Topic 4.2
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Induction and the Origins of Developmental Genetics
Scott F. Gilbert
From A Conceptual History of Modern Embryology (S. F. Gilbert, ed.) Plenum Press, NY. pp. 181-206.
Reprinted here with the kind permission of Kluwer Academic Press, Dordrecht, The Netherlands.
If ever a history of ideas in developmental genetics were to be written...it would no doubt include as one of its most important chapters an account of the intellectual role that "inductive interaction" between the fields of genetics and embryology has played in the analysis of developmental mechanisms and their genetic control in higher organisms.
-Salome Gluecksohn-Waelsch (1981)
...the outsider sees most of the game.
-Conrad Hal Waddington (1968)
1. Introduction: The Problem of Synthesis
At the turn of the past century, the field of heredity included embryology, regeneration, and genetics. Discussions of genetics necessarily entailed a theory of development, and any theory of development had to show how the eggs of different species developed in different ways. Thus, the "hereditary" theories of William Keith Brooks (1) or August Weismann (2) did not distinguish separate genetic and embryological domains. The developmental mechanics of His, Roux and Driesch likewise contained explicit genetic components whereby the hereditary determinants (thought to reside either within the cytoplasm or inside the nucleus) were seen to direct the processes of organ formation and cell differentiation.
The split between genetics and embryology emerged gradually, largely through the investigations of Thomas Hunt Morgan and his laboratory (3,4). Whereas most American and German experimental embryologists followed Boveri in thinking that the nucleus was the site of the hereditary determinants, Morgan was convinced that these determinants lay in the cytoplasm. Morgan had collaborated with Driesch on a project that involved the removal of cytoplasm from the uncleaved ctenophore egg (5). The result of such operations were defective embryos. Morgan declared that there was "no escape from the conclusion that in the protoplasm and not in the nucleus lies the differentiating power of the early stages of development." However, in 1905, his close friend, E. B. Wilson, and his former graduate student, Nettie Stevens, both provided evidence that the nucleus did indeed contain the determinants of genetics and development. They both correlated the XX chromosome composition with female animals and the XO or XY chromosome complement with male animals (6). If this were true, then the nucleus determined the sex of the individual.
Morgan responded by investigating a parthenogenetic species of aphids, eventually correlating chromosome number and sex. However, he interpreted his results as still being consistent with the cytoplasm having the controlling role in development (7). However, by 1910, Morgan had found mutations in Drosophilathat could be best interpreted as segregating with the X chromosome. Although he initially resisted this interpretation, he eventually came to see the genes as physically linked on the chromosomes. What had begun as an investigation as to whether the nucleus or the cytoplasm controlled development ended in the founding of the gene theory.
Immediately after 1911, genetics arose as a discipline within experimental embryology; but it soon evolved its own techniques, favored organisms, rules of evidence, and specialized vocabulary which separated it from the rest of embryology. Eventually, it acquired its own sources of funding and new journals (4,8). In his 1926 book, The Theory of the Gene, Morgan (9) formalized the split by declaring that genetics dealt exclusively with the transmission of hereditary traits, while embryology concerned the expression of those traits. He claimed that "the sorting out of characters in successive generations can be explained without reference to the way in which the gene affects the developmental process," and that confusion had arisen "from confusing the problems of genetics with those of development." Genetics and embryology began to go their separate ways.
But Morgan remained an embryologist, publishing Experimental Embryology the year after The Theory of the Gene. When he left Columbia University to head the Biology division at the California Institute of Technology, he returned to study the problems of ascidian development. Thus, when Morgan publishedEmbryology and Genetics in 1934, many biologists hoped that it would reunite these disciplines. This was not to be the case. It was more a joint textbook than an attempt to resynthesize the field. The synthesis would be left for others to create. In 1939, Richard B. Goldschmidt and Ernest E. Just published their respective attempts to unify the fields. Goldschmidt would have had embryology subsumed under genetics, while Just saw genetics as a rather minor subset of embryology (4). At the same time, at least three other researchers, Salome Gluecksohn-Schoenheimer (later S. Gluecksohn-Waelsch; 1907-), Conrad Hal Waddington (1905-1975), and Boris Ephrussi (1901-1979) were attempting more balanced syntheses of the two disciplines. Burian, Gayon, and Zallen (this volume) have written an account of Ephrussi's synthesis of embryology and genetics. This paper will focus on the work of Gluecksohn-Schoenhimer and Waddington.
This paper is framed by two questions: First, what were the conceptual foundations of developmental genetics and how did they come into existence? Second, how did developmental biologists learn about the operon model of microbial gene regulation which was soon to become the major paradigm of developmental genetics? Evidence will be presented that the conceptual foundations of developmental genetics originated by researchers who were trained in experimental embryology but who met with frustration in their attempts to solve questions of development using embyological techniques. In particular, much of what we call developmental genetics today was framed by Salome Gluecksohn-Schoenheimer and C.H. Waddington, two researchers who had been interested in "the organizer problem." Gluecksohn-Schoenheimer focused her research on using mutants to elucidate normal developmental pathways. Waddington's focus on embryonic cell competence caused him to propose the notion of canalization and its application in genetic assimilation. Later, this same phenomenon of competence caused him to focus on the cytoplasmic regulation of the genome and predisposed him to see the operon as a model for embryonic induction.
Salome Gluecksohn-Schoenheimer (Waelsch)
C. H. Waddington
2. SALOME GLUECKSOHN-SCHOENHEIMER AND THE PATH FROM EXPERIMENTAL EMBRYOLOGY TO DEVELOPMENTAL GENETICS
2.1 Early work on the T-locus: The search for inducer genes
The path from experimental embryology to developmental genetics—from the Freiburg laboratory of Hans Spemann to the Columbia University of Leslie C. Dunn—was first travelled by Salome Gluecksohn-Schoenheimer. Having studied chemistry and zoology at Konigsberg and Berlin, she asked Spemann to be his graduate student in 1928. She does not consider it a happy partnership. "Our first meeting made it quite clear that we were not meant for each other, but I suppose Spemann did not have enough courage to turn me down outright." Spemann, who Gluecksohn-Waelsch recalled as being prejudiced against women working in his laboratory, would not let her work on the exciting projects involving the Organizer. Instead, he gave her "a rather boring descriptive study of limb development" for her Ph.D. dissertation research (10).
Spemann, like many other embryologists of his day, had no interest in the new science of genetics. However, although he did not believe that genes played any major role in embryonic development (11), two members of his laboratory did perceive that genetics had some critical things to say concerning how organisms developed. One of these was Spemann's assistant, Viktor Hamburger. Hamburger supervised Gluecksohn-Schoenheimer's thesis and "was the only one who provided us students with some introduction to the principles of genetics." The second person was Conrad Hal Waddington, a Cambridge graduate student who came to Germany in 1932 to study Organizer phenomena and to learn the techniques of tissue grafting. He became one of Gluecksohn-Schoenheimer's closest friends, and the two of them had several discussions concerning the possible roles of genes in development. Gluecksohn-Schoenheimer decided that when she completed her dissertation, she would attempt to uncover the roles that genes played in the development of the embryo.
But neither Hamburger, who was studying the innervation of embryonic limbs, nor Waddington, who was trying to isolate the molecules responsible for Organizer function, were truly geneticists. To study animal genetics in Germany meant studying in Richard Goldschmidt's laboratory at the Kaiser Wilhelm Institute in Berlin-Dahlem. So in 1932, as she finished her dissertation, Gluecksohn-Schoenheimer went to see the "Lieber Gott von Dahlem." She recounts that she was unable to see him, however, because Goldschmidt's assistant, Curt Stern, told her that it would be useless teaching genetics to a Jewish woman. She did not see Goldschmidt until years later when, fleeing the Nazis, he came to New York City (11).
