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Genetic testing for TMEM154 mutations associated with lentivirus susceptibility in sheep.

Heaton MP, Kalbfleisch TS, Petrik DT, Simpson B, Kijas JW, Clawson ML, Chitko-McKown CG, Harhay GP, Leymaster KA, International Sheep Genomics Consorti - PLoS ONE (2013)

Bottom Line: Genetic variation in the ovine transmembrane protein 154 gene (TMEM154) has been previously associated with OPPV infection in U.S. sheep.Sheep with the ancestral TMEM154 haplotype encoding glutamate (E) at position 35, and either form of an N70I variant, were highly-susceptible compared to sheep homozygous for the K35 missense mutation.A matrix-assisted laser desorption/ionization-time-of flight mass spectrometry (MALDI-TOF MS) assay was developed to detect these and six previously reported missense and two deletion mutations in TMEM154.

View Article: PubMed Central - PubMed

Affiliation: U.S. Meat Animal Research Center (USMARC), Clay Center, Nebraska, USA. mike.heaton@ars.usda.gov

ABSTRACT
In sheep, small ruminant lentiviruses cause an incurable, progressive, lymphoproliferative disease that affects millions of animals worldwide. Known as ovine progressive pneumonia virus (OPPV) in the U.S., and Visna/Maedi virus (VMV) elsewhere, these viruses reduce an animal's health, productivity, and lifespan. Genetic variation in the ovine transmembrane protein 154 gene (TMEM154) has been previously associated with OPPV infection in U.S. sheep. Sheep with the ancestral TMEM154 haplotype encoding glutamate (E) at position 35, and either form of an N70I variant, were highly-susceptible compared to sheep homozygous for the K35 missense mutation. Our current overall aim was to characterize TMEM154 in sheep from around the world to develop an efficient genetic test for reduced susceptibility. The average frequency of TMEM154 E35 among 74 breeds was 0.51 and indicated that highly-susceptible alleles were present in most breeds around the world. Analysis of whole genome sequences from an international panel of 75 sheep revealed more than 1,300 previously unreported polymorphisms in a 62 kb region containing TMEM154 and confirmed that the most susceptible haplotypes were distributed worldwide. Novel missense mutations were discovered in the signal peptide (A13V) and the extracellular domains (E31Q, I74F, and I102T) of TMEM154. A matrix-assisted laser desorption/ionization-time-of flight mass spectrometry (MALDI-TOF MS) assay was developed to detect these and six previously reported missense and two deletion mutations in TMEM154. In blinded trials, the call rate for the eight most common coding polymorphisms was 99.4% for 499 sheep tested and 96.0% of the animals were assigned paired TMEM154 haplotypes (i.e., diplotypes). The widespread distribution of highly-susceptible TMEM154 alleles suggests that genetic testing and selection may improve the health and productivity of infected flocks.

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TMEM154 SNP maps and median-joining networks.Panel A, genomic map of TMEM154: orange arrows, 5′ and 3′untranslated regions of exons; blue arrows, exon coding regions; grey rectangles, introns or intergenic regions. Blue and red tick dots denote position and frequency of SNPs in an international panel of 75 sheep and a panel of 96 U.S. sheep [9], respectively. Panel B, high resolution map of TMEM154 regions targeted for PCR-amplification. PCR amplification primers are indicated with black arrowheads and listed in Table S3. Red lowercase letters above SNP positions are IUPAC/IUBMB ambiguity codes for nucleotides (r = a/g, y = c/t, m = a/c, k = g/t, s = c/g, w = a/t) [17] and indicate 12 sites affected by nonsynonymous substitutions. The red uppercase letters above SNP positions indicate the amino acid polymorphisms encoded at TMEM154 codons 4, 13, 14, 25, 31, 33, 35, 44, 70, 75, 82 and 102. Black lowercase letters below SNPs indicate nucleotide polymorphisms that resulted in synonymous substitutions. Panel C, the areas of circles for haplotypes 1 to 4 are proportional to the frequencies in the international panel of 75 ISGC sheep. The symbols are as follows: black circles, risk factors; white circle, non-risk factor; grey circles, risk factor status unknown; red circles, haplotypes known in U.S. sheep but not observed in the international panel of 75 ISGC sheep (risk factor status unknown); shaded square, TMEM154 haplotype predicted to have occurred but unobserved to date. Dashed grey line, haplotypes observed in wild sheep species but not domestic sheep. Haplotypes 13 and 14 were observed in one animal each and their location of origin is indicated.
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pone-0055490-g002: TMEM154 SNP maps and median-joining networks.Panel A, genomic map of TMEM154: orange arrows, 5′ and 3′untranslated regions of exons; blue arrows, exon coding regions; grey rectangles, introns or intergenic regions. Blue and red tick dots denote position and frequency of SNPs in an international panel of 75 sheep and a panel of 96 U.S. sheep [9], respectively. Panel B, high resolution map of TMEM154 regions targeted for PCR-amplification. PCR amplification primers are indicated with black arrowheads and listed in Table S3. Red lowercase letters above SNP positions are IUPAC/IUBMB ambiguity codes for nucleotides (r = a/g, y = c/t, m = a/c, k = g/t, s = c/g, w = a/t) [17] and indicate 12 sites affected by nonsynonymous substitutions. The red uppercase letters above SNP positions indicate the amino acid polymorphisms encoded at TMEM154 codons 4, 13, 14, 25, 31, 33, 35, 44, 70, 75, 82 and 102. Black lowercase letters below SNPs indicate nucleotide polymorphisms that resulted in synonymous substitutions. Panel C, the areas of circles for haplotypes 1 to 4 are proportional to the frequencies in the international panel of 75 ISGC sheep. The symbols are as follows: black circles, risk factors; white circle, non-risk factor; grey circles, risk factor status unknown; red circles, haplotypes known in U.S. sheep but not observed in the international panel of 75 ISGC sheep (risk factor status unknown); shaded square, TMEM154 haplotype predicted to have occurred but unobserved to date. Dashed grey line, haplotypes observed in wild sheep species but not domestic sheep. Haplotypes 13 and 14 were observed in one animal each and their location of origin is indicated.

