DNA Origamis to the Rescue in Lupus Research

DNA Origamis
A tantalizing question in lupus research has been how anti-dsDNA antibodies bind DNA, and whether there are physiological mimetopes by which B cells expressing these antibodies might be selected. Because no crystals could be generated, there is no available X-ray structure of anti-dsDNA antibodies or Fab fragments bound to DNA. With cryo-electron microscopy, we now have a tool comparable—and potentially superior—to traditional X-ray crystallography techniques. In this way, we compare the structure of antibodies that differ by a single or a few amino acids and are dsDNA-binding and nonbinding, respectively. However, to obtain structures of antibodies bound to DNA fragments, we have to constrain the flexibility of DNA—the part of it that is not bound to antibody. In vivo, DNA is constrained in complexes such as chromatin or retroviral pre-integration complex. Similarly, for our structural study we constrain the dsDNA in synthetic nucleosome or DNA origami structures.

During the last decade, DNA origami has progressed past an art form and is being tried for a number of applications. The specificity of the interactions between complementary base pairs makes DNA an exciting construction material for molecular origamis—creating scaffolds that hold other molecules in place or structures all on its own. For our research, we use two examples of antibody complexes with constrained dsDNA. In one example, synthetic DNA is folded into a 6-helix honeycomb structure, which forms a tube. In the other example, the antibodies are complexed with synthetic nucleosomes.

The figure shows electron micrographs (EM) of DNA-antibody complexes. In the upper panel, an antibody is bound to a 6-helical honeycomb tubular structure made of DNA. To the left of the panel is an inter­pretation of the first EM picture; antibody is in blue, DNA in grey. It appears that antibodies mostly bind to the ends of the origami structure, where the full DNA double helices are more accessible. In the lower panel, an anti­body is bound to a synthetic nucleosome. To the left, interpretation of the first EM picture. To the right, enlarged schematic of the nucleosome structure; DNA (gray) in a ring outside, and histones (blue) in the center. (In collaboration with Eugene Palovcak, Yifan Cheng, Lea Tröster, Maryam Shansab and Shawn Douglas).

Ecotropic virus in mice that are modifiers of autoimmunity

The fairly recent realization that endogenous elements play an important role in the immune system has revived a great interest in them. We now know that endogenous retroelements are adjuvants in the immune response to T cell-independent type 2 antigens; and retroelements are implicated in monogenic and polygenic (“spontaneous”) autoimmune disease. Read More »

Proof, Facts, and Alternative Medicine

Philosophers of science are a bit like Catholic priests as marriage counselors: They entertain ideas that are different from those of the practitioners. The foolhardy assumption that a hypothesis cannot be proved, but only disproved, is irrelevant to the workings of modern science, which is based on experiments. (In fact, strictly speaking, a hypothesis can neither be proven nor disproved). Experiments restrict the range of discussion on a subject. While it is axiomatic that, for a limited number of experiments there is an unlimited number of explanations, or hypotheses, the hypotheses have to be reasonable. When reasonable explanations are tested and excluded, the remaining explanation becomes the basis of the next experiment. It is this next experiment that theoreticians forget about: It is the accumulation of positive outcomes from successive experiments that establishes a fact, such as that DNA is the hereditary material. “Alternative” medicine is, at best, based on “experience,” not on experiments. While experiences no doubt constitute facts as well and, indeed, greatly outnumber facts established by experiments, experiences tend to be tied to various ad hoc explanations that fit the day. The therapy prescription to “throw enough cards (or remedies), and eventually some odds will go your way,” is questionable, as there are too many possibilities.

How inflammation leads to multiple myeloma

In mice, mineral oil causes inflammation, which leads to a lupus-like condition and to the formation of plasma cell tumors. It has long been known that chromosomal translocations (breakage of two chromosomes and rejoining the wrong ends between them) drive tumor formation and progression. But clearly, this is not enough for full-blown tumors, and it raises the question: What do inflammatory tissues provide to promote further mutagenesis? We have shown that endogenous retroviruses and retrovirus-like particles mutate DNA in such a way as to activate cancer genes.

From the peer review of the paper:

“This is a well written paper that progresses our understanding of the transforming events that may be associated with inflammation-induced cancer. Experiments described in this manuscript will fuel further investigations to determine whether human cancers associated with chronic inflammation can be associated with reactivation, replication and mobilization of endogenous retroelements, thereby possibly contributing to tumor initiation maintenance and/or progression.”

Read the paper:

It seems that lupus and HIV avoid each other. Why is that?

There is only anecdotal and retrospective evidence for it, but it seems that HIV and lupus avoid each other. One might think, no wonder: HIV patients are immune-deficient and thus may be autoimmune-deficient, too. However, there ought to be patients who suffered from lupus before they were infected with HIV. Clinician-scientists tell me that of the thousands of HIV patients they have seen, none had lupus. While this does not rise to the cogent standards of scientific evidence, it may be worth thinking about.

Mice have many endogenous retroviruses (viruses that are part of the mouse genome), and so do human beings. The endogenous retroviruses in people are not infectious, but they still can produce some proteins. These proteins may prevent completion of the infection cycle by HIV. In mice, the so-called Fv-1 resistance gene, which stymies infection by a mouse retrovirus, encodes a protein similar to that of the core of the exogenous virus. By analogy, lupus patients may express a retroviral protein that interferes with the life cycle of HIV.

“How to Understand a Friend” — almost a decade later

The following letter was written at the height of the intense discussion about intelligent design versus Darwinism. It was in response to an editorial by Cardinal Schönborn, the lead editor of the official 1992 Catechism of the Catholic Church, in the New York Times of July 7, 2005. It was not sent for personal reasons.

