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The 2011 Nobel Prize in Physiology/Medicine to Ralph Steinmann, Jules Hoffmann, and Bruce Beutler recognized a paradigm shift in our understanding of innate immunity, and its impact on adaptive immunity. The Prize highlighted the initial discoveries of Toll's role in immunity in flies, Toll-like receptors in mammals, and the establishment of dendritic cells as the initiators of adaptive immunity. This historical Commentary focuses on the developments in our understanding of innate immunity.
In 1908, the Nobel Prize in Physiology/Medicine went jointly to Ilya Ilyrich Metchnikoff, the original champion of cellular immunity, and Paul Ehrlich, then ambassador of humoral defenses, “in recognition of their work in immunity.” Metchnikoff advocated the idea that phagocytic cells, far from being harmful to the organism, as was the current paradigm, in fact constituted a first line of defense by nonspecifically ingesting and digesting invading pathogens and other foreign material []. His cellular theory of immunity, however, was challenged when Emil von Behring and Shibasaburo Kitasato discovered that immunity to tetanus and diphtheria was explained by antibodies (Abs) specific for their respective exotoxins []. Subsequently, Ehrlich proposed the “side-chain theory” to explain how Abs functioned []. However, the discovery by Almoth Wright and Stewart Douglas that “the body fluids modify bacteria in a manner which renders them ready prey to phagocytes” (where body fluids can now be interpreted as Abs in immunized animals) was the first report that both branches (cellular and humoral) of the immune system may work together []. Wright named this observation the “opsonic phenomenon,” and the factors were called opsonins (from the Greek opsono (I prepare victuals for)). Even Ehrlich, an enthusiastic believer in humoral immunity, acknowledged in his landmark review of 1908 [] that infections are cleared by cellular and humoral immunity. Nevertheless, most immunologists at that time became followers of the humoral theory to explain how immune defenses worked, mainly because Abs could be easily studied in a test tube. Therefore—and perhaps mirroring the work of the more chemically oriented Ehrlich—immunology began to shift from cellular immunology toward chemistry, led by scientists such as Karl Landsteiner, Felix Haurowitz, Michael Heidelberger, John Marrack, and Linus Pauling.
In the early 1960s, the tide changed again and immunology transformed from a chemical to a more biological discipline mainly through the work of N. Avrion Mitchison [] and Peter Medawar [] who showed that cellular rather than humoral mechanisms were sufficient to account for allograft rejection, immunological tolerance, and resistance and memory against tumors. To begin to provide a framework to understand these phenomena, the Clonal Selection Theory was formulated [] and crucial discoveries were made such as the role of the thymus and bone marrow in T- and B-cell genesis (reviewed in [[9, 10]]), as well as the MHC restriction of antigen recognition by T cells []. These studies, however, largely neglected the contribution of innate immunity during the early phases of infection, perhaps because, until recently, the necessary conceptual views and technologies were missing. Of upmost importance to the development of the field has been the infusion of molecular biology into immunology and the utilization of the central dogma of genetics, which holds that cellular information flows from DNA to RNA to protein. As a result, today's understanding of immunology merges humoral and cellular aspects, and knowledge on adaptive immune responses has advanced by quantum leaps during past decades.
The Clonal Selection Theory [] states that each lymphocyte is equipped with many identical copies of an antigen-specific receptor, and when this receptor binds its ligand with high avidity, T and B cells undergo clonal expansion and differentiation. However, for naive T cells to become activated and for adaptive immunity to be initiated, antigen must be presented by a specialized cell type called the dendritic cell (DC), as was first brought to our attention in 1973 by the Nobel Laureate Ralph Steinman, together with Zanvil Cohn []. Ralph Steinmann's contribution in transforming the “novel cell type of 1973” into one of the brightest stars of the immunology firmament has often been highlighted, for example [] and is therefore not a focus of this article.
The upregulation of costimulatory signals on DCs, induced by postulated pathogen-associated molecular patterns (PAMPs), was speculated by the late Charles Janeway [] in 1989 to play an essential role in alerting adaptive immunity []. In addition, although microbes had long been recognized as the cause of infectious diseases, and Metchnikoff's nonspecific phagocyte model as the first line of immune defense had been with us since the end of the 19th century, the fundamental question as to how the immune system perceives infection remained largely unknown. A clue came from the observation that the inbred mouse strains C3H/HeJ and C57BL/10ScCr resisted doses of lipopolysaccharide (LPS; endotoxin) that were lethal in other mice strains []. Was it possible that these inbred mice harbored a nonfunctional (mutated) receptor sensing LPS?
The critical tools provided by Christiane Nüsslein-Vollhard, Edward Lewis, and Eric Wieschaus (Nobel Prize Laureates in 1995) assisted in the revelation of how the mammalian host recognizes infection. These researchers isolated a set of master genes in Drosophila. Of note, Nüsslein Vollhard's group showed that the Toll gene controls the establishment of the dorsoventral axis in fruitfly embryos []. Using the Toll mutants generated for the embryological studies, in 1996 Jules Hoffmann and coworkers, including Bruno Lemaitre, discovered that a functioning Toll gene was essential to control fungal infections in adult flies []. In their landmark publication [], they described how the gene cassette “spätzle (Toll ligand)/Toll/cactus (the Drosophila NF-κB analogue)” controlled antifungal “defensin” production that in turn combated fungi. That the fly innate immune system relied upon germline-encoded and ligand-specific receptors to sense pathogens was a revelation to many immunologists, and advanced TLR biology at an incredible speed.
