Evolution of Complexity: Single Cell to Complex Organism - Professor Dr Christiane Nüsslein-Volhard

Professor Turi King discusses the career of developmental geneticist Professor Dr Christiane Nüsslein-Volhard, and how her passion for genetic analysis led to her 1995 Nobel Prize for Physiology or Medicine.

Episode Transcript

(00:02):
Hello and welcome. You are listening to a podcast by the Milner Centre for Evolution, at
the University of Bath. I'm Professor Turi King, your host, and today I'm talking to Professor
Christiane Nüsslein-Volhard, known as Janni. She's a developmental geneticist and winner of the 1995
Nobel Prize for Physiology or Medicine. So, Janni, you're one of the founders of the entire

(00:29):
discipline of developmental genetics, asking one of Biology's great questions. So how does a single
cell go on to develop into a complex organism?But I know you didn't start there. What did you
start your research in and what caused you to change direction?
I started studying biology, which I found pretty dull because I know the most important things

(00:52):
already from school. And then I was fascinated by physics and wanted to change subjects. But at the
end, I ended up studying biochemistry in order to get a good background in fundamental things like
physics and physical chemistry and chemistry.And then I did my PhD on molecular biology,
which was at that time very fashionable. And at the end of my studies, people thought that

(01:17):
molecular biology was already sort of fading out and the most important problems were solved,
DNA replication, RNA synthesis, translation, transcription were,
sort of solved, and people thought nothing exciting is waiting for us anymore.
And at the time in the institute, there was a group working on Hydra. The director,

(01:38):
Alfred Gierer, had already foreseen the end of the interests of molecular biology and started
studying higher organisms. At other places people worked with fish or mice or drosophila. And he
worked on Hydra because this is an animal which can regenerate from very tiny pieces,
a whole organism. Fascinating problem, but it didn't develop via eggs,

(02:00):
it just developed by regeneration, essentially.And I was fascinated by the work of these people,
and there were some very interesting postdocs from the United States,
which was sort of more modern. And I was ambitious and was attracted by these American postdocs. And
they suggested to me to work on developmental biology, and I found that fascinating.

(02:21):
And I realized, however, that Hydra was not a good subject, because they were looking for
factors supporting this regeneration potential, making heads all of a sudden, and so on. But the
assays were just very poor, and there was no good way to get at the molecular principles of this.
And I thought one had to study real embryos, and I looked around and read a book on

(02:46):
developmental biology. And they had many, many different systems where they worked on,
I mean, crickets and mice and spiders and sea urchins and tons of different things,
but no real good common theme. And I felt that the approach is, well, not leading anywhere.
And in the Institute, there was another guy who worked on DNA replication, and he made mutants,

(03:09):
looking for the real replicating enzyme. And there was a mutant which knocked out DNA polymerase,
which was the only enzyme known to make DNA at the time. And the mutant still replicated,
which meant that this could not be the proper enzyme replicating in vivo. And this,
for me, was just like a revelation. And I thought, this genetics is really the thing.

(03:35):
And I thought, you have to combine genetics with developmental biology in order to get at
the right molecules, the morphogens, which would determine pattern during embryonic development.
So, I decided one has to work on an organism where you can make mutants, and you make
mutants in genes coding for morphogens. And then maybe you can try to isolate the gene product by

(04:00):
transplanting wild type cytoplasm into the mutant embryo and then isolate using this as an assay.
There are certain genes that create little things known as morphogens,
that once they reach particular concentrations, it's like they start a cascade, don't they,
of kind of reactions in different cells.Yes, essentially, they work that way, but at that

(04:23):
time it was not very clear. I mean, Lewis Wolpert proposed positional information, and he described
these gradients as providing different information to different parts of the embryonic field. And he
had the French flag model, high concentrations would determine blue and middle concentrations
white and low concentrations red. And this was a wonderful paper, but not well received by the

(04:46):
developmental biologists. He was very disappointed because they didn't accept it. Neither did they
accept Alfred Gierer's models, which were more sophisticated and would propose also
some mechanisms by which these gradients could work, but this was all still very mysterious.
So, this is like a really basic question, isn’t it? Because you’ve got this single cell,

(05:10):
and we want to know how it develops and how the cells become specialized. So, you're trying to
look for the genes that are involved in that, that's what I want to understand.
Also, drosophila was mentioned in this developmental biology textbook I read,
but only for the adult structures. And the adult structures had been studied intensely by the group

(05:32):
in Zurich. And they come from imaginal discs, and they produced the legs, and the wings, and the
eyes and so on. So, people studied the origin of these imaginal structures in the embryo, but only
using as an assay the imaginal disc structures.And they were the first experiments at that time
which dealt with the embryos. So, transplantation experiments done by Carl Illmensee, for the

(05:57):
pole cells. I mean they were these posterior cells which would make the germline. And he
transplanted the cytoplasm from the posterior pole to the anterior pole and showed that it
contains some sort of localized determinants. And this was the other concept gradients or
localized determinants. And I thought if he can transplant things maybe this is a possibility.