In 1933, Gluecksohn-Schoenheimer and her husband, the noted physiologist Rudolph Schoenheimer, fled to America. Hamburger and Stern left the same year, eventually followed by Goldschmidt in 1936. While her husband had an appointment in Columbia University's College of Physicians and Surgeons, Salome Gluecksohn-Schoenheimer worked as a technician in the laboratory of Samuel Detwiler. This was an obvious place to work since Detwiler was interested in those problems of limb innervation which the Freiburg laboratory had helped identify. This employment did not last long. Gluecksohn-Schoenheimer recollects that she met geneticist Leslie C. Dunn at a cocktail party where he told her of his recent work on mutations of vertebrate development. Dunn had obtained a mouse strain with a dominant mutation (T; Brachyury) for short tails. Work by Dunn and his graduate student Paul Chesney had shown that the heterozygous (T/+) condition resulted in a shortened tail due to a constriction in the neural tube. Homozygous embryos died at 11 days in utero with the posterior half of their body missing. Chesney's work pointed to an earlier defect in the notochord as being responsible for the lack of a posterior neural tube. In short, it appeared that the T mutation was involved in axial determination. It was even possible that the wild-type T gene controlled the posterior inducer substance of the notochord, itself. But the project was incomplete. Paul Chesney had committed suicide.
The project was ideal for Gluecksohn-Schoenheimer. Forbidden by Spemann to work on the central problems of axial determination in amphibians, she could now study them in mice. Unable to get genetic training in Germany, she could now learn the most modern genetics in the very birthplace of the gene theory. Moreover, she found that she might be working on one of the most important genes of them all—a gene responsible for the posterior organizing substance of the mammalian embryo. Gluecksohn-Schoenheimer accepted Dunn's invitation to work in his laboratory, even though it meant working without pay for a year.
The first papers on the T-locus mutants make it clear that Gluecksohn-Schoenheimer interpreted these phenotypes as being caused by a genetic defect in the induction of the posterior neural tube by the notochord. In 1938, she concluded (12) that "our data do not give conclusive evidence for conceiving the malformations of the neural tube as secondary to the disorders of the notochord, but they point in this direction." By 1940, Gluecksohn-Schoenheimer was able to state more assuredly (13) that "in the heterozygous Brachy (T+) mouse the notochord in the posterior region of the embryo is defective and as a result the Brachy phenotype develops."
In doing these studies on the T-locus, Gluecksohn- Schoenheimer made a virtue out of necessity and founded the first version of developmental genetics. Unable to manipulate the mammalian embryo inside the uterus and placenta, she had looked to nature's own experiments. In the introduction to her first paper on the tailless mice (14), Gluecksohn-Schoenheimer presented the first programmatic statement of developmental genetics, distinguished the activities of the "developmental geneticist" from those of the "experimental embryologist," and gave a rationale for the emergence of developmental genetics out of experimental embryology.
First, one could not study mammalian development as one had studied amphibian embryogenesis. "It is not possible yet to use transplantation, isolation, or vital staining methods on mammalian embryos as they have been used on amphibian embryos." Quoting Spemann (15) in the negative sense, she wrote, "For the present, however, the experimenter is not able 'to alter the course of events at a chosen point in a chosen manner and draw conclusions on their relations from the resulting changes'." Spemann's dictum for experimental embryology would not hold for the study of mammalian development. Gluecksohn-Schoenheimer's first paper on the subject began, then, in the context of Spemann's studies, but found his methods unusable.
Second, instead of manipulating the embryo and seeing its effect on the phenotype, Gluecksohn-Schoenheimer proposed to look at the phenotypes produced by mutant genes and relate them back to their embryologic causes. "A mutation that causes a certain malformation as the result of a developmental disturbance carries out an 'experiment' in the embryo by interfering with the normal development at a certain point. By studying the details of the disturbed development it may be possible to learn something about the results of the 'experiment' carried out by the gene." In this attitude, she was closer to Boveri and Goldschmidt than to Spemann. Moreover, this program bridged the gap (as both Boveri and Goldschmidt tried to do) between genetics and embryology. Most American embryologists in the late 1930s did not think that genes acted during the early stages of development (4,8,16).
Third, Gluecksohn-Schoenheimer declared that this type of research was to be done by a new type of scientist, the developmental geneticist.
While the experimental embryologist carries out a certain experiment and then studies its results, the developmental geneticist first has to study the course of the development (that is, the results of the developmental disturbance) and can then sometimes draw conclusions on the nature of the 'experiment' carried out by the gene.
In this three-paragraph introduction, Gluecksohn-Schoenheimer had moved from experimental embryology to developmental genetics. To be sure, there was something like developmental genetics before Gluecksohn-Schoenheimer coined the expression. The subjects of Goldschmidt's "physiological genetics" would certainly fit into this catagory. But Goldschmidt refused to give credence to Morgan's theory of individualized genes and held to his own hypothesis that mutant phenotypes were caused by the timing of gene activity. In Gluecksohn-Schoenheimer, however, we see the merging of Spemann's embryological concepts (induction, regulation, etc.) with the particulate gene theory of the Morgan school—the merging of Freiburg and Columbia. The work of Landauer and Dunn on the Creeper mutation of chicks also precedes that of Gluecksohn-Schoenheimer, but the original analysis of Creeper was primarily physiological rather than developmental. The embryology of the Creeper mutants (17) wasn't accomplished by Landauer until 1944. With Gluecksohn-Schoenheimer, however, we have a programatic statement for developmental genetics as a discrete science with its own methodology.
2.2 The Genetics of Induction
Between 1938 and 1949, Gluecksohn-Schoenheimer pursued a research program explicitely linking embryonic organizers and specific genes in the mouse. The first series of these investigations looked at the interactions between dominant and recessive alleles at the T locus. The dominant T allele, as we have seen, was interpreted as affecting the notochord's ability to induce the neural tube. The recessive t-alleles, moreover, were interpreted as being involved in more general mesoderm-forming processes. In to/to embryos, the mesoderm failed to form at all, while in T/to embryos, the mesoderm and notochord of the posterior regions were seen to be defective. These mutant genes, however, were considered to be alleles at the same locus. How could one allele affect just the chordamesoderm while the other allele affect the other mesodermal areas? The answer to this genetic quandry was to be found in the embryology of the mouse (18).
The effect of the two alleles T and to on notochord and mesoderm might suggest that the two alleles act on two different structures. However, if considered from the embryological point of view, the notochord and mesoderm of the mouse have the same origin, namely in the tissue of the wall of the primitive gut.
This brought the problem back to what had been thought of as the mammalian equivalent of the dorsal blastopore lip. Indeed, Gluecksohn-Schoenheimer was trying to do with mutants what Spemann and the Mangolds had done by transplantations. As she would summarize in 1949, "The study of this material makes it very likely that in mammals the notochord plays a role in processes of early organization similar to that of the notochord in amphibians as analyzed with the techniques of experimental embryology" (19). More than that, Gluecksohn-Schoenheimer thought that she could do with the T-locus what Spemann's group and the Cambridge laboratory of Needham and Waddington could not do: Find the inducer molecule, itself. Gluecksohn-Waelsch (20) would later write, "It was therefore hoped that the identification of the mode of action of T-locus genes — and the nature of their gene products — might provide leads towards the molecular analysis of normal inductive mechanisms."
The T-locus alleles weren't the only mutations that appeared to control induction. The phenotypes caused by another, closely linked, mutation, Kinked(now abbreviated FuKi) were interpreted in terms derived directly from Spemann's work on amphibian embryonic regulation. Homozygous mutants of Kinked were found to have duplications of their dorsal axis, sometimes forming twin embryos (21).
