Genetics of resistance to bacterial and parasitic infection / edited by D. Wakelin and J.M. Blackwell.

Date:
1988
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Licence: Attribution-NonCommercial-NoDerivatives 4.0 International (CC BY-NC-ND 4.0)

Credit: Genetics of resistance to bacterial and parasitic infection / edited by D. Wakelin and J.M. Blackwell. Source: Wellcome Collection.

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    Conclusions 271 chromosome 8). The gene, which segregates for dominant resistant {Mx^) and recessive susceptible {Mx~) alleles, was later mapped to mouse chromosome 16 (Hallerei д/. 1988). The approaches used to identify the mechanism of action of the Mx gene initially followed the same theme as many of the genetic studies described within this book. Immunological correlates of resistance and susceptibility were sought by studying immunocompromised mice or mice bearing genetic immune deficiencies. Hence it was determined that T cell deficiency in nude {nu/nu) mice did not alter resistance (Haller and Lindenmann 1974), nor did expression of the xid X-linked recessive B-cell defect (Haller 1981). Resistance was also unimpaired by administration of cyclophosphamide (Haller et al. 1976) or whole-body X-irradiation (Haller 1981). The latter implied a role for the radioresistant resident macrophage population in mediating resistance, supporting the earlier observations (Lindenmann et al. 1978) of expression of resistance in peritoneal macrophages in vitro. Strangely, however, resistance and susceptibility were determined by recipient rather than donor genotypes in radiation bone marrow chimaeras (Haller et al. 1979a). Hence it was the environment of the macrophage which determined its response. An explanation for these observations was provided by further studies (Haller et al. 1979b, 1980) demonstrating that the Mx gene is regulated by IFN-a/ ß (but not IFN-7), anti- IFN antibodies rendering resistant mice susceptible in vivo. Treatment of macrophages in vitro with anti-IFN antibodies did not alter the resistant phenotype, but treatment in vivo followed by treatment in vitro (Haller et al. 1979b), or prolonged cultivation of resistant macrophages in vitro in the absence of IFN-a/ß (Haller et al. 1980), did result in a susceptible phenotype. The demonstration that expression of the Mx gene was induced by IFN-a/ß provided the breakthrough which allowed identification of the Mx gene product. It is at this point that the Mx story overtakes the rest of the field in molecular characterization of host resistance genes. Using 2-D PAGE analysis of [^^S] ^ methionine-labelled proteins from macrophages cultured in the presence or absence of IFN, Horisberger and coworkers (1983) identified the Mx gene product as a 72-5 kD protein present in IFN-treated macrophages from congenie BALB.A2G-Afx {Mx'^) mice but not from IFN-treated cells from BALB/c {Mx~) mice. Cross-immunization of BALB/c mice with extracts of IFN-treated cells from congenie BALB.A2G-A/x mice (Staeheli et al. 1985) produced a polyclonal hyperimmune serum and a monoclonal antibody, both of which immunoprecipitated a single IFN-induced 75 kD protein in Mx^ but not Mx~ cells. The protein was purified to homogeneity using the polyclonal antiserum (Horisberger and Hochkeppel 1985). The polyclonal antiserum was also used by Staeheli and coworkers (1986) to identify the Mx protein in a cell-free protein synthesis system containing RNA of around 3 kbp extracted and size-fractionated from IFN-treated Mx~^ embryo cells. A poly(A)'RNA fraction enriched 10- to 20-fold with respect to Mx mRNA activity was used to direct vector-primed cDNA synthesis using the cloning vector pHG327. The resulting cDNA library was then screened by differential colony hybridization using 32 P-labelled cDNA prepared from size-