(06:20):
So, the drosophila would be good as an organism. And maybe if one studies the
embryo instead of the adult fly, maybe one could get at these morphogenetic substances.
Because that's the thing, wasn't it. Because drosophila are a great, fruit flies, because they
reproduce relatively quickly and you can keep them quite easily, and you can look at them under the

(06:42):
microscope and you can see different eye colours, and that's the route you ended up, kind of,
going down, because they're easy to work with as a model organism, but this idea that you could…
Well, the whole genetics of higher organisms was developed in drosophila actually, it was
the best organism for that, and the recombination was discovered,
sex determination by the Y chromosome and the X chromosome was discovered in drosophila. And

(07:05):
there were lots of markers of the adult fly, where you could do sophisticated genetic experiments,
but the embryo was not explored, really. The embryo was still, sort of, a little dark corner.
And I decided I would, I would work on it.I mean, I still admire my courage, but it
was great fun, actually. There was one guy who was a hero in the field. He was Walter Gehring,

(07:30):
he was very young still, and he had just started a group. I met him and asked him whether I could
work in his lab, and he said, well, if you have a fellowship, you can do what you like. And I went
there, and it was a very good step, actually.Yeah. I was very lonely, and no one worked on
embryos when I got there, with the exception of Eric Wieschaus, because he worked on embryos,

(07:51):
but also the aim to discover the origin of the structures of the
adult fly and not really embryonic patterns.And I started doing fly genetics, and there
was one mutant which had been described in the literature, which was possibly encoding something
like a morphogen, at least the embryos which were born to mothers, which were mutant for this

(08:15):
particular gene bicaudal, developed very curious duplications of the hind end only and no head,
no thorax, and this was a fascinating phenotype. Nobody could explain it, except you can describe
it as a gradient duplication. But it was mysterious, and the mutant was
supposed to be lost, and I tried to get it back.There was a stock which was labelled bicaudal,

(08:41):
question mark, and I took this stock and tried to analyse whether the mutant was
still in this stock. And it was, and I found it and then studied this gene. But it was very
frustrating because only a very small number of the embryos showed the mutant phenotype.
I mean, the females are all the same, but only a few embryos showed the phenotype.
And I wondered whether this could be really important because the embryos can

(09:04):
also develop without this gene product, apparently. So, it was not a good mutant, but it was at
least showing that you can get these mutants.So, I tried to get new alleles, meaning hits
in the same gene, which might be more strong or would provide a higher fraction of the phenotype.
And in the course of these experiments, which was one of the first experiments I did in flies,

(09:26):
I discovered a gene which caused female sterility. And when I looked at these embryos, they had a
very weird and strong phenotype. And in this case, all of the embryos showed the phenotype,
complete absence of the ventral structures. And this was something which had never been described
before, and it wasn't fascinating phenotype. And actually, it turned out later, this was

(09:48):
one of the most important morphogens in the fly.It was my first experiment in flies, and I hit...
You hit jackpot.Yeah, I hit the jackpot.
When you get a mutation in a particular gene, you're looking at what are the changes you can
see. So, the phenotype is what you can actually, kind of, see are the changes. And you've hit on
something that has quite a serious impact on the phenotype of the fruit fly. So,

(10:12):
what the fruit fly looked like. And I know that this is something that eventually,
kind of, morphed into the work that became what you won the Nobel Prize for, with Eric Wieschaus.
What was the big question that you were trying to answer, and how were you doing it?
Eric and I chaired a lab in Heidelberg in the newly developed European Molecular Biology

(10:35):
Laboratory, which was a European place where young scientists, only junior groups were established,
and we were lucky in getting one of these junior groups, albeit joined. I mean,
we had two heads and no body essentially. We had one technician, and this was our group.
And we both had our own projects initially, but we were both very keen on embryonic development.

