BioEssu~~s Vol. 9. NO. 4 - October 1988 129 Molecular Genetics, Microbiology, and Prehistory JuL I4 w Bernard D. Davis Since the literature of molecular genetics a radical advance provided by bacterial has been growing explosively, it is small genetics : while classical genetics could xonder that those who were not present only count the various phenotypes in a at the creation of the field may have a limited population of progeny, appro- cloudy picture of its origins: events of priate selective media for bacteria, or only five years ago are already ancient selective hosts for their viruses, could be history. But a recent reprinting of the used to isolate even very rare mutants autobiography of Emil Fischer has or recombinants from an enormous i brought to light an even earlier pre- population. The resolving power of j history, in which a remarkable specu- genetic mapping was thus suddenly lation anticipated the possibility of refined by many orders of magnitude: genetic engineering. To appreciate this mutations and recombinations, pre- contribution we must first consider the viously localized only in terms of genes, obstacles that so long delayed the union could now be localized in terms of ( of biochemistry and genetics. individual nucleotides. i The Role of Bacterial Genetics : Everyone knows, of course, that the : discovery of the double helix by Watson 1 and Crick in 1953 gave birth to mo- lecular genetics. But the field had its , conception - the zygote that united its previously disparate elements - in the ! chemical identification of the pneu- , mococcal transforming principle by I Avery, MacLeod and McCarty in 1944. . Moreover, this discovery not only es- tablished DNA as the material of the ! gene: it also launched the field of i bacterial genetics. For genetic studies, ) until the recent identification of gene I sequences, had depended on the re- ; combination of different alleles from . different parents. Transformation first - demonstrated that this process occurs . U-I bacteria, and it quickly led to , Lederberg's recognition of two addi- tional mechanisms, conjugation and ~ transduction. i The importance of bacterial genetics I for the development of molecular ge- netics cannot be overemphasized. The t work of Beadle and Tatum with auxo- trophic mutants of the mold Neuro- ., spora not only provided a novel set of 1 genetic markers in a single-celled or- , ganism; it also indicated that each gene I forms a corresponding enzyme ; and 1 similar mutants of bacteria soon pro- b vided a wide variety of phenotypes that invited similar biochemical correlations. ;M oreover, while recognition of the 1 double-helical structure of DNA was ! based on biochemistry and X-ray crys- tallography. exploration of the impli- " cations of that structure benefited from Possible Reasons for the Neglect of the Avery Discovery The Avery discovery was truly revolu- tionary, not only because of its in- trinsic significance, but because the answer was so unexpected. Before then it was generally assumed that only proteins could provide the complexity required for the gene, and so the DNA in the chromosomes was presumably providing some kind of scaffolding. Yet Avery and his colleagues did not receive a Nobel Prize, though he lived for II years after announcing the role of DNA. Neither was a Nobel Prize awarded for the development of the fine-structure genetics by Benzer, Yan- ofsky and Brenner which made it possible to correlate specific changes in DNA sequence with changes in protein sequence and with altered function in the cell. It may be hard to appreciate today that this discovery-that cros- sing-over can occur between any ad- jacent bases, rather than at special sites between genes - had the two elements of a great discovery : surprise, as well as broad significance. Perhaps the problem here was that fine-structure mapping, and the collinearity of DNA and pro- teins, so quickly became taken for granted as a foundation on which so many built. Since the Nobel Committee has on the whole shown excellent judgment, except in the area of medical therapy (for example, Finsen's prize for al- legedly curing skin tuberculosis with ultraviolet irradiation, and Munoz's for prefrontal lobotomy). we must wonder why they missed Avery. Indeed, this was clearly one of their most egregious errors (though some would consider bypassing Freud's impact on literature, if not on medicine and psychology, an even greater omission). Here I would like to suggest several reasons for the slow recognition of Avery and his colleagues. " (I) A small part of the responsibility rests on Avery himself, for while his unique style was something to admire, it was not ideally designed to draw attention to such a revolutionary dis- covery. Indeed, it was antithetical to the inimitable highly competitive, impatient style set by the subsequent leaders in molecular genetics, continually shifting the direction of research as new peaks were revealed for conquest. Avery in- stead devoted his entire lifetime to the patient study, with few collaborators and little sense of competition, of factors affecting the virulence of one organism, the pneumococcus - the major cause of death in the developed parts of the world at the time when he began. What is even more relevant was his style of publication, which would clearly be very difficult to emulate in today's era of intense competition and constant pressure to justify renewal of grants. My teacher, Dubos, told me that Avery would explore a problem at length and finally, when he had the answer, would not publish these data but would per- form the `protocol experiment', with the precise number of points and con- trols needed to document his con- clusions. Moreover, he would then put the paper in the drawer for a few months in order to be able better to polish it before publication -and the yield was at most two to three papers per year. In addition, he was opposed to short notes: it did not escape his attention that the DNA discovery had deep genetic implications, which he expressed most tentatively; but he pub- lished it only as a full paper in the Journal of Esperittletttal Medicine, with- out trying to draw attention to its general significance in such a journal as Nature. A few years later, visiting what was then the world center of research on biochemical genetics, Beadle's de- partment at CalTech. I found that its 130 BioEssays Vol. 9, No. 4 -October 1988 ROOTS library, understandably, did not carry the Journal of