Role of Genetic PolymorphismsRole of Genetic Polymorphismsi R t T i A ti R t T i A tin Responses to Toxic Agentsin Responses to Toxic Agents• Definitions• “Forward genetics” and toxicologyggy• “Reverse genetics” and toxicology• Genetic markers• SNPs and their use in toxicology•Ethical, Legal and Social Issues (ELSI),g ( )“Toxicology is concerned with the interaction between xenobioticsdbi l i llldi tlidi tlddithDNAdandbiologicalmoleculesdirectlyorindirectlycodedintheDNA,andcan be regarded as a branch of GENETICS.”Michael F.W. Festing (2001)Michael F.W. Festing (2001)Gregor Mendel (1822 – 1884)TERMINOLOGYTERMINOLOGYGene: A sequence of DNA bases that encodes a proteinAllele:A sequence of DNA basesAllele:A sequence of DNA basesLocus: Physical location of an allele on a chromosomeLinkage:Proximity of two alleles on a chromosomeLinkage:Proximity of two alleles on a chromosomeMarker: An allele of known position on a chromosomeDi tNb fbibt t lllDistance:Number of base-pairs between two allelescentiMorgan: Probabilistic distance of two allelesPhAdbblh(i)Phenotype:An outward, observable character (trait)Genotype: The internally coded, inheritable informationPenetrance: No. with phenotype / No. with alleleModified from M.F. Ramoni, Harvard Medical SchoolThe 80s Revolution and the Human Genome Projectthe Human Genome ProjectGenetic Polymorphisms: naturally occurring DNA markers that id tif i f th d i di id lidentify regions of the genome and vary among individualsThe intuition that polymorphisms could be used as markers sparkled the revolutionthe revolutionOn February 12, 2001 the Human GenomeProject announced the completion of a firstProject announced the completion of a firstdraft of the human genome and declared:“A SNP map promises to revolutionize both ppmapping diseases and tracing human history”SNP are Single Nucleotide Polymorphisms – subtlevariations of the human genome across individualsModified from M.F. Ramoni, Harvard Medical SchoolDISTANCES ON A GENETIC MAPDISTANCES ON A GENETIC MAP• Physical distances between alleles are base-pairsB t th bi ti f i t t t•But the recombination frequency is not constant• A useful measure of distance is based on the probability of recombination: the Morganprobability of recombination: the Morgan• A distance of 1 centiMorgan (cM) between two alleles means that they have 1% chance of being separated by recombination• A genetic distance of 1 cM is roughly equal to a physical distance of 1 million base pairs (1Mb)physical distance of 1 million base pairs (1Mb)Modified from M.F. Ramoni, Harvard Medical SchoolMORE TERMINOLOGYMORE TERMINOLOGYPhysical Maps: maps in base-pairsHuman physical map: 3000Mb (Mega-bases)Human physical map: 3000Mb (Mega-bases)Genetic Maps:maps in centiMorganGenetic Maps:maps in centiMorganHuman Male Map Length: 2851cMHuman Female Map Length: 4296cMHuman Female Map Length: 4296cMCorrespondence between maps:Correspondence between maps:Male cM ~ 1.05 Mb; Female cM ~ 0.88MbModified from M.F. Ramoni, Harvard Medical SchoolSingle Gene(Mendelian) diseases:Simple and Complex TraitsSimple and Complex TraitsSingle Gene (Mendelian) diseases:Autosomal dominant (Huntington)Autosomal recessive (Cystic Fibrosis)(y )X-linked dominant (Rett)X-linked recessive (Lesch-Nyhan)T d 400 i ldi h b id ifi dToday, over 400 single-gene diseases have been identifiedProblem: traits don’t always follow single-gene modelsyggComplex Trait: phenotype/genotype interactionMultiple cause: multiple genes in several loci determine ppga phenotype in conjunction with non-genetic factors (accidents of development, social factors, environment, infections other factors)infections, other factors)Multiple effect: gene causes more than one phenotypeModified from M.F. Ramoni, Harvard Medical SchoolToxicology Toxicology ≈≈ GeneticsGeneticsThere is substantial polymorphism in genes that determine the response to xenobiotics both in humans and animalsThis has important implications for toxicology and pharmacology:• adverse reactions to drugs cause thousands of deaths each year and gymany of those are associated with susceptible phenotypes• are we protecting the most sensitive in human population when occupational/environmental limits of exposure are established?occupational/environmental limits of exposure are established?• how to account for strain differences in susceptibility in animal studies (1000-fold differences have been reported for TCDD LD50in rats)?• genotyping of individuals from a sample of blood DNA is becoming increasingly easy so it is possible to genotype people for loci that are thought tocontrol susceptibility to certain drugs/xenobioticsthought tocontrol susceptibility to certain drugs/xenobioticsAdapted, in part, from M.F.W. Festing, Tox. Lett. 120:293-300 (2001)…loci that are thought to control susceptibility to certain drugs/xenobiotics:certain drugs/xenobiotics:Before we can correctly interpret genotyping results we need to:• gain a much better understanding of the genetics of susceptibility • know the mode of action of xenobioticsProblem: relatively little research is done on the genetics of tibilit d t i l i t i l t bsusceptibility and toxicologists in general seem to be unaware of the extent of genetic variation in responseamong the experimental animals that are being usedProblem: modes of action of an overwhelming majority of established toxic substances are still largely unknown (not even worth mentioning scores ofunknown (not even worth mentioning scores of compounds that are being newly developed)Adapted, in part, from M.F.W. Festing, Tox. Lett. 120:293-300 (2001)Genotype-Phenotype Interactions in Complex Biological SystemsAgeEnvironmentAdapted from: Huang, 2002Genetics in ToxicologyGenetics in ToxicologyPhenotype (e.g., toxic symptoms, cancer)netics”Studying mechanismsGenes that control susceptibility/resistancerward GeStudying mechanismsof actionGenes that control susceptibility/resistance“Forenetics”Genotype (gene knockout, polymorphism, etc.)Studying mechanismseverse GePh tStudying mechanismsof action“RePhenotypeAdapted, in part, from M.F.W. Festing, Tox. Lett. 120:293-300 (2001)“Forward Genetics” and Toxicology“Forward Genetics” and ToxicologyDifferentanimalstrainsnearlyalwaysresponddifferentlytotheDifferentanimalstrainsnearlyalwaysresponddifferentlytothesame agent/dose unless the toxic insult is so dramatic that all theanimals die very quicklyExamples of strain
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