteins by Western blotting, mRNA by Northern blotting and GTPase function should identify the specific Galpha involved in this blue light response.

   The sporangiophore of  Phycomyces  may also use serotonin.  Serotonin is a well known neurotransmitter in vertebrates. We have evidence that serotonin (5-hydroxytryptamine) exists in the sporangiophore at a level of approximately 1-3 ug/g, that plasma membrane from sporangiophore binds to radiolabeled serotonin and that serotonin enhances GTPase about 80%.
 
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