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Genetics of Alcoholism Part II

Genetics of Alcoholism Part II. Ian Gizer University of Missouri-Columbia Columbia, MO, USA gizeri@missouri.edu. Definitions. Chromosomes – threadlike structures on which individual genes are located. Karyotype of normal human male. Chromosome #9.

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Genetics of Alcoholism Part II

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  1. Genetics of Alcoholism Part II Ian Gizer University of Missouri-Columbia Columbia, MO, USA gizeri@missouri.edu

  2. Definitions • Chromosomes – threadlike structures on which individual genes are located Karyotype of normal human male

  3. Chromosome #9 • Locus (location) and allele (alternative form) • Centromere, short (p) and long (q) arms Centromere p q ABO locus (9q34.1)

  4. Definitions • Chromatin: genetic material contained in chromosomes – DNA & proteins (histones and nonhistones) • Euchromatin – less condensed/light bands; coding DNA • Heterochromatin – compacted/dark bands, usually noncoding DNA Chr 21 UM Bauer

  5. Definitions • DNA: Deoxy ribonucleic acid • Purine and pyrimidine bases • Purines: Cytosine, Thymine • Pyrimidines: Adenine, Guanine • Double stranded (each strand has full information content) • Strands are held together by (hydrogen) bonds that form between the nucleotide bases of the DNA molecule Adenine (A) <====> Thymine (T) Guanine (G) <====> Cytosine (C)

  6. Definitions • Gene: A sequence of DNA (a locus on a chromosome) that is involved in (“codes for”) the synthesis of a functional polypeptide (proteins consist of one or more polypeptides, which are strings of amino acids).

  7. Gene Structure EXON – EX-pressed or coding DNA that is converted into protein INTRON – IN-active or noncoding DNA that is not converted to protein

  8. Definitions • Transcription: One of the two DNA strands is transcribed to a single-stranded nucleic acid called ribonucleic acid (RNA) RNA has the same bases as DNA except uracil (U) substitutes for thymine (T). • Translation: Conversion of the basic informational unit of 3 nucleotide bases (called a codon) into a single amino acid.

  9. Example Non-transcribed DNA strand TTT TCC Transcribed DNA strand AAA AGG Transcription mRNA UUU UCC Translation Amino Acid Phenylalanine Serine

  10. Genetic Variation • 95% - 98% of human DNA does not code directly for protein. • An estimated 99.8% - 99.9% of our DNA is common. • But then .1% of 3,000,000,000 = 3 million differences! • We are interested in these variations and the transmission and co-aggregation of these variations with AUDs.

  11. Two major types • Microsatellite/short tandem repeat (STR): a stretch of DNA that is sequentially repeated a variable number of times. • Can cause disease (e.g. CAG repeat expansion causes Huntington’s disease; • Can also be benign variation; • Assume it is close to a disease contributing gene;

  12. Single Nucleotide Polymorphism • SNPs are single base pair changes that occur as natural variation in the human genome. They can code for protein change (non-synonymous) or not.

  13. Two major methods for identifying genes associated with AUDs • Linkage • Association

  14. Linkage Analysis Aa (Bb) AA (BB) Aa (Bb) AA (BB) AA (BB) AA (BB) AA (BB) AA (BB) Aa (Bb) Aa (Bb)

  15. LINKAGE • Basic idea is identity-by-descent (IBD) or how often does an affected pair of relatives share the same ancestral DNA. If more often than expected by chance, then somewhere near this shared DNA is a gene that contributes to affection status. • Need related individuals where multiple relatives are affected. • Identifies large stretches of DNA.

  16. Linkage Analysis: The Basics IBD – An Illustration A. One allele IBS and one allele IBD. B. One allele IBS and zero alleles IBD. C. Two alleles IBS and at least one allele IBD.

  17. IBD Sharing in pairs affected for disorder Sib 1 AC AD BC BD AC AD Sib 2 BC BD 4/16 = 1/4 sibs share BOTH parental alleles IBD = 2 8/16 = 1/2 sibs share ONE parental allele IBD = 1 4/16 = 1/4 sibs share NO parental alleles IBD = 0

  18. LINKAGE via IBD Sib 1 A/B C/D Sib 2 Sib 1 Sib 2 H(0): IBD (0) = 25%; IBD (1) = 50%; IBD (2) = 25% H(A): IBD (0) < 25%; IBD (1) > 50%; IBD (2) > 25% H(A) is evidence for linkage.

  19. Linkage studies of AUDs • Most prominent is Collaborative Study of the Genetics of Alcoholism (COGA). • Has identified many important genetic regions using STRs and SNPs.