Their striking resemblance to the double-monsters obtained by constriction experiments of amphibian embryos at the two-cell stage led to the suggestion that an "organizer" region analogous to that identified experimentally in amphibians existed in mammalian embryos and that its normal functioning was severely affected in FuKi /FuKi embryos... There is no doubt that all these interpretations of mutational effect on the developmental mechanism was strongly influenced by the orientation of the particular investigators and their view of development as depending on a series of inductive interactions.
Gluecksohn-Schoenheimer interpreted all three genes (T, to, and FuKi ) as disturbing "specific organizer relationships." She interpreted the action of theKinked gene as causing constrictions analogous to those done experimentally by Spemann and Holtfreter on salamander eggs (22). These famous studies had shown that the constriction of the egg down the medial plane caused the formation of two organizers, each of which formed embryonic axes, thereby creating twin larvae. Constriction in the frontal plane, however, caused the formation of one normal larva and one Bauchstüch, an amorphous tissue mass consisting chiefly of endoderm and blood cells. Partial constrictions, moreover, caused conjoined larvae, an observation that Spemann had related to mammalian teratology (23).
According to Gluecksohn-Schoenheimer, the Kinked mutants had an inducing mesoderm that was divided in two, just like Spemann's and Holtfreter's constricted embryos. The duplicated axes formed when this constriction was in the medial plane, and the Bauchstüch-like mass seen in several of the Kinkedembryos also "might well be the result of a frontal constriction."
Gluecksohn-Schoenheimer was aware of her integrating embryology and genetics. She announced that her research on the Kinked gene "was undertaken both from the point of view of the embryologist interested in the causal analysis of development and that of the geneticist interested in the analysis of gene effects" (24). Linking organizers to genes meant linking embryology to genetics.
During this investigation of axial development, other tailless or short-tailed mutant mice were found. One of these tailless mutants was due to the Sd/Sd genotype that also caused the lack of kidneys. Gluecksohn-Schoenheimer wrote that when confronted with such cases, the developmental geneticist must reverse the order of the experimental embryologist and work backward from effect to cause. Why weren't there any kidneys in these mice? She demonstrated that the ureteric bud normally grew into the area of the metanephrogenic mesenchyme. When that occurred, the ureter continued to grow and branch, and the mesenchyme condensed into tubules. In the Sd/Sd mutant, however, the ureteric bud failed to reach the mesenchyme and no kidney was formed. In Sd/+ heterozygotes, some tips of the short ureteric bud did find there way into the metanephrogenic mesenchyme, and a small kidney resulted. "These findings," wrote Gluecksohn-Schoenheimer (25), " indicate strongly the existence of an inductive relationship between the ureter and kidney—such as has been shown experimentally to exist in other vertebrates, the chick, for example."
In Gluecksohn-Schoenheimer's work during this period, there is a reciprocity between genetics and embryology. Genetics could be used to analyze development in areas where experimental techniques had not yet been perfected. Embryology could identify the effects of these genes whose functions were necessary for the construction of the embryo. These early embryonic abnormalities "represent the end result of a chain of events at the beginning of which stands the gene. The analysis of the action of the gene is our ultimate goal" (26).
This programatic statement of 1945 differs from that of 1938. In 1938, the developmental geneticist was content to draw conclusions on the nature of the "experiment" performed by the genes. Now, the further charge was to understand the nature of gene activity. But Gluecksohn-Schoenheimer's goal would have to await the techniques of molecular biology. She did not start analyzing gene activity until the mid-1970s when she turned the direction of her laboratory from morphological mutations in mice to the analysis of the biochemical defects caused by the deletion of a specific portion of mouse chromosome 7. In Salome Gluecksohn-Schoenheimer's research on developmentally lethal genes, we see the enormous role that experimental embryology, especially Spemann's constriction and transplant experiments, had in the propounding of developmental genetics.
At the same time, Gluecksohn-Schoenheimer's friend and colleague, C. H. Waddington, would also meet with frustration in his attempts to analyze the organizer. He, too, would turn to genetics as a way of approaching the problem of induction, and he would bring to the fruit fly the same procedure that Gluecksohn-Schoenheimer was employing to study mouse development. Waddington, however, was more theoretically inclined than Gluecksohn-Schoenheimer and was eager to reunify genetics and embryology not only with each other but with evolutionary biology, as well. While Gluecksohn-Waelsch's research narrowed in on specific regions of mouse chromosomes 7 and 17, Waddington's focus became increasingly wide. We will see that Waddington's studies served as a bridge linking the induction of organs in vertebrate embryos to the induction of enzymes in Escherichia coli.
3. THE CONCRESCENCE OF GENETICS AND DEVELOPMENT: C. H. WADDINGTON
3.1 The scientist as pluripotent migratory cell
Conrad Hal Waddington identified himself as a student of "diachronic biology," a science of "embryology-genetics-evolution which again form a group whose interconnections are obvious and unavoidable" (27). That Waddington did not see these three disciplines as distinct entities is reflected in his peripatetic training as a biologist. After graduating with a first class degree in geology from Cambridge University in 1926, he began pursuing graduate work in paleontology. His thesis work involved analysing ammonites, a group of extinct cephalopods that, he would later claim, "forces on one's attention the Whiteheadian point that the organisms undergoing the process of evolution are themselves processes...The whole developmental process is preserved so that one cannot avoid examining it." Waddington admitted to being very much influenced by Whitehead during his last two years as an undergraduate (28), and his research in paleontology was partially funded by a philosophy studentship. This work in paleontology did not lead to a PhD. He had met Gregory Bateson, and the friendship between these men caused Waddington's interest to move from paleontolgy towards genetics. However, Waddington felt that it was not possible to earn a living as a geneticist in Britain during the 1920s, so he looked elsewhere (29).
In 1929, Dame Honor Fell, director of the Strangeways Laboratory, was told about a "bright young paleontologist (who also had a scholarship in philosophy) who had been reading Spemann's papers and wondered if the organ culture method developed here could be used for the experimental study of avian and mammalian embryos" (30). Indeed, she was impressed with Waddington's ideas, and soon Waddington began working at the Strangeways Laboratory (31). By 1930, at the International Congress of Experimental Cytology in Amsterdam, Waddington was able to present his first results on culturing chick embryos. Another participant at this congress, Richard Goldschmidt, invited Waddington to come to Germany. However, when Waddington received funds to work in Germany, he decided to work with Otto Mangold "for the purpose of learning the technique of amphibian operations" (32), rather than to pursue genetics in Goldschmidt's laboratory. Mangold, a former graduate student of Spemann who was now his collaborator, was working on the problems of neural induction in amphibians. Waddington would adopt this set of problems for himself. From 1932 to 1938, Waddington attempted to clarify the nature of amphibian neural induction and to transfer the techniques of amphibian experimental embryology to the study of chick development.
3.2 The Chemistry of the Organizer: Sterols and Sterility
Waddington's work on chick embryos was marked by enormous success. Using his technique of growing avian embryos in vitro, he was able to manipulate and transplant different embryonic regions from one early chick embryo to another. He discovered that the elongation of the primitive streak was directed by the underlying hypoblast, and he referred to this process as an induction. He further showed that the formation of the chordamesoderm was directed by Hensen's node, and thus reasoned that this most anterior portion of the primitive streak was analogous to the dorsal lip of the amphibian blastopore. His photographs of twin chick embryos resulting from the transplanting of an exogenous node beneath the ectoderm gave testimony to his being able to demonstrate induction in a warm-blooded animal.