(10:58):
And I worked on the maternal and Eric and other things and then in the course of these studies,
we jumped across some mutants which were not maternal but in the embryo itself and had very
dramatic phenotypes in the segmentation.And we wondered whether there would be
more of them, because in order to understand development, you cannot just start one gene,

(11:20):
you have to anticipate that there are many genes.And people at that time thought there were
millions of genes, and you couldn't figure that out because it's too difficult.
And we were just courageous and said, we do a big screen and see whether we can find all the genes,
which are significant for the embryonic development in showing a specific phenotype,

(11:41):
which would pop up in the cuticle of the larva, not the adult fly, but the larva.
So, we chose the larva as our object instead of the adult fly, and I think this was the big
step and I had developed previously methods to make visible the structures of the larva,
and this was very important to count segments, to see the polarity of the segments,

(12:03):
see the head structures and end structures. And this technique was very important and made the
larva an object for genetic analysis.And this was essentially the question,
can we use this larva and identify the genes which are really important for this pattern?
And it turned out that although many mutants in the embryo were lethal, but most of the larvae

(12:26):
look normal. I mean, I think we've defined finally, 120 genes where this was not the
case. They were not normal, they were abnormal. We could see a distinct and specific phenotype.
The phenotypes were all different. I mean, they were not always the same. And it was
not just a mess. It was very specific number of segments or polarity or lacking certain

(12:49):
structures. And it was very clear that this was a set of genes which would be really
informative for the embryonic development.This was a grand work, but we finished it.
We finished it within three years, working pretty hard, but it was great fun. It was
like looking for mushrooms essentially, you know, oh, you go around the corner and there

(13:09):
is a new thing which you discover. And it was very exciting. I mean, the phenotypes were so
curious and different that it took some while until we could sort them out and order them.
And this was also left for the next generation of geneticists and developmental biologists.
But we’ve published this work, first on the segmentation genes, because they were easy,

(13:33):
because you can count segments and you determine polarity. And it was not so difficult to order
them into classes and found that, segmentation is hierarchically organized, in to first,
large regions of the embryo, then double segments, and single segments.
And this made a splash actually, we had a paper in nature in 1980,
which was… people just jumped and can't believe it, and they were completely stunned. Well,

(13:57):
we didn't have much of an explanation, but we had these genes. And the Nobel requires that you
work on human beings. I mean that you make a step forward in understanding human beings. And this,
of course, our work was basic biology and no idea of medicine or of human development. Also,
because we knew that vertebrates were developing completely differently from

(14:20):
flies. At least that was the notion at the time.But then these mutants, were so attractive for
many, many biologists. And very quickly a large community arose of developmental
geneticists working on our mutants.And they cloned the mutants, meaning they
identified the gene products, and they discovered many very important protein families, which

(14:44):
had features which can induce structures, start signalling cascades, determine patterns, and would
distribute in the embryo in specific manners.And these gene classes then turned out to be
conserved also in vertebrates, which was completely a new finding at the time. But

(15:05):
it turned out that there is a common ancestry, and there is a sort of, toolkit which had evolved
in the two different directions, but still was recognizable also in human development.
This was then a possibility to give us the Nobel Prize, although we worked on
flies only and never touched a vertebrate.So let me take you back a bit. Explain what

(15:29):
drosophila development looks like normally and how you were trying to look for the mutations,
the differences, the genes that are telling you about what are the important genes in development.
The normal development essentially starts with this zygote, and in the freshly laid egg there is
just the zygote, meaning a fertilized nucleus, and this divides several times synchronously

(15:52):
until about 6000 nuclei are reached.And then they distribute in the cortex
of the egg to form a uniform cell layer, with the cellular blastoderm.
And these cells then acquire different fates, which you can already see in the expression of
certain genes. Although the cells still look completely undistinguishable, they're all the

(16:12):
same, but you can see that already at this stage, several genes are expressed in a differential
manner, some with very broad stripes and some with seven stripes and some with 14 stripes.
And then the cells start dividing. They start making shapes, meaning they start

(16:33):
getting different. Invaginate in the head region, invaginate in the ventral region,
invaginate in the posterior tail to make the gut, in the ventral region to make the mesoderm,
under the musculature, and so on. And in the head, the brain region also.
And these cells become different and express different genes at different

(16:53):
times. And these genes all interact and somehow determine the next step
of development, via the interaction of the cells. And the nervous system is formed, gut is formed,
in rather complicated morphogenetic movements, which took us a while to figure out. And there was
no good description at the time, but we learned.And we had these molecular probes eventually,

(17:18):
meaning that we had antibodies against the gene products, and we had messenger RNA,
antisense RNA, which you could make in vitro, following the sequence of the genes.
And we could probe and see where the genes are expressed and what happened in the mutants.
And this was very important, and many people did that. And then gradually a picture arose,

(17:40):
and we figured out how the embryo would form.And then what you want to do is introduce
mutations into genes individually, like one gene at a time, and try and see if you can work out
what is that doing in the development. And you did this on a massive scale.

