Field of Science

Who really discovered trisomy 21? (righting an old wrong)

A few days ago a French student in my Useful Genetics Coursera course posted a link to an article in Le Monde (sorry, it's both in French and behind a paywall, but this link might get you a translation).  It reported that a Jan. 31 award ceremony for the discovery of the cause of Down syndrome, part of the 7th Human and Medical Genetics Congress  in Bordeaux, had been blocked by a Down syndrome support organization (Fondation Jerome-Lejeune).  The back story is very depressing, an egregious example of a woman scientist being denied credit for her discovery.

Photo source: Le Monde
The woman is Dr. Marthe Gautier, now 88 years old.  In 1956 she was a young physician, returning to Paris from a year's study of pediatric cardiology at Harvard.  She was given a clinical/teaching position at a local hospital, with no funds for research.  The Head of the Pediatric Unit, Raymond Turpin, was interested in mongolism (as Down syndrome was then called); years earlier he had proposed that it might be caused by a chromosome abnormality.  Human cytogenetics was not well understood, but a big breakthrough came this same year, when the true chromosome number was finally established as 46 (not 48).  When Turpin complained that nobody was investigating his hypothesis, Gautier proposed that she take this problem on, since her Harvard training had introduced her to both cell culture and histology.  Turpin agreed to provide a tissue sample from a patient.

For this work she was given a disused laboratory with a fridge, a centrifuge, and a poor quality microscope, but no funding.  And of course she still had her other responsibilities.  But she was keen and resourceful, so she took out a personal loan to buy glassware, kept a live cockerel as a source of serum, and used her own blood when she needed human serum.

By the end of 1957 she had everything working with normal human cells, and could clearly distinguish the 46 chromosomes.  So she asked Prof. Turpin for the patient sample.  After 6 months wait it arrived, and she quickly was able to prepare slides showing that it had not 46 but 47 chromosomes, with three copies of a small chromosome.  But her microscope was very poor, and she could not identify the chromosome or take the photographs of her slides that a publication would need.

All this time Prof. Turpin had never visited her lab, but she'd had frequent visits from a protege of his, Jerome Lejeune.  When she showed Lejeune her discovery, he offered to take the slides to another laboratory where they could be photographed.  She never saw the slides again, but the photographs appeared in Montreal two months later (August 1958), where Lejeune announced to the International Conference of Human Genetics in Montreal that he had discovered the cause of Down syndrome!  Lejeune and Turpin quickly wrote up 'their' discovery, with Gautier as middle author, but Gautier only learned about this publication the day before it appeared in print.

These were tough times for a woman scientist in France, and Gautier decided not to fight for the credit for her discovery, instead returning to her clinical and teaching work on congenital heart diseases.

Lejeune became not just a renowned researcher but the darling of the French Catholic right-to-life movement.  You can read long flattering Wikipedia biographies in both French and English. He was showered with awards and given a prestigious Chair of Human Genetics at the Paris School of Medicine, bypassing the usual competition.

When prenatal diagnosis became available Lejeune campaigned against it on religious grounds. He became a friend of Pope John Paul II and was appointed President of the Pontifical Academy for Life (Wikipedia), the Catholic think-tank for medical ethics.  He died in 1994.  The Fondation Jerome-Lejeune was established in his honour; there's an American branch too.  This foundation provides funds for research into Down syndrome and support for families and patients, but only in the context of very strong opposition to abortion.  They're also campaigning to have Lejeune beatified by the Vatican.

But Gautier's role in the discovery of trisomy 21 was not totally forgotten.  It has been very well described in a 2009 article in the journal Human Genetics by Gautier and Peter Harper, the author of a major history of cytogenetics (paywalled but try this link to a pdf), and in a 2013 interview.  There's also a French Wikipedia page about her.  But few people in the field know about this injustice, and cytogenetics textbooks and courses still credit Lejeune for the discovery.  Gautier has no English Wikipedia page, and the Wikipedia pages on Lejeune describe her contribution as follows:
"Using a new tissue culture technique brought back from the United States by his colleague Marthe Gautier, Lejeune began working with her to count the number of chromosomes in children with Down syndrome. The laboratory notebook begun by Dr. Lejeune on July 10, 1957 indicates that on May 22, 1958, he succeeded in showing, for the first time, the presence of 47 chromosomes in a child with Down syndrome."
Not surprisingly, the Fondation Jerome-Lejeune strongly opposes any correction of the scientific record, since this would reveal the intellectual theft at the base of their hero's reputation.  That's why they sent in the bailiffs to record her award ceremony.

