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Resistance genes controlling infection

Molecular genetics of host cell responses to microbial pathogens: role of disease resistance/susceptibility loci. -

Aims: It is the aim of this short lecture course to review the current literature describing recent advances in our understanding of the immune response towards microbial pathogens brought about using molecular genetics. Study will be focused on two such genes responsible for the Lps and Ity/Bcg/Lsh phenotypes in animal models.

Objectives: Having attended both lectures and studied the cited literature it is expected you should be able to:

  1. Define the phenotypes for Lps and Ity/Bcg/Lsh gene models.
  2. Outline approaches for candidate gene identification and cloning for Lps and Ity/Bcg/Lsh gene models.
  3. Interpret models of Nramp1 gene function on the proliferative rates of microbial pathogens.

Background. Polymorphism within the genes of the host-cell immune system may render an individual as being more or less resistant to infection by microbial pathogens. In human the identification of such disease-predisposing loci is hampered due to differential exposure of the microbial pathogens in the study groups. Furthermore, pathogenic organisms of differing virulence may counter the effects of the host immune response and host polymorphism at other loci may influence the phenotypic indicator being used. Therefore to understand at the molecular level and elucidate the role of genes involved in the immune response, investigators have used animal models to identify candidate susceptibility loci. These two lectures will focus on two genes which operate within the innate immune response in mouse, and identified by positional cloning using models of infection,.

Lps gene. A series of defence mechanisms are operated following infection by Gram-negative bacteria by sensing the presence of lipopolysaccharide (LPS), a major component of the cell wall of the invading pathogen. In humans, monocytes and macrophages respond to LPS by inducing the expression of cytokines, cell-adhesion proteins, and enzymes involved in the production of small proinflammatory mediators. Under pathophysiological conditions, LPS exposure can lead to an often fatal syndrome known as septic shock. Responses of myeloid cells to LPS require a plasma protein called LPS-binding protein (LBP) and the glycosylphosphatidyl-inositol-anchored (GPI-) membrane protein CD14. However, the mechanism by which the LPS signal is transduced across the plasma membrane remained unknown until the recent cloning of the Lps gene and concurrent work on related proteins.

Prior to Lps identification Yang and co-workers showed that Toll-like receptor 2 (TLR2) is a signalling receptor that is activated by LPS in a response that depends on LPS-binding protein and is enhanced by CD14. A region in the intracellular domain of TLR2, with sequence identity to a portion of the interleukin (IL)-1 receptor that is implicated in the activation of the IL-1-receptor-associated kinase, is required for this response. The results indicate that TLR2 is a direct mediator of signalling by LPS.

Mutations of the gene Lps selectively block LPS signal transduction in two strains of mice (C3H/HeJ and C57BL/10ScCr) rendering them resistant to the effects of endotoxin, but susceptible to Gram-negative infection. The co-dominant Lpsd allele of C3H/HeJ mice was shown by Poltorak and co-workers to correspond to a missense mutation in the third exon of the Toll-like receptor-4 gene (Tlr4), a gene strongly related to TLR2 by sequence and by implication, function. The predicted changes substitute a proline for histidine at residue 712 of the polypeptide chain. C57BL/10ScCr mice are homozygous for a null mutation within Tlr4.

Toll was initially identified as a instrumental protein in the development of the Drosphila body plan and later in life is involved in defensive responses to fungal infection. Toll is an integral membrane protein with an intracellular domain conserved with the Il-1R. Subsequently 5 human Toll-like receptors have been cloned by Rock.

References:

Poltorak, A. et al. (1998). Defective LPS signaling in C3H/HeJ and C57BL/ScCr mice: Mutations in Tlr4 gene. Science 282, 2085.

Yang, R-B. et al. (1998). Toll-like receptor -2 mediates lipopolysaccharide-induced cellular signalling. Nature 395, 284.

also see (1998). J.Exp.Med. 188, 11 2091-2097

Muzio, M. et al. (1998). The human toll signaling pathway: divergence of nuclear factor kappaB and JNK/SAPK activation upstream of tumor necrosis factor receptor- associated factor 6 (TRAF6). J.Exp.MEd 187, 2097-2101.

Gerard C. (1998). For whom the bell tolls. Nature 395, 217.

Rock, F.L. et al. (1998). A family of human receptors structurally related to Drosophila Toll. Proc.Natal.Acad.Sci. USA. 95, 588.

