And that was the genius of John Gurdon’s work. When he performed his experiments what he was attempting was exceptionally challenging with the technology of the time. If he failed to generate toads from the adult nuclei this could simply mean his technique had something wrong with it. No matter how many times he did the experiment without getting any toads, this wouldn’t actually prove the hypothesis. But if he did generate live toads from eggs where the original nucleus had been replaced by the adult nucleus he would have disproved the hypothesis. He would have demonstrated beyond doubt that when cells differentiate, their genetic material isn’t irreversibly lost or changed. The beauty of this approach is that just one such toad would topple the entire theory – and topple it he did.
John Gurdon is incredibly generous in his acknowledgement of the collegiate nature of scientific research, and the benefits he obtained from being in dynamic laboratories and universities. He was lucky to start his work in a well set-up laboratory which had a new piece of equipment which produced ultraviolet light. This enabled him to kill off the original nuclei of the recipient eggs without causing too much damage, and also ‘softened up’ the cell so that he could use tiny glass hypodermic needles to inject donor nuclei. Other workers in the lab had, in some unrelated research, developed a strain of toads which had a mutation with an easily detectable, but non-damaging effect. Like almost all mutations this was carried in the nucleus, not the cytoplasm. The cytoplasm is the thick liquid inside cells, in which the nucleus sits. So John Gurdon used eggs from one strain and donor nuclei from the mutated strain. This way he would be able to show unequivocally that any resulting toads had been coded for by the donor nuclei, and weren’t just the result of experimental error, as could happen if a few recipient nuclei had been left over after treatment.
John Gurdon spent around fifteen years, starting in the late 1950s, demonstrating that in fact nuclei from specialised cells are able to create whole animals if placed in the right environment i.e. an unfertilised egg[6]. The more differentiated/specialised the donor cell was, the less successful the process in terms of numbers of animals, but that’s the beauty of disproving a hypothesis – we might need a lot of toad eggs to start with but we don’t need to end up with many live toads to make our case. Just one non-murderous doctor will do it, remember?
So John Gurdon showed us that although there is something in cells that can keep specific genes turned on or switched off in different cell types, whatever this something is, it can’t be loss or permanent inactivation of genetic material, because if he put an adult nucleus into the right environment – in this case an ‘empty’ unfertilised egg – it forgot all about this memory of which cell type it came from. It went back to being a naive nucleus from an embryo and started the whole developmental process again.
Epigenetics is the ‘something’ in these cells. The epigenetic system controls how the genes in DNA are used, in some cases for hundreds of cell division cycles, and the effects are inherited from when cells divide. Epigenetic modifications to the essential blueprint exist over and above the genetic code, on top of it, and program cells for decades. But under the right circumstances, this layer of epigenetic information can be removed to reveal the same shiny DNA sequence that was always there. That’s what happened when John Gurdon placed the nuclei from fully differentiated cells into the unfertilised egg cells.
Did John Gurdon know what this process was when he generated his new baby toads? No. Does that make his achievement any less magnificent? Not at all. Darwin knew nothing about genes when he developed the theory of evolution through natural selection. Mendel knew nothing about DNA when, in an Austrian monastery garden, he developed his idea of inherited factors that are transmitted ‘true’ from generation to generation of peas. It doesn’t matter. They saw what nobody else had seen and suddenly we all had a new way of viewing the world.
Oddly enough, there was a conceptual framework that was in existence when John Gurdon performed his work. Go to any conference with the word ‘epigenetics’ in the title and at some point one of the speakers will refer to something called ‘Waddington’s epigenetic landscape’. They will show the grainy image seen in Figure 1.1.
Conrad Waddington was a hugely influential British polymath. He was born in 1903 in India but was sent back to England to go to school. He studied at Cambridge University but spent most of his career at the University of Edinburgh. His academic interests ranged from developmental biology to the visual arts to philosophy, and the cross-fertilisation between these areas is evident in the new ways of thinking that he pioneered.
Figure 1.1 The image created by Conrad Waddington to represent the epigenetic landscape. The position of the ball represents different cell fates.
Waddington presented his metaphorical epigenetic landscape in 1957 to exemplify concepts of developmental biology[7]. The landscape merits quite a bit of discussion. As you can see, there is a ball at the top of a hill. As the ball rolls down the hill, it can roll into one of several troughs towards the bottom of the hill. Visually this immediately suggests various things to us, because we have all at some point in our childhood rolled balls down hills, or stairs, or something.
What do we immediately understand when we see the image of Waddington’s landscape? We know that once a ball has reached the bottom it is likely to stay there unless we do something to it. We know that to get the ball back up to the top will be harder than rolling it down the hill in the first place. We also know that to roll the ball out of one trough and into another will be hard. It might even be easier to roll it part or all of the way back up and then direct it into a new trough, than to try and roll it directly from one trough to another. This is especially true if the two troughs we’re interested in are separated by more than one hillock.
This image is incredibly powerful in helping to visualise what might be happening during cellular development. The ball at the top of the hill is the zygote, the single cell that results from the fusion of one egg and one sperm. As the various cells of the body begin to differentiate (become more specialised), each cell is like a ball that has rolled further down the hill and headed into one of the troughs. Once it has gone as far as it can go, it’s going to stay there. Unless something extraordinarily dramatic happens, that cell is never going to turn into another cell type (jump across to another trough). Nor is it going to move back up to the top of the hill and then roll down again to give rise to all sorts of different cell types.
Like the time traveller’s levers, Waddington’s landscape at first just seems like another description. But it’s more than that, it’s a model that helps us to develop ways of thinking. Just like so many of the scientists in this chapter, Waddington didn’t know the details of the mechanisms but that didn’t really matter. He gave us a way of thinking about a problem that was useful.
John Gurdon’s experiments had shown that sometimes, if he pushed hard enough, he could move a cell from the very bottom of a trough at the bottom of the hill, right the way back up to the top. From there it can roll down and become any other cell type once more. And every toad that John Gurdon and his team created taught us two other important things. The first is that cloning – the recreation of an animal from the cells of an adult – is possible, because that’s what he had achieved. The second thing it taught us is that cloning is really difficult, because he had to perform hundreds of SCNTs for every toad that he managed to generate.
6
Key papers from this programme of work include: Gurdon et al. (1958)