Forward genetic analysis of Mammalian Resistance to Viral Infection

Scripps Research Institute, The, La Jolla CA

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Abstract

Using an unbiased approach (random germline mutagenesis in mice), we have set out to identify host genes[unreadable] that are indispensable for survival early in the course of infection with a large DMA virus (mouse[unreadable] cytomegalovirus; MCMV). In selecting lethality as the endpoint of our screen, we have assured that all[unreadable] targets identified will be essential for defense, without regard to the level at which they function.[unreadable] Approximately 1 in 33 mice born to an ENU-mutagenized sire carries a recessive mutation that is lethal in[unreadable] the context of MCMV infection, suggesting that a large number of genes serve non-redundant functions in[unreadable] host resistance to this pathogen. By screening 12,170 G3 germline mutant mice from 2,028 micropedigrees,[unreadable] we have so far resolved a total of 31 transmissible mutations that markedly impair the innate immune[unreadable] response to MCMV. 8 additional mutations that impair MCMV resistance were identified by cross-screening[unreadable] mice identified in other innate immunity screens, so that 39 mutations have been collected in all, and at the[unreadable] present rate of screening, 25 new mutations should accrue each year. A total of 9 mutations have been[unreadable] identified already, and we have begun to map, assess allelism, and positionally clone those that remain.[unreadable] Informed by parallel efforts in Drosophila (Project 2) and aided by anticipatory resequencing,[unreadable] candidate analysis and positional cloning will be greatly accelerated. Paralogues of mutations that cause[unreadable] MCMV susceptibility will be targeted in mice (Project 3), and using both genetic and biochemical[unreadable] approaches, the mechanisms and pathways utilized in host resistance will be deciphered. Based on[unreadable] comparative estimates of saturation mutagenesis in which lethality is taken as an endpoint, we believe that[unreadable] approximately 11% of all host resistance genes have been struck in our primary screen as performed to[unreadable] date. As a first approximation, offer that about 280 genes comprise the MCMV resistome. The proteins that[unreadable] we identify may fulfill sensing or post-sensing functions. Some may offer very specific protection against[unreadable] MCMV; others may offer protection against many microbes, viral and non-viral alike. Some may fulfill[unreadable] evolutionarily conserved functions (in Drosophila and in mice) while some may be species-specific. We[unreadable] propose to continue the screen, which is the first of its kind in mammalian genetics, and in terms of hit rate,[unreadable] one of the most productive ever described in any model organism. Our goal is to elucidate a large fraction of[unreadable] the proteins that create resistance to viral infection in the mammalian host, and ultimately, to understand[unreadable] how these proteins work.