Mentions: The fidelity of genetic testing is enhanced by knowing the position and frequency of polymorphisms in the populations to be tested. If not accounted for, nucleotide variation at neighboring sites may cause base-pair mismatching with oligonucleotides used in DNA testing and significantly decrease the genotyping accuracy in some populations. Moreover, characterizing nucleotide variation in many previously untested breeds allows discovery of TMEM154 missense mutations. For these reasons, the same set of 75 whole genome sequence data was also used to identify novel polymorphisms in the TMEM154 gene region. An analysis of nucleotide differences among the 75 animals revealed approximately 1500 variant sites in a 61,663 bp region containing TMEM154, of which, only 128 had been previously reported [9]. The five wild sheep accounted for 13% of the nucleotide differences observed, however, no heterozygous animals were observed in wild sheep and it was assumed these were mostly species-related nucleotide differences. The positions of these 1,500 SNPs and their minor allele frequencies (MAF) are shown in Figure 2A (blue dots). For comparison, frequency data from 96 rams from 10 U.S. sheep breeds are shown for 128 SNPs (red dots). In regions of TMEM154 where data are available from both panels of sheep, there was a trend towards more SNPs and higher MAFs in the international panel of 39 breeds compared to the U.S. sheep from 10 breeds.


Genetic testing for TMEM154 mutations associated with lentivirus susceptibility in sheep.

Heaton MP, Kalbfleisch TS, Petrik DT, Simpson B, Kijas JW, Clawson ML, Chitko-McKown CG, Harhay GP, Leymaster KA, International Sheep Genomics Consorti - PLoS ONE (2013)

TMEM154 SNP maps and median-joining networks.Panel A, genomic map of TMEM154: orange arrows, 5′ and 3′untranslated regions of exons; blue arrows, exon coding regions; grey rectangles, introns or intergenic regions. Blue and red tick dots denote position and frequency of SNPs in an international panel of 75 sheep and a panel of 96 U.S. sheep [9], respectively. Panel B, high resolution map of TMEM154 regions targeted for PCR-amplification. PCR amplification primers are indicated with black arrowheads and listed in Table S3. Red lowercase letters above SNP positions are IUPAC/IUBMB ambiguity codes for nucleotides (r = a/g, y = c/t, m = a/c, k = g/t, s = c/g, w = a/t) [17] and indicate 12 sites affected by nonsynonymous substitutions. The red uppercase letters above SNP positions indicate the amino acid polymorphisms encoded at TMEM154 codons 4, 13, 14, 25, 31, 33, 35, 44, 70, 75, 82 and 102. Black lowercase letters below SNPs indicate nucleotide polymorphisms that resulted in synonymous substitutions. Panel C, the areas of circles for haplotypes 1 to 4 are proportional to the frequencies in the international panel of 75 ISGC sheep. The symbols are as follows: black circles, risk factors; white circle, non-risk factor; grey circles, risk factor status unknown; red circles, haplotypes known in U.S. sheep but not observed in the international panel of 75 ISGC sheep (risk factor status unknown); shaded square, TMEM154 haplotype predicted to have occurred but unobserved to date. Dashed grey line, haplotypes observed in wild sheep species but not domestic sheep. Haplotypes 13 and 14 were observed in one animal each and their location of origin is indicated.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3569457&req=5

