Discovering Protein-based Inheritance through Yeast Genetics

I will describe here the way in which I was able to discover that yeast has prions (infectious proteins), proteins that are the carriers of genetic information, and thus are acting as genes (1). This startling finding involved inventing new genetic approaches, but was possible because of my particular background and interests. Our work has broadened the “prion” concept (originally thought to be restricted to a special mammalian disease), established that there are such things as genes made of protein, and led to an understanding of how proteins can encode and transmit heritable information. My undergraduate degree was from Cornell University in mathematics. This early interest proved important in guiding me into genetics, which I have always viewed as the “logic of life” and, more recently, as I have become involved in solid-state NMR studies. I received an M.D. degree from Georgetown University, and, following a medical internship, I became a postdoctoral fellow with Herb Tabor at the National Institutes of Health (NIH). This was my first real experience doing science, and Herb's critical, careful attitude toward research was my guide in establishing my own approach. I found that adenosylmethionine decarboxylase, the product of which is an intermediate in spermidine biosynthesis, has a pyruvoyl residue as a prosthetic group (2) and that histidine ammonia-lyase has a similar prosthetic group, dehydroalanine (3). As a postdoctoral fellow with Jerry Hurwitz at Albert Einstein College of Medicine in New York, I learned about nucleic acid enzymology from “The Boss” and worked on Escherichia coli DNA polymerase II and in vitro DNA replication (4, 5). I was impressed that the dnats mutants, isolated by Jacob, Bonhoeffer, Carl, Wechsler, and others, were the key to the biochemistry of DNA replication, coupled with in vitro replication systems selected to require the dna gene products (6). In vitro complementation could be used to purify the proteins shown by in vivo studies to be responsible for the biochemical reaction. These experiences having provided me with a firm biochemical background, I then took the 3-week Cold Spring Harbor Laboratory course on yeast genetics, taught by Fred Sherman and Gerry Fink. I began my independent work in the laboratory of Jerry Hurwitz, who generously allowed me to start working on yeast genetics for a year until my job at the NIH began in 1973. I started out purifying DNA polymerases from yeast, similar to what I had been doing as a postdoctoral fellow with Jerry, but I decided that yeast genetics might be a better route to answering even many biochemical issues. I isolated mutants that could take up dTMP for labeling cellular DNA. These mutants were not widely used for this purpose, but proved quite interesting, particularly tup1 mutants, which had a mating-type α-specific mating defect and a sporulation defect (7) and proved to be a subunit of a transcriptional repressor (8, 9). The key role that bacteriophage had played in the development of an understanding of bacterial genetics led me to focus on the non-chromosomal genetic element determining the killer trait of yeast (10–12), then newly discovered to involve a double-stranded RNA (dsRNA) in virus particles (13–15). We isolated mutants in host genes necessary for propagation of the killer toxin-encoding M dsRNA (a satellite of the L-A dsRNA virus), mutants that could propagate M dsRNA but could not express the killer phenotype (kex mutants), and superkillers (ski). In an era before yeast cloning, we genetically mapped them as a means of identification.