Charles Janeway and his collaborator Ruslan Medzhitov cloned a human (h) TLR (as recounted in []—and following on from Janeway's speculation on PAMPs and the adaptive immune system [] noted above) at the time that the Hoffmann group's results in flies were published []. One year later, Janeway and Medzhitov published data showing that enforced expression of a constitutive active hTLR (it happened to be TLR4) caused NF-κB-dependent cytokine production and induction of costimulatory molecules []. This discovery triggered a further explosion in the field of innate immunity, since it was the first to link TLRs with activation of innate immune cells resulting in the upregulation of costimulatory molecules.
In 1985, Bruce Beutler and colleagues reported that LPS—the major glycolipid constituent of the outer membrane of Gram-negative bacteria—induces the pro-inflammatory cytokine “tumor necrosis factor” []. Using LPS-resistant C3H/HeJ mice, Beutler's group searched for the postulated LPS receptor via a positional cloning approach. His group discovered in 1998 that TLR4 is required for LPS recognition: a missense mutation in the third exon of TLR4 ablated LPS recognition in C3H/HeJ mice []. Since LPS can induce lethal sepsis, Beutler's milestone discovery was the first to link the TLR system with recognition of structurally defined molecules of utmost biological relevance.
In generating TLR pathway gene knockout (KO) mice, Shizou Akira and his group were central in profoundly advancing our knowledge of TLR immunobiology. While an early study from this group confirmed that TLR4 recognizes LPS [], the group's ever expanding stock of KO mice allowed them (and many others) to identify the ligands of other TLR family members and to dissect the TLR-signaling pathways, yielding either the induction of pro-inflammatory cytokines or type 1 interferons (reviewed in []). Of note, Akira has been extraordinarily generous in sharing his KO mice with the scientific community, and deservedly he is one of the most highly cited biologists in the world.
In summary, the pioneering work of Akira, Beutler, Hoffmann, and Medzhitov—initially together with the late Charles Janeway—has brought about an overwhelming paradigm shift in how we view the immune system. Innate immune cells (and many other cells, as it turns out) express evolutionary-conserved, germline-encoded receptors that recognize pathogen-derived ligands, which function as powerful adjuvant both to induce innate immunity in all organisms and to alert adaptive immunity in vertebrates. Thus, the original question posed at the end of the 19th century regarding how the host perceives infection appears to have been solved.
While they were the first to be discovered, TLRs are not the only pattern-recognition receptors (PRRs), and subsequent work has uncovered a plethora of recognition molecules. TLRs and C-type lectin PRRs are membrane-bound, found at the cell surface and in endosomes. Many additional PRRs are found in the cytoplasm, including the “retinoic acid inducible gene I-like receptors,” “nucleotide binding domain leucine rich repeat containing receptors” (NLRs), and several other DNA sensors that signal through a crucial adaptor (STING, stimulator of IFN genes) associated with the ER membrane (reviewed in []). In fact, STING has recently been shown also to function as a direct sensor of cyclic di-GMP (a conserved signaling molecule restricted to bacteria) []. In addition, the pioneering work of the late Jürg Tschopp [] highlighted the caspase 1-activating function of the “inflammasome,” formed in the cytosol after ligand-driven oligomerisation of certain NLRs []. Once activated, caspase 1 controls maturation of members of the interleukin (IL)-1 family, and IL-1 is known to drive fever, a characteristic ofinflammation (reviewed in []).
Unforeseen, a second paradigm shift (the first being the identified link between innate and adaptive immunity) has appeared on the horizon in recent years. There is now compelling evidence that germline-encoded PRRs not only perceive pathogen-induced inflammation, but also “sterile (auto)inflammation” by sensing metabolically altered self-components (reviewed in [[30, 31]]), including modified lipids [] and proteins [].These data have supported Matzinger's view that “danger” as sensed by the innate immune system comes mainly “from the inside” []. Autoinflammatory responses have been linked, for example, to type 2 diabetes (see the clinically relevant effects of IL-1 blockers []) and to certain aspects of this metabolic syndrome []. Furthermore, chronic autoinflammation is considered as hallmark of age-associated arteriosclerosis [].
A third paradigm shift has arisen more recently. PRRs such as TLRs do not discriminate between commensals and pathogens in the gut microbiota. However, there is increasing evidence that TLR signaling in the intestinal epithelium shapes not only intestinal function (reviewed in []), but also the induction inflammatory Th17 T cells and that of regulatory T cells (reviewed in []). Thus, T-cell functions appear to be imprinted not only in the thymus but also in the gut.
On the morning of 3rd October 2011, we celebrated the announcement that Ralph Steinmann along with Bruce Beutler and Jules Hoffmann had been awarded the Nobel Prize for Physiology and Medicine. Soon after, however, we mourned that Ralph Steinmann had passed away 3 days prior to the announcement. Ralph Steinmann was awarded one half of the Nobel Prize “for his discovery of the DC and its role in adaptive immunity,” since he unraveled their professional antigen-presenting function that shapes adaptive immune reactivity and tolerance. Jules Hoffmann and Bruce Beutler shared the other half of this Nobel Prize for their discoveries on how Toll (in flies) and TLRs (in mammals) activate innate immunity. Here, I have discussed my view of innate immunity's path to the Nobel Prize, and pointed out the evolving paradigm shifts in how we have viewed immunity over the past century. Obviously, the Nobel Prize decision highlighted the biological importance of the initial discoveries, but these discoveries now impact tremendously on our understanding of age-related autoinflammatory diseases, intestinal function, and the putative interdependence of the gut's microbiota and adaptive immunity. We all look forward to this century's discoveries.