Genetic Mapping of Innate Immune Defects.

Septic shock is a very dangerous medical condition that isdescribed in your textbook on page 135. It is essentially an acutehyper-activation of the macrophage innate immune response,resulting in systemic (instead of localized) production ofcytokines, with deleterious effects. Normally, local cytokineproduction at an infection causes “leakiness” in blood vessels, sothat leukocytes can exit and migrate to the infection site(erythrocytes also leak out in the process, leading to the“redness” characteristic of inflammation). However, when cytokinesare released throughout the body (“systemically”), then all theblood vessels become leaky (“vascular permeability”), leading toprecipitous drops in blood pressure, and frequently organ failureand death. Septic shock has a 50% mortality rate, and is theleading cause of death in intensive care units (ICU’s) inhospitals.

One of the most common triggers of septic shock is LPS. Innormal Gram-negative infections, this outer-membrane molecule isonly present at the site of infection; however, if the bacteriareach the blood-stream, then LPS can quickly disseminate andtrigger massive cytokine release throughout the body. Thisdangerous property of LPS led to its original name, “endotoxin”.(“endo” was meant to distinguish it from secreted “exo” bacterialtoxins; unlike exotoxin virulence factors like cholera toxin,LPS/endotoxin is not a virulence factor evolved to exploit hostcells).

Mice were often used to study septic shock, and a mouse strainthat was resistant to septic shock was discovered in the 1960s.Normally, mice injected with LPS would succumb quickly to septicshock and death, but these mutant mice (C3H/HeJ strain) showed noeffect upon LPS injection. On the other hand, when infected withlive Gram-negative bacteria, the C3H/HeJ mice were unusuallysusceptible to infection and death.

In some ways, the C3H/HeJ mice were analogous to the boysdiagnosed with CGD (chronic granulomatous disease) in the 1960s, inthat both had heritable conditions that impaired immunity. Aheritable defect implies the existence of a mutated gene thatnormally contributes to the affected process.

2a. The typicalprocess for identifying an unknown gene that has a “phenotype” ofinterest is to “map” it to a particular chromosome, and then narrowits location to a smaller and smaller portion of this chromosome.Using animal models, this involves many generations, and tests ofeach generation to see which regions of which chromosome are alwaysinherited in the individuals that exhibit the trait. This impliesthat chromosomes are not inherited intact from generation togeneration: if chromosomes are strands of nucleotides covalentlylinked to each other, how is it that they are not inherited intactfrom parent to child?

2b. When mapping atrait in humans, instead of following inheritance through multiplegenerations (even if the disease status of ancestors is recorded,mapping is not feasible within family trees since DNA samples ofancestors were not preserved), researchers instead work withmultiple independent families that seem to exhibit the same geneticdisorder. For CGD genetic mapping, it was fortuitous that in the1960s the disorder was identified in multiple unrelated patients.What feature of the CGD disease made identifying the affected genea little easier by allowing the researchers to narrow their focusto a single chromosome?

2c. Is CGD arecessive or dominant genetic disorder?

2d. What traits ordefects were observed in patients with CGD?

2e. Identifying thegene affected in CGD patients led to the discovery of an enzymecomplex important in innate immunity. What is the name of thisenzyme complex, and what does it do?

2f. The C3H/HeJ micehad a genetic syndrome with both beneficial and detrimental traits.In your own words, describe the beneficial vs detrimentalaspects.