Meanwhile, research into the chemical nature of the amphibian organizer substance was becoming more and more frustrating. In 1933, Waddington and Joseph and Dorothy Needham showed that the activity of the newt organizer could be duplicated by ether extracts of adult newts. This ether extract could turn presumptive epidermis into neural tissue. Since it did not specify the regional properties of the neural tissue (i.e. brain or spinal chord), Waddington referred to it as the "evocator" (33). (The other molecules that would then specify the regional properties of the neural tube were called the"individuators"). The properties of this lipid fraction suggested that the evocator was a sterol, and, indeed, both natural and artifical steroid hormones were found to induce neural tubes.
This initially caused great excitement, and Waddington and Joseph Needham spent over three years attempting to biochemically characterize the active agent in the ether extracts. If the natural inducer substance turned out to be a sterol, Waddington and Needham would have linked embryology to two of the most exciting areas of contemporary physiology and biochemistry—steroid synthesis and reproductive endocrinology. Such a conclusion would have also been helpful in retaining an active sponsor for their research, as the Rockefeller Institute was keenly interested in sterol biochemistry and was already funding the technician for Waddington and Needham's research. A sterol inducer made good sense, and Needham outlined these reasons in his 1936 volume Order and Life (34). Sterols had been shown to provide the biochemical framework for (a) both the male and female sex hormones, (b) cancer-producing hydrocarbons, (c) vitamin D, and (d) pharmacologically active cardiac glucosides. Moreover, (e) Emil Witschi, one of the pioneering researchers on sexual development, had shown that sterols might be stored in eggs.
However, there was a problem. Some of the neural-inducing hormones were so unlike one another that there seemed to be no structural specificity. As Waddington and Needham end their discussion of one of their papers in 1935 (35):
Dodds has metaphorically spoken of these synthetic substances as skeleton keys, which can unlock several doors...Here the skeleton key is so unlike the householder's latchkey that one wonders whether the house has been entered through the back-door, or, in an even more unorthodox manner, through a window.
During the same year, Waddington also published two papers on the competence of ectoderm to respond to neural and lens inducers; but the pivotal paper showing the central importance of competence was not to appear until 1936. This paper by Waddington, Joseph Needham, and Jean Brachet (36) concerned the activation of the evocator substance. In this paper they retreated from the notion of a specific evocator substance released from the chordamesoderm which induces the ectoderm above it to become neural. Rather, they produce evidence that the evocator "substance, which is present throughout the whole embryo, was liberated or activated in one particular region, the organization center, by reason of a gradient system." The first evidence for this came from Holtfreter's 1933 experiments wherein he had demonstrated that non-inducing regions of the amphibian gastrula could acquire the ability to induce when the cells were killed. Thus, non-inducing tissue contained the active inducing factor, in "some form in which it could not exert its normal influence." Julian Huxley (1930) had speculated that the dorsal lip "organization center" was merely a region of extremely high metabolic activity. It was this quantitative difference, rather than any chemically qualitative ones, which distinguished it.
This speculation fit into the paradigm of C. M. Child's axial gradients which was very influential during the 1930s. Watanabe and Child (1933) had recently proposed that gradients of respiratory rates might have profound effects in the determination of embryonic cell fate. To this end, "An attempt was made to raise the respiratory rate of isolated fragments of ectoderm by the use of dyes [methylene blue and cresyl blue] which act as respiratory catalysts, and the treated ectoderm was then tested for inducing capacity." The answer was ambiguous. First, the tissues incubated in methylene blue dye gave obvious neural induction, but a sensitive manometer showed no difference in the respiration rate between the inducing tissues and the non-inducing ones. The researchers dismissed as "ridiculous" the notion that methylene blue resembled the natural evocator. Rather, they interpreted their findings as showing that the evocator was present in the competent tissue in an inactive state. A substance could cause neural induction either by releasing the natural evocator from its inhibitor or by resembling the evocator itself. This allowed them to draw a distinction between the chemicals they thought resembled the evocator (such as sterols) and those chemicals which non-specifically release the natural sterol inducer from its inhibitor (such as methylene blue) (37).
By 1938, the chemical identification of the evocator substance was still stalled. Waddington showed that estrone, a natural estrogen, seemed to be among the most potent inducers of neural tissue in amphibians, but the biochemists had not come any closer to finding the natural inducing molecule. As Yoxen (38) has shown, the project to characterize the organizer molecule had by this time become untenable because of the non-specificity of inducers. Moreover, the departmental politics at Cambridge University stopped the Rockefeller Foundation from continuing its support of the project (39). That same year, using Rockefeller Foundation support, Waddington travelled to the United States. Here he worked with L. C. Dunn at Columbia University and renewed his acquaintance with Salome Gluecksohn-Schoenheimer. He made trips from New York City to visit Ross Harrison at Yale, Curt Stern in Rochester, and Streeter and Metz at Johns Hopkins. In January, 1939, Waddington began a very productive three-month stay at the California Institute of Technology. In this extremely active genetics group (including Columbia University transplants Sturtevant and Dobzhansky), Waddington started applying to Drosophila the same type of developmental genetics that Gluecksohn-Schoenheimer had recently pioneered on mice. He analyzed 24 alleles known to cause deformation of the wing, and he showed their earliest visible deviations from wild type while they were in the pupa. The wing, he said (40), "appears favorable for investigations on the developmental action of genes."
3.3 Genes as Organisers
Not everybody believed that the "developmental action of genes" was important or that it even existed. In 1924, Spemann alerted his fellow-embryologists to the threat posed by geneticists who felt that the genes explained all of development. "They have cast their eye on us, on Entwicklungsmechanik," he wrote (16). In 1937, Ross Harrison gave a lecture at the zoological sciences section of the American Association for the Advancement of Science warning the geneticists to stay on their own turf. He warned of the dire consequences of "this threatened invasion" caused by the "Wanderlust" of geneticists. Embryologists such as Spemann, Harrison, and Lillie ignored genetics, saying that development could not be controlled by genes that were the same in every cell type; while geneticists such as Goldschmidt laid claim to development, certain that the ontogeny of the embryo could be viewed as an epiphenomenon of gene expression (4).
Aside from Waddington, Gluecksohn-Schoenheimer, and Dunn, only a few others were actually looking at the "developmental action of genes" at this time. Goldschmidt certainly was, but his version of genetics, summarized inPhysiological Genetics (1938), was not the genetics of the Morgan school. Boris Ephrussi was another researcher actively studying the developmental action of genes. As will be detailed by Burian, Gayon, and Zallen (this volume), Ephrussi (41) undertook experiments on Drosophila eye pigments "to lay a bridge between causal embryology and genetics." Ephrussi's techniques were those of the embryologist (transplantation) rather than those of the geneticist (breeding), and he explicitely looked at these phenomena in terms of development, using the term "inducer" to refer to the substance made by the wild type cinnabargene.
We have no lack of information concerning Waddington's views on this subject. Between 1938 and 1940, Waddington wrote two textbooks and two review articles concerning the developmental action of the genes. The first of these was in his textbook An Introduction to Modern Genetics. (This was quite a presumptive undertaking for a 33 year-old paleontologically trained embryologist who had yet to publish his first paper in genetics!) The second review was written for The School Science Review on "Embryonic Organizers." Another article, bearing the provocative title "Genes as Evocators in Development" was delivered at the first Growth Symposium in 1939 and published in the supplementary edition of Growth. Finally, Waddington's synthesis of genetics and embryology is given full treatment in Organisers and Genes (1940).