(18:01):
Yeah. Our first question was simply, at the time people hadn't studied these genes at all. They
were just very few known, which were studied intensely. But we figured that if you only
study one particular gene, you have to know all the others which would interact with them. So,
we first did this survey, and essentially, I left this field and said, let the other people

(18:24):
work out what these genes are doing. I want to go back to the original question, to the morphogens.
And this meant maternal information, and this was not solved yet. And so,
we did screens for the maternal genes. This was much more difficult, and it took us
another couple of years until we also had the set of maternal genes, which were determined.

(18:46):
And they were all encoding morphogens, and they were showing that the embryo essentially is built
by the maternal information, which provides gradients of positional information. And there
were three different sorts of gradients. I mean, it was not that simple. It was not just anterior
posterior, it was a bit more complicated, but we discovered four systems making gradients in the

(19:10):
egg. So, this was already the information which was given to the embryo when the egg was laid.
And then on the basis of these gradients, the psychotic genes we had discovered earlier,
they worked on and built the shape of the embryo.And the really interesting thing is, I know you
went on and worked with zebrafish because you were interested in the vertebrates,

(19:30):
but what you found and you talked about this a little bit earlier, is your finding that rather
than it being different across various kind of groups of organism, you're finding, as you say,
it's like a tool kit of sets of genes that are, kind of, conserved across different groups. And
it's the same ones across all of them.To a large extent, yes. I mean,

(19:52):
we essentially discovered in the fly the genes which made the most important signalling systems,
also invertebrates. But at the time when we had done this work on flies, the homology was
not known. It was not known that they were the same origin. There was never the same genes,
it was just the genes derived from some common ancestry, would determine both vertebrates and

(20:16):
invertebrates. This was not known, and we were wondering how much what we found was also hold
for the development of higher organisms, higher organisms, meaning vertebrates essentially.
At the time people worked on frogs mostly, and they have big eggs, and they have a
generation time of two years, and you can't do genetics, absolutely not. And it was also clear

(20:39):
that the way embryonic development in frogs was described was completely different from flies.
In flies one always talked about genes influencing each other and the determination by genes,
and in frogs one always talked about factors. So, factors determining and the factors which
you could isolate by these transplantation and cutting experiments. But they didn't get

(21:00):
anywhere the frog people. Although there were some cute experiments where it was shown that
certain tissues from particular regions would have long ranging influences, fascinating stuff,
but people hadn't been able to isolate anything on the basis of these experiments.
So, I thought arrogantly, genetics is the way and if you can't do genetics then forget it,

(21:23):
you can't get the answer to these factors, which was not true. They did
finally find the right determinants.So, I decided one had to do genetics
on a vertebrate organism, and this is where I decided to work on zebrafish,
which was already being studied in Eugene, Oregon by George Streisinger, and colleagues.

(21:44):
So, I started developing the genetics of the fish, which was a much bigger operation. But we finally
did large scale screens, and we finally ended up with 1200 or so new genes and some of which
later on turned out to be homologs to, what we had found in flies, but many were new,
because vertebrates have, apart from some basic principles which are common to both classes of

(22:10):
phyla, there are some big differences to. And there were some structures which just
happened to be vertebrate specific, and there's lots of cell migration involved,
for example. And so, the classes we found were much more divergent than what we found in flies.
But again, we had just a collection of genes and then provided that to the people and said,

(22:30):
look work on them. And of course, in the case of the flies, just Eric and I worked on it,
and then Gerd Jurgens who joined as a postdoc later. But in the case of the fish, we had a large
group of people who were involved, and we also had invited guests to analyse some of the genes, which
came from outside and took the genes with them and worked on them, for example on ears, and on blood

(22:53):
and on heart, which we were not interested in.And in my lab only a few people
remained to continue working on these genes, whereas most of the work was
done in laboratories of my ex-collaborators, who set up their own labs with these mutants.
It sounds like you've had some really lovely collaborations, and I saw this lovely interview

(23:15):
of you with Eric, and you're talking about how you built this microscope so you could
both look at the same time, and you could be looking down the microscope, you've been
mutating little genes and trying to work out what the differences are, and you would be,
kind of, competing against each other a little bit to, kind of, who could spot which one first.