  20. COGA strategy 1. Ascertain multiplex alcoholic families Polydiagnostic interview Electrophysiological data 262 Families, 2282 individuals 2. Linkage analyses to identify chromosomal regions  allele-sharing among affecteds within a family 3. Association analyses to identify specific genes Gene A Gene B Gene C

  21. LOD score LOD = Likelihood of Odds; LOD of 3.0 means it is 1000 times more likely than expected by chance that there is linkage. Log101000 = 3 Higher the LOD, more likely genes are nearby Williams et al., 1999

  22. Irish affected sib pair study Prescott et al., 2006

  23. Problems with Linkage • Methodological problems; • Need BIG sets of families; • Home in on a big chunk of DNA – possibility of hundreds of genes!!! 1 cM (centiMorgan) is approximately equal to 1 Megabase or 1000000 bp!!!! Genes may be anywhere in the 50cM region

  24. Association Analysis Cardon & Bell, 2001 Nat Rev Genet

  25. Association • Family Based(transmission disequilibrium test) • How often is the risk allele transmitted to an affected child from a parent who is heterozygous (A/a) for the SNP? A/a a/a a/a A/a A/a aa A/a A/a a/a A/a

  26. Association • Case/Control Design • Is the prevalence of the risk allele greater in affected versus unaffected people? a/a a/a A/a A/a A/a a/a A/a A/a

  27. Which Genes should I look at? • Genes in a linkage region • Genes that metabolize alcohol (candidates) • All genes

  28. Genes in the linkage region GABRA2: gamma-amino butyric acid receptor A, subunit 2 gene

  29. GABAmajor inhibitory neurotransmitter of the central nervous system • GABA & Alcohol (Buck, 1996; Grobin et al., 1998) • motor incoordination • anxiolytic effects • sedation • ethanol preference • withdrawal signs • tolerance & dependence • GABAA receptor agonists tend to potentiate the behavioral effects of alcohol, while GABAA receptor antagonists attenuate these effects

  30. GABRA2 and AUDs • Region contains: • GABRG1 • GABRA2 • GABRA4 • GABRB1 Edenberg et al., 2004

  31. Many replications… • Many studies now show an association between SNPs in GABRA2 and AUDs. • SNPs are also associated with drug dependence, nicotine dependence, conduct problems and antisocial personality disorder – likely to be general vulnerability to thrill seeking. • Replicated in family-based and case-control studies.

  32. Genes that metabolize alcohol ADH cluster(1a,1b,1c,4,5,6,7)

  33. Flushing Response • Dysphoric effects that occur w/i 15 minutes of drinking: • Heart palpitation • Facial reddening • Nausea, dizziness • There are large ethnic group differences in rate of flushing – metabolic not cultural

  34. Pathway of Alcohol Metabolism Alcohol Acetaldehyde Acetate ADH ALDH

  35. ALDH2 Deficiency • ADH1B*2, ADH1C*1 code for protein subunits that have greater enzymatic activity, suggesting faster conversion to acetaldehyde • ALDH2*2 – inactive enzyme, can’t break down acetaldehyde • Causes facial flushing, nausea

  36. ADH1B(2)*2  faster to acetaldehyde ADH1C(3)*1  faster to acetaldehyde ALDH2*2  slower breakdown acetaldehyde PROTECTIVE EFFECTS ADH2*2 less common in alcoholics ADH3*1 less common in alcoholics ALDH2*2 less common in alcoholics

  37. Wall et al. (2001)

  38. Wall et al. (2001)

  39. Edenberg et al., 2006

  40. MacGregor et al., 2009

  41. 3 Alcohol Dependence rs3762894 Withdrawal rs2066702 Severity 2 -Log10p 1 0 ADH5 ADH4 ADH6 ADH1A ADH1C ADH7 ADH1B Gizer et al., 2011

  42. Examine ALL genes • Called GWAS: Genomewide association study; • Saturate the genome with a million SNPs and then test association with each SNP. • Maybe find something new!

  43. Treutlin et al., 2009

  44. Bierut et al., 2010

  45. Problems with association studies • Population stratification (only when using unrelateds) –when an association between a SNP and AUDs is due to ethnic variation in that SNP. • P-values need to be adjusted for testing many markers (e.g. 0.05/#markers tested). • Replication in other samples. • What does the gene/SNP do in the etiology of AUDs?

  46. ENDOPHENOTYPES • Inherited mediators; • Associated with, but not a consequence of, alcoholism; • Transmitted in families of alcoholics; • Present when disorder is not in active phase; • Heritable; • Examples: EEG, P300, Subjective response to alcohol. Irv Gottesman

  47. Why study EEG for AUDs? • EEG (Electro-encephal0grams) of waves suggest that certain EEG activity is associated with risk for AUDs; • EEG is heritable; • In families with AUDs, unaffected relatives of AUD individuals have distinct EEG patterns; • EEG pattern is not modified when an individual goes into recovery; • EEG is an ENDOPHENOTYPE for AUDs

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