One of the major themes in these reviews is the formal equivalence of organisers and genes. Since Waddington viewed development as a succession of distinct alternative conditions, "it can best be symbolized as a system of branching paths. The characteristics of each path will depend upon the developmental potencies of the tissue, that is to say, they will be under the control of the genes. We may also expect to find genes which act in a way formally like that of evocators, in that they control the choice of alternatives. Genes of this sort are in fact known. One good example is aristopedia in Drosophila melanogaster..." (42). Thus, Waddington developed lists of analogies. The gastrula ectoderm had the alternatives of becoming either epidermis or nerve tissue. The evocator acted to make it nerve, and without this inducer, it would become skin. In like manner, he argued, the anterior imaginal tissue of theDrosophila larva could become either leg or antenna. The wild type aristopediagene acted to make this tissue develop into antenna. In its absence (as when this gene mutates), the same tissue would develop into leg structures. Thus, the gene was acting as an organiser. Similarly, the genes that controled the sexual phenotype of animal gonads and the color of Drosophila eyes also functioned as organisers. The male or female phenotype was determined from orginally bipotential gonads. The choice of one of the two alternative paths was seen to be directed either by internal genes or by the external environment (where he mentions the case of Bonellia) (43). Ephrussi's studies on eye pigmentation were also interpreted in this manner. The eyes could become red or cinnabar colored. In the presence of the wild-type gene, the eyes become red. If this gene is not present, the alternative phenotype is chosen (44).
The ectoderm of the amphibian gastrula has two alternative methods of change open to it; it may become epidermis or if the evocator is added to it, it may become neural tissue. The case is exactly parallel to that of the pigment system of Drosophila at one of its branch points.
Thus, Waddington was able to speak (45) of "evocator genes which shunt development along one or the other of the possible developmental branches." This is illustrated in figure 2.
Figure 2 Branch-track diagram showing the fate of antennal imaginal tissue inDrosophila. The wild type aristopedia (ssa) gene would cause the cell fate to become antenna. However, the mutant allele at this locus would cause the cell fate to travel down the other path to become leg. Other mutations could act to modify these new conditions. From Waddington (27) For a more detailed discussion of Waddington's bifurcation diagrams, see Gilbert, 66).
To Waddington, the embryologist and the geneticist were studying the same phenomenon (46).
The similarity between the theoretical schemes we have arrived at on embryological and genetic grounds is immediately apparent. In embryonic development we are confronted with alternative modes of development, the choice between which is taken in reference to an external stimulus, in inductive development or to an internal one, in mosaic development. In considering the effects of genes, we find alternatives the choice between which may be taken in response to diffusible substances, as in the Drosophila eye colours, or apparently in response to internal factors as in aristopedia. It is clear that we have merely followed two different methods of approach to the same phenomena, and that the two schemes are in fact identical.
Since "both methods of approach to the study of development formulate the main problems in the same kind of way," Waddington hoped (47) that "genetics and embryology can collaborate in finding the answers." Indeed, Waddington begins Organisers and Genes by declaring a truce between the geneticists and the embryologists. "A coherent theory of development cannot be founded on the known properties of genes; in fact, it seems much more hopeful to try to fit our somewhat scanty knowledge of the developmental action of genes into a framework founded in the first instance on the direct experimental study of development "(48). Both the geneticist and the embryologist had valid research programs and a great deal more work to do in order to explain development.
3.4 The primacy of competence and the foundations of genetic assimilation
Spemann's 1938 Silliman Lecture series Embryonic Development and Induction, although far more complete than Waddington's Organisers and Genes, spent hardly any print on the concept of the competence of the tissues to respond to induction. Waddington, however, devotes more space to this concept than to induction, itself. There are several reasons for this. One is that Waddington wrote, not of induction, but of the "evocator-competence system." Second, his own experiments had convinced him that the evocator was not specific. Rather, specificity resided in the competent cells. Many things could induce, but only certain tissues could respond to the inducer. This competence to respond to an evocator in a particular way was linked directly to gene action (49).
The evocator is merely a differential; it is the competence which is responsible for the details of the developmental process and thus of the kinds of tissues produced. Since it is the genes which control the character of the animal and its tissues, it must in general be genes which determine the properties of the competence.
Waddington's idea of "competence" differed from the analogous German term Reaktionsfähigkeit, which implies a passive ability to respond to the stimulus given it. For Waddington (50), competence is actively achieved by "a complex of reactions between substances which form an unstable mixture, which may at certain times have two or more alternative modes of change." Thus, competence is manifest in an unstable (i.e. multipotential) system which an inducer can push to one equilibrium or another.
This view, that genes control competence, and that inducers merely push an unstable system into one of two alternative equilibria leads to the notion that competence, rather than induction, per se, is the central aspect of determination. This view had several important consequences. First, it explained the notions of potency and segregation, so dear to the American embryologists; for once a tissue acquired its competence, the stimulus that induces it could be changed (51).
The transition from typical inductive development to cases of double assurance show that the processes occuring in mosaic development, where no inducer is involved, are of the same general nature as those involved in competence; only in the former the factor that decides which mode of development will be followed by a piece of tissue is already present within it, instead of having to diffuse in from the surroundings.
Since the competence could be achieved independently from an evocator, and since different evocators could induce the same determinative process, a given competent tissue could transfer its ability to respond from one inducer stimulus to another. These evocators could be intracellular (as in mosaic development), intercellular (as in inductive development), or even environmental (as in the case of Bonellia sex determination). This will form the basis of Waddington's concept of genetic assimilation. Here, a tissue that is competent to respond to environmental evocators becomes competent to respond to internal evocators as well. In other words, an environmentally induced response can become an embryologically induced response.
Waddington used as an example the formation of calluses on the underside of the ostrich. Ostriches are born with the calluses already in place, and thus appear to be a case of Lamarckian evolution wherein the adult skin would form calluses upon abrasion and the offspring would inherit such properly positioned calluses. Waddington was able to explain this by invoking a transfer of competence from an external inducer to an internal inducer. First, the belly skin of the ostrich has to be able to develop calluses in response to friction. This is a genetic competence that this region of skin has acquired through natural selection. Then, during evolution, the skin—which has already obtained the competence to form calluses—becomes able to respond to another, internal, inducer. The skin is thereby able to form calluses through embryonic induction. What had appeared to be a case of Lamarckian "inheritance of acquired characteristics through use" can be explained by development and natural selection (52).
Presumably its skin, like that of most other animals, would react directly to external pressure and rubbing by becoming thicker...This capacity to react must itself be dependent upon genes...It may then not be too difficult for a gene mutation to occur which will modify some other nearby area of the embryo in such a way that it takes over the function of external pressure, interacting with the skin so as to "pull the trigger" and set off the development of the callouses.
This genetic assimilation hypothesis, based on the ability of competent tissue to switch evocators, operated according to strictly Darwinian natural selection, and could also, wrote Waddington, "provide a plausable account of the result in terms of orthodox genetic and embryological mechanisms." (53).
As biology became increasingly molecular, Waddington would use the concept of competence and masked evocators to account for that most "Lamarckian" of molecular phenomena then known; adaptive enzymes in yeast and E. coli (54). Here, an "inducer" molecule would influence gene activity by interacting with cytoplasmic components that could activate or suppress specific genes in the cell. While Waddington did not have anything to do with the discovery of the operon, he followed this work closely and brought it into discussions of developmental biology. He clearly saw the bacterial operons as models for the elements of embryonic induction. But in order to appreciate how bacterial operons might be used to model embryonic blastomeres, he had to possess two general concepts that were not generally held at this time. First was a view of organisms and cells as "systems" of interacting parts. While Waddington would eventually expand this notion into a full-fledged cybernetics of development, his earlier views of systems come from Whitehead's philosophy. Second, he had to believe that the cytoplasm could activate the genome. This went against the grain of genetics which held the nucleus to control the cytoplasm (see 4,8). Yet this would be crucial in his analysis of induction and provides the link between the embryological induction of organs and the microbial induction of enzymes.