O'Brien A.D. et al. (1980). Genetic control of susceptibility to Salmonella typhimurium in mice: The role of the LPS gene. J.Immunol. 124, 20.

Ity/Bcg/Lsh gene.

Ity/Bcg/Lsh is the name given to a phenotype identified between inbred strains of mice associated with the differential proliferative rates of obligate intracellular macrophage pathogens including: Salmonella typhimurium (Ity); Leishmania donovani (Lsh); Mycobacterium bovis, intracellulare and lepraemurium (Bcg). The identified gene responsible for the phenotype was termed Nramp1 (Natural resistance-associated macrophage protein 1), the prototypic member of a small family of genes exhibiting strong sequence conservation from yeast to man. Nramp1 is polymorphic in mouse, a G169D substitution corresponds precisely to the in vivo phenotype; the G169 allele corresponds to low proliferative rates of endocytosed pathogens and is termed resistant, the D169 allele is associated with high prolierative rates and termed susceptible.

Nramp1 operates to suppress the proliferation of intracellular pathogens prior to the acquisition of adaptive immunity and is a major component of the innate immune response. Nramp1 expression is restricted to the macrophage cell in mouse and encodes a transporter molecule, but expressed within an intracellular vesicular membrane. A major advance in our understanding of the function of the Nramp1 protein at the biochemical level has come from studies of the sequence-related members of the gene family; Nramp2 and the SMF genes from yeast. Nramp2 was originally isolated by low-stringency molecular hybridisation, and as an orphan gene. Recent work has re-identified this gene, by positional candidate cloning, as the carrier of a mutation (G185R) within strains of mice (mk) and rat (Belgrade) exhibiting a phenotype of impaired intestinal iron transport and analogous to the human disease microcytic anaemia. Secondly, functional cloning identified DCT1 as the rat-related Nramp2 gene using an assay of iron uptake. By implication, from the strong sequence conservation, a related iron transport function for Nramp1 is predicted. There is a basis for a link between iron transport and disease resistance since iron with-holding has been a proposed antimicrobial mechanism and iron adminstration appears to suppress Nramp1 antimicrobial function. Models of Nramp1 function and relationship to pathogen proliferation will be discussed.

References.

Blackwell, J.M. (1996). Structure and function of the natural-resistance-associated macrophage protein (Nramp1), a candidate protein for infectious and autoimmune disease susceptibility. Mol.Med.Today. 2, 205.

Vidal, S.M., Malo. D., Vogan, K., Skamene, E. & Gros, P. (1993). Natural resistance to infection with intracellular parasites: isolation of a candidate for Bcg. Cell 73, 469.

Vidal, S., Tremblay, M., Govoni, G., Gauthier, S., Sebastini, G., Malo, D., Skamene, E., Olivier, M., Jothy, S. & Gros, P. (1995). The Ity/Lsh/Bcg locus: natural resistance to infection with intracellular parasites is abrogated by disruption of the Nramp1 gene. J.Exp.Med. 182, 655.

Gunshin, H., Mackenzie, B., Berger, U.V., Gunshin, Y., Romero, M.F., Boron, W.F., Nussberger, B., Gollan, J.L. & Hedlger, M.A. (1997). Cloning and characterization of a mammalian proton-coupled metal-ion transporter. Nature 388, 482

Fleming, M.D., Trenor C.C., Su, M.A., Foenzler D., Beier, D.R., Dietrich, W.F. & Andrews N.A. (1997). Microcytic anaemia mice have a mutation in Nramp2, a candidate transporter gene. Nat.Genet. 16, 383

Fleming, M.D., Romano, M.A., Su, M.A., Garrick, L.M., Garrick, M.D. & Andrews, N.C. (1998). Nramp2 is mutated in the anemic Belgrade (b) rat: evidence of a role for Nramp2 in endosomal iron transport. Proc.Natl.Acad.Sci.USA. 95, 1148

Gomes, M.S. & Appelberg, R. (1998). Evidence for a link between iron metabolism and Nramp1 gene function in innate resistance against Mycobacterium avium. Immunol. 95, 165-168.

Weinberg, E.D. (1992). Iron depletion: a defense against intracellular infection and neoplasia. Life. Sci. 50, 1289-1297.

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Dr. H. Barton 2006