Dolly the sheep has mystified the concept of cloning

Prof. Dr Martin Simon on the history and development of cloning

25 years ago, the first mammal was cloned at the Roslin Institute near Edinburgh in Scotland, a sheep that became world famous with the name "Dolly". A process thought to be impossible became reality. Martin Simon, a molecular and cell biologist from Wuppertal, talks about the opportunities and risks of duplication, which has been practised in agriculture for centuries.

The entire scientific world looked to Scotland in amazement on 5 July 1996 when the birth of a sheep was announced in Edinburgh. What was unusual about it were the circumstances, for the animal, named Dolly, was the first mammal cloned from a differentiated somatic cell. "It was a major event", knows Wuppertal molecular and cell biologist Martin Simon, "it was a masterpiece both in terms of craftsmanship and science, but it also mystified the concept of cloning". Many ethically sensible and necessary discussions were initiated on it, interestingly often analogous to science fiction literature, which has often rehashed and critically discussed the topic.

Core transfer is tricky

The scientific community was very surprised, Simon knows, because the process was a combination of a successful nuclear extraction, but also a nuclear transplant. "An adult somatic cell was introduced into a foreign egg cell, meaning there was no genetic manipulation." This differentiation is important, he said, because the genome was not tampered with; the researchers did not add, remove or change any genes. An intact cell nucleus of a cell of an adult sheep was transferred into an egg cell; in the process, a reprogramming of the cell nucleus took place. At that time, this was a special dexterity, because today we have special micromanipulators with joystick control that can position an extremely thin needle precisely to the nanometre. These needles are also optimised so that the cell nuclei are not damaged.  This precision did not exist in 1996 and one should not forget the several hundred failed attempts that preceded the Dolly success. "We only see the result of what worked," says Simon, "but this nucleus transfer is tricky. Implanting embryos into a so-called surrogate mother is also complicated. That's not 100% efficient even today, there's also only a small percentage of about 25% of embryos that actually grow."

Identical twins - a natural clone

However, natural clones have also existed since time immemorial, he said, and these are identical twins. "With the identical twin, we are talking about what makes a clone!" The concept of clone in relation to the mammal has been misled by science fiction literature, he said. "In the case of a genetically identical organism - which is what every identical twin has, quite naturally - this mystification forgets all about epigenetics. Genetically identical does not mean phenotypically identical or identical in nature and appearance. After all, since the first human genome projects were published around 2000, we have also learned that genetics is not everything." There is also information transfer outside the DNA, explains Simon, which also has a very strong influence on us. That's why identical twins are different, he says. "One twin might become more corpulent, the other one runs eight kilometres every day and stays slim. And already we have a difference." In the course of life, these differences accumulate.

There are no identical clones

Strictly speaking, the miracle sheep was not an identical clone either, because our mitochondria, the so-called power plants of the cells, are inherited exclusively on the mother's side. "Our mitochondria have their own genome. I.e. if I transfer a cell nucleus into an egg cell from another organism, as was done with Dolly, then I transfer the chromosomal nuclear balance, but the mitochondria are inherited purely on the mother's side and are therefore the mitochondria of the donor organism. In that case, one cannot speak of genetic equality, because strictly speaking, only the nucleus is identical." But the epigenetic differences must also be taken into account. "Our cells also undergo genetic changes during ageing," says the scientist. We all accumulate small genetic changes in our body cells in the course of our lives, both due to errors in copying the DNA during normal cell divisions, and indicated by the environment, e.g. UV radiation.  It is also normal "that the so-called telomeres of the chromosome ends shorten a little with each individual cell division. This also happened with the cells of the dolly mother. So the telomeres were significantly shorter than in a freshly fertilised egg cell. This also accounted for the age difference. The cloned sheep Dolly had just been born through the surrogate mother, but its cells were at the cellular level of a six to seven year old sheep, namely the donor. So you can produce genetically identical individuals, but you can't expect them to be identical to their donors." In addition, in humans there is the fact that neither the essence nor the experience can be passed on and it is almost impossible to eliminate this age effect from the somatic cells.

No clone warrior in sight

It is just as impossible to create adult clones at the moment, says Simon, because "even if we were to take a somatic cell from me, it would mean that it would have to be transferred into a human egg cell, which would then transfer it into a surrogate mother. But in the end it would be a small child. I wouldn't have an identical clone." In Simon's opinion, we are a long way from the sci-fi imaginings of the Star Wars saga, in which adult clone warriors are created, equipped with abilities that then no longer need to be trained.