3.5 Concrescence, Chreods, and Canalization
Waddington was a member of the Cambridge-based Biotheoretical Gathering that was trying to provide a molecular basis for embryology. This group was self-consciously grounded in dialectical materialism, organicism, and a belief that the process philosophy of Alfred North Whitehead was important for studying emergent processes such as evolution and development (55,56,57; see Abir-Am,57a). To be sure, different members of this group placed different import on each of these characteristics, and it is also probable that each of them interpreted dialectical materialism differently and saw different parts of Whitehead's philosophy as being important. Joseph Needham, for example, placed Whitehead's work in the context of dialectical materialism, the latter being (58) "a theory of transformations of the way in which the qualitatively new arises, of the nature of change in the world." But to Waddington, Whitehead ("to whose writings I paid much more attention during the last two years of my undergraduate career than I did to textbooks in the subjects on which I was going to take my exams") had superseded dialectical materialism with a fuller view of nature (28, 59).
But what did Waddington take from Whitehead's philosophy? Undoubtedly, the notion that there were no "things" except in their interelationships was extremely important to Waddington (60), and he notes this in his autobiographical sketch (61).
As far as scientific practice is concerned, the lessons to be learned from Whitehead were not so much derived from his discussion of experiences, but rather from his replacement of "things" by processes which have an individual character which depends upon the "concrescence" into a unity of very many relations with other processes."
Indeed, Waddington stressed the role that concrescence had played in his biological thought. "In the late thirties I began developing the notion that the process of becoming (say) a nerve cell should be regarded as the result of a large number of genes, which interact together to form a unified 'concrescence'." He also claimed that this view motivated his initial work inDrosophila. Waddington retrospectively linked his appreciation of concrescence to his work on competence. Moreover, he appreciated the formal similarity of his masked evocator hypothesis to that of Jacob and Monod's operon (62).
Again a few years earlier it came to be apparent that the "gene concrescence" itself undergoes a process of change; at one embryonic period a given concrescence is in a phase of "competence" and may be switched to one or another of a small number of alternative pathways of further change—but the competence later disappears...
We showed that, in these terms, the specificity resides in the cells that react to induction—we called it "the masked evocator." This is very similar to the situation discovered by Jacob and Monod many years later in bacteria, where again the specific repressor molecules are internal to the cells which react to enzyme-inducing substances.
Waddington claimed that these were not merely retrospective glosses placed on his earlier work. Rather, "I tried to put the Whiteheadian outlook to actual use in particular experimental situations." Waddington wrote that he used the neologism chreod (or creode) as a substitute for the Whiteheadian term "concrescence." Before using "chreod" he had no shorthand way for saying "a stabilized or buffered pathway of change". This concept was also depicted by a visual image that Waddington called "The Epigenetic Landscape" and is shown in Figure 3.
Figure 3 Representation of the epigenetic landscape. The ball represents cell fate. The valleys are the different fates the cell might roll into. At the beginning of its journey, development is plastic, and a cell can become many fates. However, as development procedes, certain decisions cannot be reversed. From Waddington (77).
The epigenetic landscape was a way of depicting the branching patterns of development and at the same time portraying the different stabilities of these pathways by depth and contours. This conceit is first seen in the Introduction to Modern Genetics (63) as a representation of development "not as a branching line on a plane but by branching valleys on a surface." The depth and contours of the "geological model represent probability, so that the valley bottom is really a representative of an equilibrium." The molding of these contours would be accomplished by various genes, "and genes like [the Drosophila eye pigment gene] vermillion which have their effects at certain branching points are like intrusive masses that can direct the course of the developmental process down a side valley."
Waddington likened the establishment of cell fate to a ball rolling down the valleys of the epigenetic landscape. At certain times (as in competent tissue), two downhill paths are possible, and the presence of an evocator could deflect the tissue into one or the other of these paths. Regulation took place only when the path of development took the embryo into a valley with "gently sloping sides". However, as development proceeded, the originally wide and gently sloping valley branches into subdivisions having far steeper sides. Here, regulation cannot take place.
Implicit in this model is the notion of canalization. (Indeed, Waddington quotesOrganisers and Genes when he mentions canalization in his 1956 book, Principles of Embryology, even though canalisation is not explicitely mentioned in the earlier work). Canalisation is made explicit shortly afterwards, though, in 1941, when Waddington stated (64) that "developmental reactions...are in general canalized. That is to say, they are adjusted so as to bring about one end result regardless of minor variations in conditions during the course of the reaction." Canalisation can be considered a buffering of the developmental pathways such that minor mutations would not greatly affect the course of development. Canalisation would limit variations in development such that "If wild animals of almost any species are collected, they will usually be found to be 'as like as peas in a pod'." The canalised paths were themselves thought to be selected by natural selection. Waddington related this to genetic assimilation, stating that once a developmental path had been canalysed (as in the ostrich's ability to form a callus by friction), mutations would be able to switch development into that path.
Once within a canal, though, it is difficult to get out. Thus, canalisation is not unlike the current notion of "developmental constraints." Indeed, it was Waddington who suggested (65) in 1938 that all vertebrates were constrained to have a notochord since that transitory organ induced the neural plate, and who similarly claimed that birds (and presumably humans) had to have non-functional pronephric kidneys since they would give rise to the Wolffian ducts.
Waddington did not tell his readers from whence he derived the term and concept of canalisation. Looking at the pictures of the epigenetic landscape (especially John Piper's rendering of it on the frontispiece of Organisers and Genes), one would think that it followed from this visual image (66). However, it. too, probably came from Whitehead's Process and Reality, where Whitehead discusses "canalisation" as well as concrescence. In fact, Whitehead's use of canalisation would be difficult for an embryologist to miss, for the term is used in discussing the development of the animal body and the emergence of ordered mentality. "Apart from canalisation," wrote Whitehead, "depth of originality would spell disaster for the animal body", for life is a passage from physical order to random mental processes to "canalised mental originality". For Whitehead, canalisation provided both the limits and the magnification of the creative urge, allowing things begun to come to completion (67). Thus we see that Waddington's idiosyncratic approach to development—concrescence, canalisation, and genetic assimilation—arose from his placing fundamental emphasis on competence (rather than on induction) and in his placing these observations in the context of a Whiteheadian philosophy of organismal change. In the next section, I attempt to show that his analysis of competence was, itself, a product of his Whiteheadian philosophy.
3.6 SYSTEMS IN DIALOGUE
Whitehead was a philospher of systems in the process of becoming. Indeed, the first three elements of his philosophy were the concepts of system, process, and the creative advance into novelty (68). For Whitehead, no thing existed except in relation to other things, and these nexus were always changing, allowing new nexus to form. All relationships were in the process of becoming, and all things were linked within systems. These were very useful concepts for biologists who were engaged in studying process, interrelationships, and the emergence of new forms. Many biologists in the Biotheoretical Gathering were deeply influenced by these notions (69).
This certainly seems to be the case in Organisers and Genes, which reads like a Whiteheadian primer on embryology. Throughout this book, Waddington stressed "interrelationships," "causal networks'" and "interconnections." (All these terms can be found on the first page!) He sought (70) to identify—not the inducer—but the "causal network underlying this particular process of differentiation," and hoped "to know the whole complex system of actions and interactions which constitute the differentiation."
This tendency to think in terms of process, system, and interaction also distinguished his approach to induction from those of most other investigators (71). In 1940 he was not talking so much about the inducer as he was about "The evocator-competence reaction." Similarly, he was not so much concerned with the action of genes as with "the system of tracks and their genetic control." For Waddington, the parts of the embryo were always in dialogue. The evocator was nothing without the competent responding tissue, and the responding tissue was nothing without the evocator. They were linked in a system. Moreover, Waddington interpreted the results of embryology to show that these components interacted to effect development. Waddington looked upon an organism as a developing system, and claimed that natural selection worked upon aspects of development. Evolution was accomplished through hereditable changes in an organism's development. He credited this idea to his Whiteheadian outlook, and called his first paper on evolutionary topics (1941) "the evolution of developing systems."
