He did so by identifying spontaneous mutations in the gene coding for mouse Toll-like receptor 4 (Tlr4) in two unrelated strains of LPS-refractory mice and proving they were responsible for that phenotype.

[2] Subsequently, and chiefly through the work of Shizuo Akira, other TLRs were shown to detect signature molecules of most infectious microbes, in each case triggering an innate immune response.

During these years, he spent much time hiking in the San Gabriel Mountains, and in regional national parks (Sequoia, Yosemite, Joshua Tree, and Grand Canyon), and was particularly fascinated by living things.

His introduction to experimental biology, acquired between the ages of 14 and 18, included work in the laboratory of his father, Ernest Beutler, then at the City of Hope Medical Center in Duarte, CA.

Beutler also worked in the City of Hope laboratory of Susumu Ohno, a geneticist known for his studies of evolution, genome structure, and sex differentiation in mammals.

[14] He further suggested that H-Y antigen, a minor histocompatibility protein encoded by a gene on the Y chromosome and absent in female mammals, was responsible for directing organogenesis of the indifferent gonad to form a testis.

While a college student at the University of California at San Diego, Beutler worked in the laboratory of Dan Lindsley, a Drosophila geneticist interested in spermatogenesis and spermiogenesis in the fruit fly.

Beutler’s focus on innate immunity began when he was a postdoctoral associate and later an assistant professor in the lab of Anthony Cerami at Rockefeller University (1983-1986).

With J.-M. Dayer, Beutler demonstrated that purified TNF could cause inflammation-associated responses in cultured human synoviocytes: secretion of collagenase and prostaglandin E2.

The human p75 receptor chimeric protein was later used extensively as the drug Etanercept in the treatment of rheumatoid arthritis, Crohn's disease, psoriasis, and other forms of inflammation.

Beutler reasoned that in finding the LPS receptor, insight might be gained into the first molecular events that transpire upon an encounter between the host and microbial invaders.

[39] Utilizing positional cloning in an effort that began in 1993 and lasted five years, Beutler, together with several postdoctoral associates including Alexander Poltorak, measured TNF production as a qualitative phenotypic endpoint of the LPS response.

Analyzing more than 2,000 meioses, they confined the LPS receptor-encoding gene to a region of the genome encompassing approximately 5.8 million base pairs of DNA.

[2][40][41] They also showed that while mouse TLR4 is activated by a tetra-acylated LPS-like molecule (lipid IVa), human TLR4 is not, recapitulating the species specificity for LPS partial structures.

[43] Jules Hoffmann and colleagues had earlier shown that the Drosophila Toll protein, originally known for its role in embryogenesis, was essential for the antimicrobial peptide response to fungal infection.

Aware of this work, Charles Janeway and Ruslan Medzhitov overexpressed a modified version of human TLR4 (which they called ‘h-Toll’) and found it capable of activating the transcription factor NF-κB in mammalian cells.

This suggested that other TLRs (of which ten are now known to exist in humans) might also act as sensors of infection in mammals,[48] each detecting other signature molecules made by microbes whether or not they were pathogens in the classical sense of the term.

[49][50][51][52][53][54][55][56] After completing the positional cloning of the Lps locus in 1998, Beutler continued to apply a forward genetic approach to the analysis of immunity in mammals.

[67] feeble was identified in a screen in which immunostimulatory DNA was administered to mice intravenously with measurement of the systemic type I interferon response.

[71] Other genetic screens in the Beutler laboratory were used to identify genes that mediate homeostatic adaptations of the intestinal epithelium following a cytotoxic insult;[72][73][74][75][76][77][78] prevent allergic responses,[79] diabetes,[80][81] or obesity;[82][83][84] support normal hematopoiesis;[85][86][87][88][89][90][91][92][93][94] and enable humoral and cellular immunity.

In the course of their work, Beutler and his colleagues also discovered genes required for biological processes such as normal iron absorption,[99] hearing,[100] pigmentation,[101][102] metabolism,[82][84][103][104][105] and embryonic development.

In a laboratory setting, it accelerates positional cloning approximately 200-fold, and permits ongoing measurement of genome saturation as mutagenesis progresses.

[116] In addition, machine learning software, trained on the outcome of many thousands of experiments in which putative causative mutations were re-created and re-assayed for phenotype, is used to assess data quality.

[110][120] AMM has also permitted high speed searches for mutations that suppress or augment disease phenotypes; for example, the development of autoimmune (Type 1) diabetes in mice of the NOD strain.

Beutler has collaborated with Dale L. Boger and his research group to identify synthetic small molecule agonists of mammalian TLRs, which may be used in combination with defined molecular antigens to precisely target and coordinate innate and adaptive immune responses.

Beutler and colleagues also showed, again using X-ray crystallography combined with biological assays, that endogenous sulfatides are capable of binding to the TLR4-MD2 complex, causing its activation.

[133] Ernest Beutler was a hematologist and medical geneticist famed for his studies of G-6-PD deficiency,[134] other hemolytic anemias,[135][136] iron metabolism,[137] glycolipid storage diseases,[138] and leukemias,[139][140] as well as his discovery of X chromosome inactivation.