I'm of course outraged to learn about this situation, and this post is one attempt to set the record straight.  My other venue is Wikipedia, which I've been learning to edit.  So far I've added sentences crediting Gautier with the discovery to the Wikipedia entries on Down Syndrome and on Jerome Lejeune.  Someone else had added a mention of the dispute to his French entry.  I'm going to expand these each into a paragraph.  I've also created an empty page for Marthe Gautier and requested that the Wikipedia translation people fill it from her French entry.

So please spread the word.  Marthe Gautier discovered that trisomy 21 is the cause of Down syndrome, and Jerome Lejeune's saintly reputation is based on scientific fraud.

Later:  I've corrected an error: Gautier could not identify the trisomic chromosome with her poor microscope.  And here are links to news articles about this controversy, in Nature and Science.

Plans for RNA-seq analyses

I should have posted this after last week's lab meeting but am only now getting to it.  I sensibly took snapshots of the whiteboard at the end of that lab meeting, so I could check what we'd decided.

The issues:  We have several Haemophilus influenzae mutants whose gene-expression profiles we want to examine, either during competence development in the MIV starvation medium or during normal growth in the rich culture medium sBHI.  For most of these (i) we want samples from several timepoints over a few hours, (ii)we want wildtype controls done in the same experiment, and (iii) we want three replicate samples from experiments done on different days.  And it would be nice to have multiples of 24 samples, since the kits and sequencing are most efficient with that.

That's a lot of constraints , but we came up with a plan that meets them all:  The first two days are the samples that have already been prepared and sequenced; the other 6 days are for me to generate the samples.

The samples will consist of viable cells frozen in glycerol (one or two 1.5 ml tubes), and duplicate pellets of cells that have been briefly incubated with a RNA-protection reagent to stabilize their RNA.  Later the frozen viable cells will be thawed and transformed to check that they have the expected level of competence - I can probably do all of them in a few days.

After all the samples have been collected, the RNA-prep pellets will be thawed and the RNA isolated using a kit.  After a quick check of RNA concentration and size, the contaminating DNA is removed by treatment with Turbo-DNase.

All the samples are then checked for concentration and quality using a special something (the post-doc recommends, using equipment in another lab), then treated to remove the bulk of the ribosomal RNA (this kit costs a lot, I think $200 per sample), then rechecked for concentration and quality using the special something.

Finally the samples are ready to be made into multiplexed sequencing libraries (expensive service on campus) and then sequenced (another service on campus).

To order (first check what we have on hand):
  •  RNA protection reagent for 72 samples
  • RNA prep kits for 72 samples
  • Turbo-DNase for 72 samples
  • Does the 'special something' require special reagents?
  • rRNA-removing kits for 72 samples
The reasons for doing these experiments are described briefly in this post.  

Because my hydroxyurea experiments found that cell doubling is substantially faster when cultures are at very low cell density, for the first samples in experiments F. G and H I'll use large volumes of cells at a lower cell density than the usual OD600=0.2.  Getting the right density will be a bit tricky since we can't use OD600 to check densities of very dilute cultures.  In the table above I said I'd use an OD600 of 0.02, but I'll start by growing cells from a lower density to about this OD and then diluting them 16-fold so they'll have another four doublings before I sample them.  The hydroxyurea-experiment cultures were much more dilute than this, but I need to balance culture density and culture volume so I'll have enough cells for the RNA extractions.  With this plan I'll be collecting cells from about 2 x 250 ml of culture for the first samples, and combining the remaining volumes in the two flasks for the later samples.

Other experiments to do:  We need to sequence the genome of strain RR753, to identify candidate mutations that cause its hypercompetence.  Before doing this I should recheck it and the backcross strain I made - if they both are hypercompetent we'll want to sequence both.