The relationship between inducer and competent tissue paralleled that of the genes and the cytoplasm. The genes and the cytoplasm were in continual dialogue, and in Whiteheadian fashion Waddington claimed (72), "Neither cytoplasm nor nucleus can be disregarded: In fact the most important subject to discuss is how they affect each other." Moreover, just as Waddington saw the competent tissue as having primacy over the inducer, so he appears to give the primacy of the "interacting system of nucleus and cytoplasm" to the cytoplasm. This empowerment of cytoplasm marks a major difference between him and most geneticists, just as his empowerment of competent tissue separated him from most embryologists. In both cases, he "championed" the view of the "passive" partner. The inducer was thought to call forth a response from the competent induced tissue, and genes were believed to control the cytoplasm. Genes may be equivalent to organisers, but in both cases, what mattered was the responsive partner. The reasoning, too, may be similar. In induction, the competent tissue can be induced by several inducers. The specificity is in the tissue that is competent to be induced. Similarly, within the cell, all the genes are the same. The specificity resides within the cytoplasm.
In his 1956 book, Principles of Embryology, Waddington provided a provocative title to his chapter on developmental genetics: The Activation of Genes by the Cytoplasm. The normal "genetic" roles have been reversed! What was Waddington's evidence for this reversal? Within this chapter, he presented four modes in which the cytoplasm had been seen to control which genes were expressed by the which cell. First, in mosaic eggs, the cytoplasm of the cell obviously controlled the genes expressed by its nucleus. Second, in induction, a diffusible substance is thought to affect "the interacting system of nucleus and cytoplasm" in the competent cell. Third, in Drosophila, the chromosome puffs, believed to represent active genes, were different depending on the cell that they were in. Fourth, Sonneborne's studies on Paramecium had demonstrated that "it is the condition of the cytoplasm which determines which of the loci shall be in operation." This demonstration of the cytoplasmic control of G-antigen synthesis was particularly important, since Waddington called it "a clean-cut example of the activation, by different types of cytoplasm, of different specifically corresponding genes." (73).
Waddington saw the Paramecium experiments as being important to embryology, even though Paramecium was a unicellular protist (74). The fact that cytoplasmic control of nuclear genes "occurs, not in different parts of the same body but in various members of unicellular organisms does not make the phenomenon any the less relevant to the normal processes of development." Thus, in 1956, Waddington was already using microorganisms as models for the cellular differentiation of multicellular organisms, and he states explicitly (75) that in order to study differentiation, one has to consider "the genetics of microorganisms as well as higher forms."
Waddington's 1956 model of cell differentiation started from the premise of a system wherein the cytoplasm controls differentiation by either interacting with the genes directly or with the immediate product of the genes (76). The question then became how the constitution of the cytoplasm changed such that one canalised system would be followed rather than another. He built a model wherein the genes compete for raw materials within the cell. He likens this to animals competing for limited food reserves, and points out that slight changes in the initial conditions can effect a great difference in the final state. The evidence for this model comes from microorganisms, specifically from Spiegelman's studies on adaptive enzyme production in yeast. If the yeast were grown on two substrates for which it originally lacked the appropriate degradative enzymes, they would gradually synthesize those enzymes. However, if those yeast cells were starved for nitrogen, the two proteins enter into competition for the small nitrogen reserve. Moreover, when one of them is synthesized, its synthesis appears to be autocatalytic.
Thus, Waddington can bring his reader back to the original problem of induction and competence. In evocation, the environmental influence (the evocator) acts on the cytoplasm of the competent cell, "causing it to adopt one or the other of the alternative paths of development open to it. The fact that the reacting tissue retains its own specific characteristics...shows that the developmental paths are under genetic control and that evocation involves the differential activation of a particular set of genes." (77). The problem of induction therefore centers around the mechanism by which crucial changes in the responding cells' cytoplasm allow one reaction to outcompete its rivals.
From the Literature Cited in Principles of Embryology, we know that Waddington was already very much aware of Monod's work on adaptive enzymes in E. coli. When Jacob and Monod published their studies on the operon model in 1961, they had a ready audience in Waddington. Waddington's New Patterns of Genetics and Development was written that same year, and he began his book by showing the relevance of Jacob and Monod's "repressor-operator system" to embryology. New Patterns of Genetics and Development was Waddington's revision of Organisers and Genes on its twenty-first birthday. Like the earlier book, it attempted to integrate genetics and embryology. But genetics had become molecular genetics, while embryology had not made many significant conceptual advances in those twenty-one years. Waddington conscientiously attempted to make the operon his basic paradigm of differentiation (78), and he explicitly linked the "induction" of bacterial enzymes with the "induction" of the neural tube (Figure 4). Within a year of its being publicized, the operon model was being used as a paradigmatic model for cell differentiation.
Figure 4 Eukaryotic embryonic induction modeled on the lac operon of Jacob and Monod. Evocator substance from one cell (below) could be freed to diffuse into neighboring cell and either directly activate a series of genes or interfere with a repressor of the transcription of those genes. From Waddington (78).
In the work of C. H. Waddington from 1938 to 1962, we see the question of differentiation taken from the embryonic induction of vertebrate organs to the molecular induction of microbial enzymes. The steps can be summarized as follows:
Waddington's Whiteheadian outlook caused him to see dialectical interactions, rather than vectoral influences, between the nucleus and cytoplasm and between the inducer and the responding competent tissue. The lack of specificity in the inducer and the lack of gene specificity in the nucleus caused him to emphasize the roles of the cytoplasm in the nucleocytoplasmic system and the roles of the competent tissue in the inductive system. The emphasis on competence led to the concept of genetic assimilation wherein the inducer is changed from an external one to an internal one. Both the old and new inducers were thought of as small molecules coming from outside the cell and which were able to switch the genes of a competent cell from one canalised pathway to another. This led to Waddington's being ready to use adaptive enzymes of microorganisms as instructive analogues of eukaryotic cellular differentiation.
The pathway forged by Waddington could not have been made by a person who was purely a geneticist or purely an embryologist. Waddington's goal of synthesizing genetics, embryology, and evolution was critical for his ability to connect the disparate studies, and he was concerned that the success of molecular biology might overshadow embryological studies. He ends New Patterns in Genetics and Development:
I should like to see the present fashion for molecular genetics diluted by the diversion of rather more attention to fundamental embryology. Genetics has had its breakthrough, and those who want quick results can probably get them most easily by exploiting this. But the next breakthrough we need, to round off our understanding of fundamental biological processes, is an embryological breakthrough. Let us hope that we get it soon.
Acknowledgements
I would like to thank the following people for their suggestions on earlier versions of this manuscript: Pnina Abir-Am, Richard Burian, Anne Fausto-Sterling, Brian Goodwin, Donna Haraway, Larry Holmes, Stuart Kauffman, Jan Sapp, and Fred Tauber. I would also like to thank Dr. Salome Gluecksohn-Waelsch for consenting to be interviewed and Dr. Michael Somers for the use of his books.
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29. Yoxen, E. 1986. Form and strategy in biology: Reflections on the career of C. H. Waddington. In: A History of Embryology op. cit., pp. 309-329.
30. Ibid., p. 313.
31. If Waddington might be compared to a pluripotential stem cell migrating through various morphogenetic fields, then it is of interest that three of his major evocators were women. Waddington traced his interest in evolution to Miss G. L. Elles, his interest in genetics to Miss E. R. Saunders, and his training in embryology to Dame Honor Fell. He was called to the attention of Dame Fell by another woman, Miss Sidney Cox. (Robertson, A.,1977, Conrad Hal Waddington: 8 November 1905- 26 September 1975. Biograph. Mem. F.R.S. 23: 575-622.)
