Expression variation of TLR as candidate genes underlying trypanotolerance QTL in mice

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Abstracts / Veterinary Immunology and Immunopathology 128 (2009) 211–347

precursor. Our results demonstrate the power of RNAi coupled with microarray analysis for dissecting the genetic and cellular processes leading to cell transformation by retroviruses. doi:10.1016/j.vetimm.2008.10.015 Monocytes from disease resistant and susceptible cattle display distinct transcriptome profiles during Theileria annulata infection K. Jensen 1 , R. Talbot 2 , D. Waddington 1 , E.J. Glass 1,∗

Comparative genome analysis of trypanotolerance QTL Joseph Nganga 1,2,∗ , Mabel Imbuga 2 , Fuad A. Iraqi 3 1

International Livestock Research Institute, P.O. Box 30709, Nairobi, Kenya 2 Jomo Kenyatta University of Agriculture and Technology, P.O. Box 62000, Nairobi, Kenya 3 Department of Human Microbiology, Sackler Faculty of Medicine, Tel Aviv University, Ramat Aviv, Tel Aviv 69978, Israel Keywords: Trypanosomosis; Quantitative trait loci; Homology; Single nucleotide polymorphism

1

Roslin Institute, Roslin, Midlothian EH25 9PS, UK ARK-Genomics, Roslin Institute, Roslin, Midlothian EH25 9PS, UK Keywords: Macrophages; Disease resistance; Protozoan; Transcriptome 2

Species: Ruminants The tick-borne protozoan parasite Theileria annulata causes a debilitating and often fatal disease of cattle called tropical theileriosis. The disease has a global economic impact on livestock production as it is endemic in many areas of the world from the Mediterranean basin to China. Control strategies have so far failed to eradicate T. annulata or its vector. An attractive alternative control strategy is to breed for combined resistance and productivity in cattle by using pre-existing genetic resistance. We have identified a Bos indicus breed of cattle that originates from Pakistan, the Sahiwal, which is resistant to tropical theileriosis. Our current studies aim to identify the genes underlying the resistance of Sahiwal cattle to T. annulata.T. annulata principally infects bovine macrophages and the pathology of the disease is associated with the intramacrophage stage of the parasite. Therefore we have focussed our studies on macrophages and monocytes derived from resistant (Sahiwal) and susceptible (Holstein-Friesian, B. taurus) breeds of cattle. We have undertaken a global analysis of the transcriptional response of Sahiwal and Holstein-Friesian derived monocytes to T. annulata infection using a bovine macrophage specific microarray developed in our laboratory. This approach has identified over 60 expressed genes that exhibit breed-specific differential expression either in resting monocytes or during T. annulata infection, which may be related to T. annulata resistance. Many of the differentially expressed genes are cell-surface expressed proteins, e.g. prion protein and ICAM1, which are involved in the interaction of macrophages with other immune cells. Further analysis of the pathways leading to variation in expression of these molecules, may reveal the underlying causal genes for resistance and susceptibility to T. annulata, and provide new approaches for disease control.

Species: Ruminants Certain breeds of domestic ruminants show remarkable resistance to the effects of African trypano-somosis. Unlike susceptible animals, trypanotolerant animals control parasitemia and do not show severe anaemia or production loss. Identification of trypanotolerance genes in cattle is hampered by cost and breeding time. Marked differences between inbred strains of mice in their response to T. congolense infection can be exploited in the analysis of the genetic basis of the infection. Murine trypanotolerance QTLs have been identified on chromosome 17, 5 and 1, and designated as Tir1, 2 and 3, respectively. Tir1 and 2 have been fine mapped to a confidence interval of 1 cM. In order to find the mouse homologous region on the bovine genome, nucleotide sequence across 95% CI of Tir2 and 3 were used in the selection of candidate genes. Homologous sequences were used in the definition of synteny relationships and subsequent identification of the shared disease response genes. The homologous genes within the human genome were then identified and aligned to the bovine radiation hybrid map in order to identify the mouse/bovine homologous regions. This revealed homology between murine and bovine QTL on Tir3 while the region on Tir2 is linked to innate immune response. doi:10.1016/j.vetimm.2008.10.017 Expression variation of TLR as candidate genes underlying trypanotolerance QTL in mice Joseph Nganga 1,2,∗ , Fuad A. Iraqi 3 1

International Livestock Research Institute, P.O. Box 30709, Nairobi, Kenya 2 Jomo Kenyatta University of Agriculture and Technology, P.O. Box 62000, Nairobi, Kenya 3 Department of Human Microbiology, Sackler Faculty of Medicine, Tel Aviv University, Ramat Aviv, Tel Aviv 69978, Israel Keywords: QTL; Expression variation; Toll like receptors; Interleukins

doi:10.1016/j.vetimm.2008.10.016 Specie: Other (mice) After QTL mapping and physical representation of the particular chromosomal fragment spanning trypanosomosis resistance loci Tir2 and 3 in mice, possible candidate genes were selected. These appeared to be linked to the innate immune response. Plausible trypanotolerance can-

