1999 E.B. Wilson lecture: the cell as molecular machine.

In 1955, I began graduate studies at the University of Chicago in the Committee on Biophysics. The program allowed considerable freedom in choosing a research project. When I had made up my mind, I went to see Dr. William Bloom, the distinguished histologist, and told him that I wanted to work on the mechanism of mitosis. He handed me his copy of Wilson’s (1925) book, The Cell in Development and Heredity, and said, “Read it, and when you finish, if you still want to work on such a difficult problem, come back and see me, and we’ll talk. ” On his retirement, Dr. Bloom gave me this copy of Wilson (1925) and I have been rereading parts of it while preparing this lecture. My title was intended to make the topic appear fashionable but the idea of the cell as a machine is not new. Loeb in 1906 referred to the cell as a “chemical machine” and Wilson tells us that “the specificity of each kind of cell depends essentially on what we call its organization, i.e., upon the construction of the cell machine” but “ the differences between the cell and even the most intricate artificial machine still remain too vast by far to be bridged by our present knowledge. ” At that time, the research group of William Bloom, Raymond Zirkle, and Robert Uretz was investigating the mechanism of mitosis by using microbeam irradiation by protons and UV light. The newt was used as the source of material because the cells are big and beautiful. They do not round up during mitosis and one can see the stages of the process in the living cell with a clarity that is without parallel. Mitosis is the prime example of the operation of the cell as a molecular machine. The explanation of the formation of the mitotic apparatus and the movements of the chromosomes had made little progress since 1925, the year of the third edition of Wilson’s (1925) book. Franz Schrader’s (1953) critical discussion of the current state of the field was a particularly valuable introduction to the problem. The contrast between the mode of thought at that time and the molecular biology of today is very striking. A description in terms of the controlled assembly and disassembly of fibers consisting of globular protein subunits and the action of local force generators, which move organelles along tracks, would not have been possible in 1955. In this lecture I wish to focus more on the development of these ideas than on an account of the work in my laboratory. My Ph.D. dissertation was a very modest contribution to solving the mechanism of mitosis. There was a shortage of actual numbers to test models of mitosis and my work was primarily concerned with measuring the rates of the processes by using polarized light microscopy to visualize the spindle. The rate of growth of the spindle in prophase and the elongation in late anaphase and the rate of chromosometo-pole motion for various conditions provide data for thinking quantitatively about the mechanism. A constant growth rate of the spindle of 1 to 2 mm/min could be explained by assembly of filaments from a pool of subunits. Later studies on microtubule polymerization did yield velocities in this range. The amount of energy expended against viscous drag on the chromosomes was calculated from the velocity of chromosome movement and an estimate of cell viscosity obtained from the Brownian movement of vesicles. The amount of energy is very small, equal to the hydrolysis of 25 to 50 molecules of ATP for a movement of 20 mm. The velocity did not depend on the size of the chromosome, as had been noted previously, and it also did not depend on the viscosity of the cell. I concluded that mechanical forces did not determine the velocity and that the important energy term was “the change in internal energy of the spindle itself. ” Perhaps I was on the right track. Today we would say that velocity is determined by the ATP turnover rate of the motor and that the viscous drag on the organelle is usually small compared with the free energy of ATP hydrolysis. In this system the rate of disassembly of the microtubules could also be significant in limiting the velocity. I spent 2 yr in the laboratory of Francis Schmitt at Massachusetts Institute of Technology as a postdoctoral fellow, learning protein chemistry and tracking down the giant squid in Chile that was our source of axoplasm. Peter Davison and I worked on the properties of the neurofilament protein that is now known to be a member of the family of intermediate filaments. Its function in neurons was not clear then and it may still be in doubt. On my return to the University of Chicago I set up my own laboratory and initially my students and I studied the birefringence of filamentous proteins but my main interest was still mitosis. I decided that the two most important problems to investigate were first, What is the protein that is assembled into the fibers of the spindle and second, What is the force generator? Corresponding author. E-mail address: ewt1@midway.uchicago. edu.