pone-0055490-g002: TMEM154 SNP maps and median-joining networks.Panel A, genomic map of TMEM154: orange arrows, 5′ and 3′untranslated regions of exons; blue arrows, exon coding regions; grey rectangles, introns or intergenic regions. Blue and red tick dots denote position and frequency of SNPs in an international panel of 75 sheep and a panel of 96 U.S. sheep [9], respectively. Panel B, high resolution map of TMEM154 regions targeted for PCR-amplification. PCR amplification primers are indicated with black arrowheads and listed in Table S3. Red lowercase letters above SNP positions are IUPAC/IUBMB ambiguity codes for nucleotides (r = a/g, y = c/t, m = a/c, k = g/t, s = c/g, w = a/t) [17] and indicate 12 sites affected by nonsynonymous substitutions. The red uppercase letters above SNP positions indicate the amino acid polymorphisms encoded at TMEM154 codons 4, 13, 14, 25, 31, 33, 35, 44, 70, 75, 82 and 102. Black lowercase letters below SNPs indicate nucleotide polymorphisms that resulted in synonymous substitutions. Panel C, the areas of circles for haplotypes 1 to 4 are proportional to the frequencies in the international panel of 75 ISGC sheep. The symbols are as follows: black circles, risk factors; white circle, non-risk factor; grey circles, risk factor status unknown; red circles, haplotypes known in U.S. sheep but not observed in the international panel of 75 ISGC sheep (risk factor status unknown); shaded square, TMEM154 haplotype predicted to have occurred but unobserved to date. Dashed grey line, haplotypes observed in wild sheep species but not domestic sheep. Haplotypes 13 and 14 were observed in one animal each and their location of origin is indicated.
Mentions: The fidelity of genetic testing is enhanced by knowing the position and frequency of polymorphisms in the populations to be tested. If not accounted for, nucleotide variation at neighboring sites may cause base-pair mismatching with oligonucleotides used in DNA testing and significantly decrease the genotyping accuracy in some populations. Moreover, characterizing nucleotide variation in many previously untested breeds allows discovery of TMEM154 missense mutations. For these reasons, the same set of 75 whole genome sequence data was also used to identify novel polymorphisms in the TMEM154 gene region. An analysis of nucleotide differences among the 75 animals revealed approximately 1500 variant sites in a 61,663 bp region containing TMEM154, of which, only 128 had been previously reported [9]. The five wild sheep accounted for 13% of the nucleotide differences observed, however, no heterozygous animals were observed in wild sheep and it was assumed these were mostly species-related nucleotide differences. The positions of these 1,500 SNPs and their minor allele frequencies (MAF) are shown in Figure 2A (blue dots). For comparison, frequency data from 96 rams from 10 U.S. sheep breeds are shown for 128 SNPs (red dots). In regions of TMEM154 where data are available from both panels of sheep, there was a trend towards more SNPs and higher MAFs in the international panel of 39 breeds compared to the U.S. sheep from 10 breeds.

Bottom Line: Genetic variation in the ovine transmembrane protein 154 gene (TMEM154) has been previously associated with OPPV infection in U.S. sheep.Sheep with the ancestral TMEM154 haplotype encoding glutamate (E) at position 35, and either form of an N70I variant, were highly-susceptible compared to sheep homozygous for the K35 missense mutation.A matrix-assisted laser desorption/ionization-time-of flight mass spectrometry (MALDI-TOF MS) assay was developed to detect these and six previously reported missense and two deletion mutations in TMEM154.

View Article: PubMed Central - PubMed

Affiliation: U.S. Meat Animal Research Center (USMARC), Clay Center, Nebraska, USA. mike.heaton@ars.usda.gov

ABSTRACT
In sheep, small ruminant lentiviruses cause an incurable, progressive, lymphoproliferative disease that affects millions of animals worldwide. Known as ovine progressive pneumonia virus (OPPV) in the U.S., and Visna/Maedi virus (VMV) elsewhere, these viruses reduce an animal's health, productivity, and lifespan. Genetic variation in the ovine transmembrane protein 154 gene (TMEM154) has been previously associated with OPPV infection in U.S. sheep. Sheep with the ancestral TMEM154 haplotype encoding glutamate (E) at position 35, and either form of an N70I variant, were highly-susceptible compared to sheep homozygous for the K35 missense mutation. Our current overall aim was to characterize TMEM154 in sheep from around the world to develop an efficient genetic test for reduced susceptibility. The average frequency of TMEM154 E35 among 74 breeds was 0.51 and indicated that highly-susceptible alleles were present in most breeds around the world. Analysis of whole genome sequences from an international panel of 75 sheep revealed more than 1,300 previously unreported polymorphisms in a 62 kb region containing TMEM154 and confirmed that the most susceptible haplotypes were distributed worldwide. Novel missense mutations were discovered in the signal peptide (A13V) and the extracellular domains (E31Q, I74F, and I102T) of TMEM154. A matrix-assisted laser desorption/ionization-time-of flight mass spectrometry (MALDI-TOF MS) assay was developed to detect these and six previously reported missense and two deletion mutations in TMEM154. In blinded trials, the call rate for the eight most common coding polymorphisms was 99.4% for 499 sheep tested and 96.0% of the animals were assigned paired TMEM154 haplotypes (i.e., diplotypes). The widespread distribution of highly-susceptible TMEM154 alleles suggests that genetic testing and selection may improve the health and productivity of infected flocks.

Show MeSH
Related in: MedlinePlus