Abstracts / Veterinary Immunology and Immunopathology 128 (2009) 211–347

didate genes within the loci include TLR1, 5 and 6. TLRs are critical in the regulation of pro-inflammatory cytokine secretions. In an effort to find an association between the genes and the disease, expression patterns of TLR genes mapping to trypanotolerance QTL were investigated using quantitative real time PCR. Susceptible and resistant mice infected with T. congolense portrayed diverse TLR expression patterns. Up regulation of different TLR seemed to coincide with up regulation of IL-10 and TNF in susceptible and resistant strains, respectively. T. congolense infection therefore induces a response characterized by changes in TLR1, 5 and 6 expression in liver and spleen tissues which appear to regulate cytokine profiles in mice. These phenomena may be responsible for the diverse disease pathology evident in different mouse strains and may account similar trends observed in livestock breeds. doi:10.1016/j.vetimm.2008.10.018 Immunoglobulin allotype-defined pedigreed rabbits and genes of immunological interest in 2× rabbit genome assemblies (workshop)

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ing. When I typed serum samples from such animals in 1995, 6/12 were heterozygous for the VH1a types a1 and a2 and for the linked C gamma hinge region d11/12 types. Two of 12 rabbits were also heterozygous for CK1 allotypes (b4/b5). A search for VH genes in the Trace Archive of Oryctolagus cuniculus (WGS) finds perfect matches at http://www.ncbi.nlm.nih.gov/blast/mmtrace.shtml, to previously published sequences of both VH1a1 and VH1a2. Selective maintenance of heterozygosity reported in wild rabbit populations, also occurred at the heavy chain locus during inbreeding. The genomic sequences of the hinge region and rabbit CK were not found although there are some VK sequences in the trace archive. Links at the Rabbit Genome Resources site http://www.ncbi.nlm. nih.gov/projects/genome/guide/rabbit/ to searches for genes in the assemblies of the 2× WGS sequence at Ensembl and UCSC (BLAT) are valuable for discovering predicted mRNA sequences and exon boundaries useful in designing primers for quantitative reverse transcriptase PCR. A full rabbit genome sequence will aid discovery of genetic contributions to animal and human disease susceptibilities. Supported by the Intramural Research Program of the NIH, NIAID.

Rose G. Mage Molecular Immunogenetics Section, Laboratory of Immunology, NIAID, NIH, Bethesda, MD 20892, USA Keywords: Pedigreed NIAID rabbits; Immunoglobulin allotypes; 2× to 7× coverage; Genome assemblies E-mail address: [email protected]. Species: Rabbit Although genomic sequence of the rabbit is currently “unfinished” at 2×, deeper 7× coverage is expected soon. The January 2006 document “Increasing sequence coverage from 2× to high coverage (6–7×) for selected mammalian Species,” that recommended rabbit be sequenced more deeply, describes the NIAID allotype-defined rabbits at p.14. These animals represent a valuable resource for future SNP discovery. They have polymorphisms of a variety of immune system genes including variants (allelic allotypes) of the VH, CH, and CL regions of antibody molecules. The colony also contains descendants of rabbits formerly at the Basel Institute for Immunology, including the mutant VH1a2-deleted Alicia, CK1 splicing defective Basilea, and several VH-CH recombinant heavy chain types. These rabbits are now available to interested individuals, particularly to sites where breeding colonies can be established. A relational database (computer program 4D) contains more than 45 years of breeding records and other information about animals in the colony. The whole genome shotgun (WGS) “unfinished Oryctolagus cuniculus database” of 2,076,044,328 letters in 719,158 sequences (ACCESSION AAGW00000000) has serious gaps, yet the information has already proven useful for immunological as well as in silico studies. At the Broad Institute, the DNA of the “Thorbecke Inbred Rabbit” chosen for sequencing was found by preliminary sequencing to have less heterozygosity than outbred NZW from the same company (Covance). Ancestors of this strain accepted skin grafts after 20 generations of inbreed-

doi:10.1016/j.vetimm.2008.10.019 Genomic organization and expression of T cell receptors (TCR) in the South American opossum Zuly E. Parra ∗ , Michelle L. Baker, Jonathan Trujillo, April Lopez, Alana Sharp, Jennifer Hathaway, Robert D. Miller Department of Biology, University of New Mexico, Albuquerque, NM 87131, USA Keywords: T cell; TCR; Opossum; Marsupial Species: Others T cells play an important role in regulation and effector functions in the adaptive immune response of all jawed vertebrates. There are two subsets of T cells according to their antigen receptors: ␣␤T cells and ␥␦T cells. Here we present an analysis of the complete genomic content and organization of the TCR loci in a model marsupial Species. The South American opossum (Monodelphis domestica) is used as model for both UV induced melanoma and is reservoir for the parasite that causes Chagas disease in humans. Better understanding of opossum T cell diversity would improve their utility for such research. We found that the organization and complexity of the opossum TCR␣ and ␦ loci are similar to that of human, with TCR␦ nested within TCR␣. Similarly, the opossum TCR␤ locus resembles that of humans, but with two additional clusters of gene segments. The opossum TCR␥ locus has a translocon organization, different from that of human where it is clustered. As we reported recently, opossums have a fifth TCR (TCR␮) which does not have a homologue in placental mammals. TCR␮ occupies a distinct locus from those encoding the conventional TCRs and it has a cluster rather than the usual translocon organization. Opossums like all marsupials give birth to relatively immature young so we investigated the timing of TCR expression in ontogeny. We found that some