(23:36):
I don't know whether the competing is the right word, I think challenging each other. I mean,
it was very important because you get tired easily, it's quite a tedious task. And to
look at it and sometimes you are… but when you are two of you, then you must not be beaten by
the other one. So, you must recognize the things at the same time, sort of.

(23:57):
And it was quite amazing, and the big luck that we both were quite good at it,
and we were probably as good. So, there was not one overriding the other one,
it was more a challenge and a stimulation.And that must have been fantastic because
you've got this was amazing…Yeah, it was a big luck. We knew
very well at that time that no one else in the world would have been able to do such an

(24:22):
experiment. We were just unique in this respect because we knew the embryo and we knew the larva.
But of course, is it important? Does it interest people? This was another big question because who
is interested in fly larvae? But when the first genes were cloned, lots of molecular biologists
jumped because this was the system where you could discover, for example, cascades

(24:46):
of transcription factors determining structures.So, Mark Ptashne and other transcription factor
people jumped into the field of drosophila, because these genes were really much more
informative, and had phenotypes which you could check when you did your molecular experiments,
that this system developed into one of the most fashionable systems in biology, I think, Yeah.

(25:12):
I mean, I can, as a geneticist, see just how exciting that must have been to be able to go,
oh my god, there's a little mutant there, I wonder what's doing that? And that understanding
of really fundamental biology, how things…Yeah, and then the nice thing is also that
we had this large catalogue, we had the interactors. So, when you studied one gene,

(25:34):
the genes they interacted with were already there, you just had to identify them, and
you didn't have to find them yourself. And it was very successful and very challenging for people.
So, Phil Ingham, our lovely head of department, who I know knows you well, was telling me a story
that when you were thinking about having people come and work in the lab, I don't know if he got

(25:58):
this right, but that you would have people come to your house and cook, and that was
almost part of the initiation and the checking.No, it’s a little exaggerated. Traditionally,
and the tradition started already in Heidelberg. At Christmas time I invited the people from the
lab to come to my place in the evening and make Christmas cookies. And there I just observed

(26:23):
that the people were very good in the lab, they were also very good at making these cookies. And
some who were, sort of, doing shortcuts all the time and messed up the experiments they
also did that when they did the cookies and they were never pretty, as pretty as others.
Yeah, and sometimes we had people who were really clumsy, and I thought maybe it's the best when you
interview them to let them peel an apple and see how they behave. But I'd never did that, really,

(26:48):
but I thought it would have been a...Good idea?
Yeah.So, you've had this
incredible scientific career, which only really just finished last year. And if you look back
over your life, what’s the thing you are most proud of, coming out of your scientific career?
I think the discovery of the cuticle pattern, as an assay to look at mutants,

(27:14):
this was probably the biggest impact, I think, and it was done in my first half year in Basel,
actually, when I had this mutant with these mirror images, and people couldn't describe it because
they couldn't see the segments well in the embryo.And I had to see the segments, absolutely. So,
I squashed the embryos and found the segments. And then I developed this way of making these preps,

(27:39):
also from the literature of course, other people had similar tasks,
but this was probably the biggest impact.It’s that being able to even visualize it,
which you weren't able to do before.And the combination, I mean, to really
jump on the combination between development and genetics, which other people hadn't done,
although they could have seen that, because the molecular biologists work on bacteria and phages,

(28:05):
which are easy to work with because they are haploid, whereas higher organisms are
diploid. So, it's more complicated, you have to learn them and then learn yours,
and it’s sort of more complicated. But somehow the molecular biologists didn’t grasp that.
And actually, the first paper where I described bicaudal and also this dorsal mutant, which the
first morphogenetic mutant I found in Basel was published in a meeting report. I mean,

(28:31):
it was Society of Developmental Biology yearbook, and it was not easily accessible. But this is the
paper which probably founded the drosophila development of the genetics of the larva.
One of the biggest fields in genetics.Yeah.
Started with you.Yeah.
Janni, it has been so much fun talking to you. Thank you so much.

(28:54):
This was a podcast by the Milner Centre for Evolution. I'm Turi King and thank you for
listening. If you have any thoughts or comments on this or any other
episodes, please contact us via ourX channel @MilnerCentre.
For more information about the Milner Centre for Evolution, you can visit our website.