Dr Zon & The Zebrafish: The Origin Story

As promised, here is part two of our talk with Harvard and HHMI investigator Leonard Zon, in which he shares with us why he left frogs, how he came to work on zebrafish, and the importance of getting by with a little help from your friends. (Click here for Part I).

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Illustration by Madalena Parreira.

You spent some time working on Xenopus?

LZ When I started my lab, I thought I wanted to do mouse genetics. I went over to MIT on a Friday afternoon and dissected out day 7.5 mouse embryos, looking for the very first blood island. It took us 6 hours and at the end of that time I had a dish that had 6 embryos in it. I realized that everything that I wanted to do wasn’t going to be possible. So I was pretty depressed. I came back to the lab and I saw a friend of mine, Celeste Simon, and she said that I looked terrible. I said I’d had this bad experience with what I wanted to start my lab in. I didn’t think it was practical anymore. She said, “Well I can’t help you with that. But I’m having a party at my house and I’d like you to come”.

So I was there at the party and I had a beer in my hand and this guy walks up, and his name is Jerry Thomsen, who’s at Stony Brook. Jerry worked on frog embryos in Doug Melton’s lab. So we started talking and I told him about my mouse experience. He said, “You know, you really have to think about an externally fertilized animal, because you could have thousands of embryos, like a frog. Then everything will be one-cell, two-cell, four-cell… and eventually you would make blood and you could study that process, in a facile system”. The next week I made an appointment with Doug and started doing frogs. A couple of years in, I realized that the frog didn’t have very good genetics. There were no morpholinos or anything at that time, so everybody was not really doing anything except injecting a dominant negative construct in the frog. I felt like I needed to switch systems, so I decided to switch to the zebrafish.

I could tell you more about how I did that, or we could go on. I had kind of an amazing week that transformed my group into a zebrafish lab.

That sounds like an interesting story.

LZ I went to a hemoglobin switching meeting and presented my frog work. After the presentation this transgenic mouse person, Frank Grosveld, came up to me and said, “You know, I really felt like your talk was fantastic. The frog is a good system but, you’re never going to do genetics. Have you heard about the zebrafish as a model system?

I actually had been thinking about switching to zebrafish already because Janni (Christiane) Nusslein-Volhard was going to start doing zebrafish. So I’d been thinking about it. We spent actually about an hour together. He helped crystalize the reasons for going into a new model system, how it could be a good thing. So I came back really charged up about zebrafish. Literally the next day, I had a call from Bill Dietrich, who’s an investigator at Northeastern University. He worked on the Antarctic icefish. The icefish has lost its red blood cells, because it is a competitive disadvantage in cold water to have red blood cells, you end up stroking out. So he convinced me that I should study cold adaptation of transcription in this icefish. I said, “I don’t care about icefish, what about zebrafish?” He said, “That sounds really interesting, sign me up for a sabbatical.” So I had my first person on day 2. Then, on day 3, an investigator in Wally Gilbert’s lab at Harvard called. He had been doing a few zebrafish experiments and found a mutant fish that had no blood. They were going to throw it out, but they said, “We heard you like these things, so you can have it.” So that was it.

Now you have over three thousand tanks?

LZ Yeah, it’s actually three thousand. It’s amazing. Three hundred thousand fish. It’s actually split in two facilities right now.

You identified the first human disease-causing gene in zebrafish. Can you tell us a bit about that?

LZ It was very interesting in the early days. We had very few reagents to do research. We had a mutant where we did a chromosomal walk of a megabase. Literally took us about three years. We found the gene. This mutant was Sauternes. Sauternes ended up being an ALAS2 mutant- this is one of the major steps in heme biosynthesis. It was a human disease gene. So we really found the first human disease-relevant gene in fish. That was really exciting. We published a Nature Genetics paper. About half of the community was very excited about it, that it was possible to clone this type of gene, and the other half was “Well, we already knew about that gene, because it was already a human disease gene. Will you ever find anything novel?”

Luckily our second gene was Weissherbst. We did a comparative approach to actually clone this gene. The human locus and the fish locus had the genes in the same order. So it was nice because we had candidates to walk through this locus. We found this gene which we named ferroportin, which is an iron exporter. This mutant had iron in the yolk, but it couldn’t get it to the baby, because it missed the transporter to do that. That was a very exciting gene because it was completely novel.

When we looked at it, we realized that it was evolutionarily conserved, because the human gene was there. We made antibodies to the human gene product, and we found that it had binding in the placenta. This yolk sac iron transporter and the placental iron transporter were the same. Mothers get their babies iron through this ferroportin. Then we started staining adult tissues and we found that in the duodenum there was specific staining for ferroportin. What we realized was that there was this basolateral iron transporter in the gut that many people had hypothesized to exist, and shown it should be there. It turned out to be ferroportin. That was also exciting, and later we found mutations in people that had hemochromatosis. So it became not only a novel gene, but the first time that a zebrafish mutant predicted a new human disease.

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