Domestic and farm animal cloning

After Dolly, researchers have cloned other mammals such as mice, goats, cattle, pigs, cats, dogs, horses or even monkeys for research purposes. Not all results have been satisfactory, ethically questionable or carried out for economic reasons. "A good example is cats," Simon tells us, "with them it is very exciting, because there is an epigenetic phenomenon. In female mammals, an X chromosome is silenced. In us, the sex definition is determined by the XY system.  But female mammals have two X chromosomes, one of which is silent in the nucleus," he explains. "In cats, it is now the case that with one X chromosome, the coat colour is defined. And this chromosome activation happens randomly. It is not controlled and it also happens during the embryonic phase, when the embryo gradually forms from the cell cluster. And if now one X chromosome codes for a grey coat colour and the other X chromosome codes for a red coat colour and this all happens by chance, they can also generate a genetically identical cat, but which later looks totally different. That's why the cat cloning industry has actually not been successful, because people won't pay money for a grey cat if they wanted a red cat." Significantly more work goes into cloning strong bulls because they can generate large amounts of semen for the industry. And it has also been tried with racehorses, but the psychological characteristics of the animal are not transferable either. In all these processes, which could certainly also be seen as crossing borders, it is often a question of the expected benefit. The reliable breeding bull is a sales argument, Simon explains, because it always delivers the same semen quality, whereas he sees no need for dog cloning of millionaire pop stars.

A mammoth for the city zoo?

Due to the melting of the permafrost, more and more 'fresh mammoths' are coming to light in Russia. There are also efforts to transplant cells of this extinct animal into today's elephants using the CRISPR method and have them reproduced. Is science allowed to interfere with evolution in this way? According to the scientist, such an animal would only end up in a zoo and serve as a sensational object. "The mammoth's ecosystem no longer exists, and even if intact cells could be found, it would be a very questionable undertaking," he says. Besides, one or two individuals would not be enough to build up a complete population with this gene pool. "The consideration makes more sense in relation to the many species we are eliminating at the moment. Because perhaps at some point it may be necessary to repopulate a renaturalised ecosystem with a species that is adapted accordingly. However, it would make more sense at the moment not to have to resort to this option at all by intensifying species protection and stopping the destruction of ecosystems instead of thinking about artificial ecosystems à la Jurassic Park."

Reproductive cloning versus therapeutic cloning

"When we get to human cloning, we need to distinguish reproductive cloning and therapeutic cloning," says Simon. "The reproductive would be when you create a new human being, i.e. transferring the procedure of somatic nucleus transfer from the clone sheep Dolly." In addition to the ethical dubiousness, there would be the huge amount of eggs needed.  This approach is clearly excluded by the German constitution. Therapeutic cloning has to be distinguished.  "In the classical variant, a somatic cell nucleus is also transferred into an egg cell. However, the resulting embryo is not transplanted into a surrogate mother, but embryonic stem cells are taken, which are then to be differentiated with various growth factors into specific cells/tissues. This procedure is also prohibited by the German constitution as it involves embryonic stem cells." If epigenetics were better understood, however, the ethically critical step of embryonic stem cells could also be bypassed.

"The goal would be the induced re-programming of a somatic body cell, from which one could then regrow individual cell types or tissues in the culture dish. The individual factors, such as growth factors, that are necessary for this are not yet understood enough to carry this out. Similarly, in somatic nuclear transfer, it is not yet well understood how the egg cell reprograms the transplanted nucleus." Simon cites, for example, the transplantation of new nerve cells into a severed spinal cord to restore this connection as fully legitimate work in the medical sense of this type of therapeutic cloning. Reprogramming somatic cells into stem cells to differentiate new cells without an embryonic intermediate stage would be a possibility for the scientist, after which organ transplantation with the patient's own tissue would be possible.

Agriculture works permanently with clones

The layman may not really be aware of it, but working with clones has been part of agriculture for a long time. "Z. For example, all the normal propagation of cuttings," Simon explains, "these are genetically identical organisms, or plant parts. Winegrowers do it by default, which means that usually the entire vineyard is a clone of an individual vine. It is propagated via cuttings. And we also talk about cloning when we plant potatoes," the expert knows.

One has to differentiate between what is nature-made and what is man-made, Simon emphasises. "Nature is constantly working with clones. Every single-celled organism, every bacterium, undergoes rapid mitotic division, i.e. it functions clonally, because that is its reproduction strategy. The situation is different with multicellular organisms, including humans, which reproduce sexually and thus constantly recombine genetically.

"The reproductive cloning of Dolly has given the topic new impetus, it has triggered basic research," Simon sums up, "but things change. People used to try to explain everything through genetics and completely underestimated epigenetics." Today, new epigenetics programmes examine the differences between the various cells outside of DNA in order to clarify the question of which factors determine the activity of a gene and thus the development of the cell.

"Cloning represents an extraordinary test of human restraint and wisdom, but also of institutional development. In many ways, this will shape the morality of the 21st century," says Glenn McPhee, director of the Centre for Bioethics at the University of Pennsylvania. We can only hope that we will pass this test.

Uwe Blass (conversation from 14.12.2021)

Martin Simon studied at the TU Kaiserslautern until 2005 and then became a junior professor at Saarland University in 2012. Since 2018, he has been head of the Molecular Cell Biology and Microbiology Department in the Faculty of Mathematics and Natural Sciences at Bergische Universität.