2011 Nobel Prize in Medicine: a tribute to endotoxin research!

J. Hoffmann, B. Beutler, R. Steinman

Finally I got the Nobel Prize!! Actually, innate immunity, my research topic, got the prize.  The innate immune system is the main weapon that keeps organisms without infections. A proof-of-concept example is the pioneering work carried out by Jules Hoffmann´s team showing how fruit flies combact infections. Their work led to the identification of the receptor, named Toll, used by the flies to detect pathogens. A similar receptor, called Toll-like receptor (TLR) 4, turned out to be expressed by our cells and its activation cause inflammation, an essential response to eliminate invading microorganisms. Bruce Beutler´s studies using knock-out mice made an elegant clear-cut case. However, we should not forget that the work and insights of Charles Janeway were the driving force for this line of research. World-class scientists continuing his fundamental work are Ruslan Medzhitov, Shizou Akira, Douglas Golenbock, Terje Espevik, Luke O´Neill,...just to mention a few. However something is missing...A dogma in Biology is the receptor-ligand notion. In other words, there is always a ligand, at least one, for a given receptor. Let´s put in Nobel Prize context decades of outstanding research on the TLR4 ligand: the lipopolysaccharide.

Lipopolysaccharide (LPS) is the major component of the surface of Gram-negative bacteria. Except for few bacterial examples, LPS is essential to gram-negative bacteria, whose death results if it is mutated or removed. LPS is the prototypical endotoxin because it binds the so-called TLR4 LPS receptor complex hence leading to the secretion of pro-inflammatory mediators and compounds with microbicidal actions. The medical syndrome caused by a response out of balance is called endotoxic shock or septic shock with a mortality rate as high as 50%. 

Decades of outstanding research.
The initial emphasis on LPS research was placed on elucidating chemical structures of LPS with the aim of defining the sections of the molecule responsible for the biological activities. Essential were the methods to purify LPS described by Otto Westphal, Otto Lüderitz and Chris Galanos published during the 60s. Mostly scientists working at the former USSR and Germany elucidated the first chemical structures. In this field, outstanding work has been carried out by Yuri Knirel, Eugeny Vinogradov, Kazyyoshi Kawahara, Hubert Mayer, Ann-Marie Staub, Ernst Rietchel, Otto Holst, Antonio Molinaro...

Elegant structure-function studies were carried out by Ulrich Seydel and Klaus Brandenburg to demonstrate the connection between the LPS supramolecular structure, the so-called entodotoxin conformation, and the bioactivity of the molecule. Whereas Martti Vaara, Hiroshi Nikaido and Robert Hancock pubished seminal studies on the contribution of LPS to the stability of the bacterial membrane.

The advent of bacterial genetics brought into the LPS field the first mutants lacking only certain parts of the molecule. Further, the first studies were published identifying the gene clusters necesary for LPS biosynthesis. Carl Schnaitman, John Klena, Peter Reeves, Joseph Lam, Chris Withfield, Mikael Skurnik, Miguel Valvano, and Richard Moxon should receive credit for their otstanding contributions. This approach, combining genetics and biochemistry, has led Joanna Goldberg, Juan Tomás, and Derek Hood to elucidate the make-up necesary for LPS biosynthesis in important human pathogens.

Without any doubts, Chris Raetz , who recently passed away, is one of the most influential scientist on the LPS field. Chris identified the pathway necesary for the biosynthesis of lipid A, discovered the gene for each enzyme in the pathway, and purified and crystallized the proteins all the while seeking ligands with pharmacological potential. His pioneering studies on Salmonella and E. coli lipid As have paved the way to similar studies on other pathogens such as Vibrio, Pseudomonas, Helicobacter, Bordetella, Acinetobacter, Klebsiella, Brucella carried out by Stephen Trent, Emile Caroff, Ilka Helander, Samuel Miller, and my group among many others. 

Future directions.  

What´s next? I envision that next years will bring the identification of new enzymes and/or activites important for LPS biosynthesis in other bacteria than Enterobactericeae. Whole-genome sequencing will certainly facilitate these analysis. Evidently, these studies may have important biotechnological applications which have not been really pursued. The tools are in placed to design and produce chimeric LPSs which maybe used as novel immunadjuvants or as treatments for septic shock. 

The contribution of LPS to virulence is still far from clear. I expect exciting studies on this topic specially on the field of plant pathogens. The means used by bacteria to regulate LPS expression and how LPS expression is coordinated with that of other virulence factors requires atention. However, in my view, perhaps the most exciting studies will address the LPS expression and structure in vivo at the single bacterial level. It is possible that the LPS structure may even change as a result of the interaction with the host immune system.

After more than 70 years of LPS research, we know a great deal about this exciting molecule. However, there are still many opened questions that should keep us working for a long time. Unfortunately, nowadays LPS is not an attractive topic for young scientists and might be even difficult to get a faculty position with a research programm based on LPS. I hope that I will not be the last one to switch the light off.

* This post is a posthumous tribute to Chris Raetz, a great scientist who had a profound effect on my career and the field of lipid biochemistry.