Experimenfal Medicine. (2) The cleft between genetics and bacteriology was an even deeper pro- blem. No chromosome had yet been seen in bacteria. Moreover, genetic adaptation in these organisms was gen- erally confused with physiological ad- aptation, because overnight outgrowth of a variant seemed too rapid for a Darwinian process (until the power of rapid exponential growth and sharp selection was later understood). Ac- cordingly, the study of bacterial vari- ation was not yet linked to genetic concepts: inheritance in bacteria was generally ascribed to a vague plasticity, in the absence of evidence for the linked genes found in higher organisms. The `medical ' basis of Avery's dis- covery further deepened the cleft. At that time bacterial capsules were studied only as virulence factors, and not as products of metabolic pathways; hence there was no basis for interpreting their formation in terms of specific enzymes, which might have linked transformation to the biochemical genetics recently initiated by Beadle and Tatum. Hotch- kiss's subsequent transformation of more conventional biochemical traits eventually eliminated this barrier, but transformation probably long con- tinued to seem strange to most bio- chemists. The pneumococcus was then also very strange to most Drosophila- and maize geneticists; it was not obvious, as it is now, that the evolutionary con- tinuity of bacteria and eukaryotes im- plies shared common basic features of inheritance. To be sure, two insightful geneticists, George Beadle and Herman Muller, quickly pointed out in reviews the potential significance of pneu- mococcal transformation for their field; but their impact does not seem to have been large. (3) A more important reason for the delayed general appreciation of Avery was the unexpected nature of his conclu- sion, which inevitably generated skep- ticism. Probably the greatest source of this skepticism, as candidly revealed in Maclyn McCarty's modest history of the discovery,' was a colleague at the Rockefeller Institute, Alfred Mirsky, who had been concentrating on chro- mosomes and nucleic acid for many Years. He clearly did not enjoy being upstaged by these three medical micro- biologists, all without a background in genetics, and self-taught in their bio- chemistry. What is more, Mirsky was an urbane, widely traveled man, and he gave many seminars emphasizing that the activity of even the purest DNA preparations might be due to con- taminating protein. (It took painstaking experiments by Hotchkiss to show that the traces of amino acids, always present in hydrolysates of the DNA, were products of breakdown of purines.) Avery, who never went to meetings or traveled to give seminars, simply waited for Nature to settle the controversy. This it did - but too late for the Nobel Committee. (4) An additional factor was that genetics was then a very specialized field, and a member of the Nobel Committee later explained to me that it was hardly represented at ail in Sweden. And despite the aura of immortality surrounding the Nobel Prizes, the finite interests and background of the mortals composing the awarding committees obviously affect the selection. (5) The role of phage geneticists in the skeptical reaction has elicited a good deal of controversy. This brilliant group had set out to study phage because this simplest of all organisms seemed most likely to reveal the nature of the gene. Since theirs was a much more logical approach than the ser- endipitous one that worked for Avery, it is understandable that they could easily find grounds for doubt about the genetic significance of his discovery. Only in 1952 did the phage group place its imprimatur on Avery's conclusion, when Hershey and Chase showed that labeled DNA of an infecting phage entered the host cell while the dif- .ferentially labeled protein remained out- side. But while this pioneer phage experiment was a most important one, it was not nearly as clean as the Avery experiment: the entry of the DNA was accompanied by about 20% of the phage protein, while the purified pneu- mococcal factor contained no detectable protein, and the activity was destroyed by DNase but not by protease. (6) As has often been pointed out, the Watson-Crick discovery of DNA structure had a tremendous and im- mediated impact because its functional implications - for both gene replication and mutation - were obvious, while the Avery discovery implied only that DNA was important. But that does not explain why so few people took up that provocative lead in the next few year This was a classical case ofconservatisr in scientific fashions, perhaps parti explained by the other features of tt Avery discovery that I have just de: cribed. Emil Fischer's Speculation about Genetic Engineering These ruminations on early histor cover ground that will be familiar t many readers. But I would like i addition to call attention to a Ies familiar speculation by Emil Fischer i 1914, foreshadowing genetic engineer ing. Fischer was a giant who gave u much of what we know about th organic chemistry of sugars, peptides lipids and nucleic acids. Springer Verla, has just republished his posthumou autobiography, Aus meinem Leger: with a scholarly prologue by Bemharc Witkop of the NIH. Witkop notes thi following passage, where Fischer2 wa, discussing the methylated purines tha he had been synthesizing: With the synthetic approaches to this groq we now are capable of obtaining numerou: compounds that resemble, more or less natural nucleic acids. How will they affec- various living organisms? Will they TV rejected or metabolized or will they partici- pate in the construction of the cell nucleus' Only the experiment will give us the answer I am bold enough to hope that, given tht right conditions, the latter may happen ant that artificial nucleic acids may be assimi lated without degradation of the molecule Such incorporation should lead to pro- found changes of the organism, resemblin: perhaps permanent changes or mutations a: they have been observed before in nature. Of course, this prophetic speculation. not yet ripe for testing, fell by the wayside and was not a contribution toward the development of genetic engineering. Nevertheless, it reminds us that highly intelligent individuals, deeply immersed in an area of science, can offer judgments that are remarkably far-sighted. REFERENCES 1 MCCARTY. M. (1985). ?h Transforming Princ:plr. Norton. 2 FISCHER, E. (1914). Berichre 477. 3196. Medical School, Boston. M.4 02 I I 5, L:S.-I.