32. Waddington, C. H., Needham, J., and Brachet, J. 1936. Studies on the nature of the amphibian organization centre III. The activation of the evocator.Proc. Roy. Soc. (Lond.) B: 120: 173-198.
33. Waddington, C. H., Needham, J., Nowinski, W. W., and Lemberg, R., 1935, Studies on the nature of the amphibian organization centre I. Chemical properties of the evocator. Proc. Roy. Soc. (Lond.) B 117: 289-310.
34. Needham, J. 1936. Order and Life. Yale University Press, New Haven.
35. Waddington, C. H. and Needham, D. M., 1935, Studies on the nature of the amphibian organization centre II. Induction by synthetic polycyclic hydrocarbons. Proc Roy. Soc. (Lond.) B 117: 310-317.
36. Yoxen, E., 1986, op. cit.
37. A similar hypothesis has recently been proposed for the release of an internal regulatory protein that is essential for B cell differentiation. Here, the regulatory protein NF-kB is found in every cell; but it is usually complexed with its inhibitor, IkB.The inhibitory complex can be dissociated by several externally applied reagents, but occurs naturally only during lymphocyte development. (Baeuerle, P. A., and Baltimore, D., 1988, Science 242: 542-545).
38. Yoxen, E., 1986, op. cit.
39. Abir-Am, P., 1988, The assessment of interdisciplinary research in the1930s: The Rockefeller Foundation and physico-chemical morphology, Minerva 26: 153-176.
40. Waddington, C. H., 1939, Preliminary notes on the development of wings in normal and mutant strains of Drosophila. Proc Nat. Acad. Sci. USA 25: 299-307.
41. Beadle, G. W. and Ephrussi, B., 1935, Differenciation de la couleur de l'oeil cinnabar chez la Drosophile (Drosophila melanogaster), Comp. Rendes Acad. Sci. Paris 201: 620-622.
42. Waddington, C. H., 1939, Genes as evocators in development, Growth, suppl. 1: 37-44.
43. Waddington, C. H., 1939, An Introduction to Modern Genetics, Macmillan, New York.
44. Ibid., p. 184.
45. Waddington, C. H., 1940, Organisers and Genes, op. cit., p. 88.
46. Ibid., p. 92.
47. Ibid., p. 184.
48. Ibid., p. 3. This insistance upon placing the mechanism of gene action in the context of embryonic development continued to be a theme in Waddington's work. In Principles of Embryology, Macmillan, New York (l956), he would claim that "whatever the immediate operations of the genes turn out to be, they most certainly belong to the catagory of developmental processes and thus belong to the province of embryology." The problem of gene activity "is essentially an embryological problem."
49. Ibid., p. 54.
50. Ibid., p. 55.
51. Ibid., p. 50.
52. Waddington, C. H., 1941, Canalization of development and the inheritance of acquired characteristics, Nature 150: 563-565.
53. That was not, however, how Waddington's "genetic assimilation" was usually interpreted. While Lerner (The Genetic Basis of Selection, 1958) and Merrell (Evolution and Genetics, N.Y., 1962) saw genetic assimilation as a way of explaining so-called Lamarckian inheritance in terms of orthodox Darwinism, two of the major interpreters of evolutionary theory, Theodosius Dobzhansky and Ernst Mayr, interpreted Waddington's theory as being a failed attempt to support Lamarckian inheritance. Both Mayr (Animal Species and Evolution, 1963) and Dobzhansky (Genetics of the Evolutionary Process, 1970) claimed that there was no genetic assimilation, and what Waddington saw in his experiments was merely the selection of pre-existing variants in the population. Waddington's term, "genetic assimilation" was poorly chosen in that it did convey a notion that physiological responses could be readily fixed in the genome. Given that this type of assimilation was the basis for the neo-Lamarckian research program of Trofim Lysenko and that many British Marxists were supporting Lysenko, it is not hard to understand how Waddington's views could be thought of as subscribing to a goal-directed inheritance of acquired characteristics. Gilbert (manuscript submitted) attempts to show how "transfer of competence" (the basis for Waddington's genetic assimilation) can explain several problems in developmental biology.
54. The neo-Lamarckian character of French studies on "adaptive' enzymes has been noted by Sapp, reference 8, op. cit., p. 135.
55. Abir-Am, P.,1987, The Biotheoretical Gathering, transdisciplinary authority, and the incipient legitamation of molecular biology in the 1930s: New historical perspective on the historical sociology of science, J. Hist. Sci. 25: 1-71.
56. Haraway, D., 1976, Crystals, Fabrics, and Fields: Metaphors of Organicism in Twentieth Century Developmental Biology, Yale University Press, New Haven.
57. Wersky, G., 1978, The Visible College: The Collective Biography of British Scientific Socialists of the 1930s, Holt, Rinehart, and Winston, New York.
57A. Abir-Am, P. 1991. The philosophical background of Joseph Needham's work in chemical embryology. In A Conceptual History of Modern Embryology. (S. F. Gilbert, ed.) Plenum Press, NY. Pp. 159-179.
58. Needham, J., 1943, A biologist's view of Whitehead's philosophy (1941), in: Time: The Refreshing River, Macmillan, New York, p. 188.
59. Waddington, C. H. , unpublished lecture notes: Marxism and Biology, University of Edinburgh archives. Courtesy of Dr. P. Abir-Am.
60. Waddington, C. H., 1970, Behind Appearances, MIT Press, Cambridge, p. 114.
61. Waddington, C. H., 1975, autobiographical notes, op. cit., p. 3.
62. Ibid., p. 10.
63. Waddington, C. H., 1940, Modern Genetics, op. cit., p.187.
64. Waddington, C. H., 1941, Canalization, op. cit.
65. Waddington, C. H., 1938, The morphogenetic function of a vestigial organ in the chick, J. Exp. Biol. 15: 371-384.
66. Stream images were not uncommon in developmental biology at that time. They were used by Lillie, Just, Goldschmidt, and Childs, among others.(Gilbert, S. F., Epigenetic landscape design, manuscript submitted).
67. Whitehead, A. N., 1929, Process and Reality, Cambridge University Press, Cambridge, pp. 127, 151.
68. Ibid., p. 151.
69. Waddington, C. H., 1975, autobiographical notes, op. cit., p. 5.
70. Waddington, C. H., 1940, Organisers and Genes, op. cit., p. 3-4.
71. Ross Granville Harrison may have had a similar approach to induction-competence. In his 1933 review, "Some difficulties with the determination problem" (American Naturalist 67: 306-321) Harrison wrote that no one factor determines a tissue to the exclusion of other factors. Waddington paraphrased this when he wrote (Organisers and Genes, p. 4) that "No 'stimulus', no simple cause, is itself an adequate explanation of anything."
72. Waddington, C. H., 1956, Principles of Embryology, Macmillan, New York, p. 348.
73. Ibid., p. 360.
74. There is some historical irony here. In 1896, E.B. Wilson used protist models of differentiation to show that the nucleus dominated the cytoplasm. (For details, see 8, and Sapp, this volume).
75. Waddington, C. H., 1956, Principles of Embryology, op. cit., p. 350.
76. Waddington postulated intermediates between the genes of the nucleus and the cytoplasmic proteins they produce. These intermediates were necessitated by the findings that cell cytoplasm could still synthesize new proteins even after their nuclei had been centrifuged away. He speculated that these intermediates might be plasmagenes (i.e., copies of genes residing within the cytoplasm) and even mentioned the possibility of their being RNA (Principles of Embryology, pp. 355, 406).
77. Waddington, C. H., 1956, Principles of Embryology, op. cit., p. 412.
78. Waddington, C. H., 1962, New Patterns in Genetics and Development, Columbia University Press, New York.