“How to Understand a Friend”

Dear Christoph,

Your article in the New York Times of last Thursday has had serious percussions in Austria, and it has been noticed elsewhere in the world as well. If it were possible to create a bridge between the catholic and the scientific world in this specific matter, I think it would have to be built on your interpretation of the word “design.”

In your rebuttal of the statements you cite in your opening paragraph, you say that “the Catholic Church, while leaving to science many details about the history of life on Earth, proclaims that by the light of reason the human intellect can readily and clearly discern purpose and design in the natural world, including the world of living things.”

As I have understood Darwin, he did give many details about the history of life, but his main point seemed to be that the structures we see can be understood in terms of their natural history. This history was driven by variation and “natural” selection (as opposed to “artificial” selection by breeders, for example). Darwin did not know about mutations; the knowledge about, and the understanding of these, are among the great triumphs of biology of the twentieth century.

Yes, there clearly is “purpose and design in the natural world, including the world of living things.” No biologist would argue with that! But now, how is this purpose and design accomplished? And here your understanding of what science ought to be departs from what today’s biologists think. Yes, indeed, as you have formulated so clearly, most biologists think that “purpose and design are achieved by an unguided, unplanned process of random variation and natural selection.” Read More »

Hybridoma history, as written by the journal Nature

If journalism is “the first rough draft of history” (Philip Graham), then we might be doomed, at least in a small way. An editorial referring to—and advertising—the Nature PastCast makes the point, “If those who do not learn from the past are truly doomed to repeat the same mistakes, then Nature has done its bit over the past few months to help them avoid that fate.” It goes on to talk about the hybridoma technique, published in 1975 by Köhler and Milstein in Nature. This technique enables the easy generation of monoclonal antibodies, an over-$50 billion-industry today. Curiously, the editorial informs us that:

“The revolution was nearly stillborn: the published technique to make monoclonal antibodies could not be reproduced. ‘The crisis was such,’ says historian Lara Marks at King’s College London, ‘that the authors considered withdrawing their article from Nature’. It was years before the investigations paid off and the huge medical potential of monoclonal antibodies started to crystallize. ‘Nowadays it seems prophetic,’ says Greg Winter, who made the first monoclonal antibody for use in humans, ‘but at that point I don’t think anyone had realized the importance.’ ”

If my recollection of the events serves me right, almost everything in this account is wrong. Read More »

The Bureaucratization of Science

“Science has always been bad” – Charles Steinberg

In November of 1895, when following up on a radiation experiment “that needed further investigation,” the fifty-year old Wilhelm Röntgen saw the bones of his fingers on a fluorescent screen. Worried about his sanity, he locked himself in his lab for two weeks to understand what was wrong with his worldview, or with himself. He finally emerged and, with trepidation, led his wife to his experimental setup, for her to confirm what he had seen. Only then did he tell the world about his discovery of X-rays.

Some twenty years earlier, Friedrich Miescher recovered a precipitate that contained phosphorus and nitrogen, but not sulfur, from cell nuclei. He called the substance “nuclein” (now known as DNA) and submitted a paper describing his finding for publication. The editor of the journal, Hoppe Seyler, found the discovery hard to believe, and only published the manuscript (ref. 1) after he repeated Miescher’s experiment and convinced himself of its validity.

These are but two examples of the many conscientious laborers in the garden of truth. Today, on the side of the experimental scientists, there are still many Röntgens among us, though clearly, it would be difficult for the modern editors to do as Hoppe-Seyler did. But what can they do, and what have they done, to ensure that the experiments and the conclusions of the manuscripts they publish are basically correct? In light of the recent, disputed paper by Obokata et al. describing stimulus-triggered acquisition of pluripotency, or STAP cells (ref. 2), and the ever-increasing number of rules, experiments, and requirements thrust upon authors to avoid such scandals, it is important to consider the role of the journal in curating science: Read More »

“The opposite of a fact is falsehood, but the opposite of one profound truth may very well be another profound truth.” — Niels Bohr

As scientists, we believe in cause and effect. If this is so, then everything is predetermined, at least at the macroscopic level. This would also mean that we are just molecular machines that have no responsibilities because we have no free will. In society, we would regard a person with such a radical view as a sociopath. We argue that we are indeed responsible for our actions and thus rightly punished for our crimes. How can these two views co-exist? Obviously, the human brain has evolved to observe, interpret and react to the outside world, but this does not mean that our soul, our thoughts, and our language are in perfect sync.

“From such crooked wood as that which man is made of, nothing straight can be fashioned.” — Immanuel Kant.

Clinical trial initiated, based on an experiment performed in the Wabl lab

In May 2014, a clinical trial of reverse-transcriptase inhibitors in Aicardi-Goutières syndrome (AGS) is scheduled to start at the Necker Hospital in Paris. The orphan disease AGS is a Mendelian inflammatory disorder of the brain and skin. It is a clinical and radiological mimic of congenital infection, and it is associated with increased levels of the antiviral cytokine interferon alpha. Sadly, thirty-five percent of children with this disease die before the age of 10. A subgroup of patients carry a mutation in a gene that encodes the enzyme Trex1. Mutations in this gene are also responsible for 2% of lupus cases. The investigators of the trial write that they were “inspired by the really fantastic paper [from our lab: http://www.retrovirology.com/content/8/1/91], … which indicates that the accumulation of cytosolic DNA in Trex1-null cells can be ameliorated by inhibiting endogenous retro-element cycling.” In that paper we showed that mice with this lethal enzyme defect can be rescued by FDA-approved drugs (retroelement inhibitors) and suggested that such treatment might also ameliorate AGS in humans.