Genomics and Personalized Medicine

Warfarin and Personalized Medicine

• Normal dose of anti-coagulant drug warfarin can lead to excessive bleeding
• Genotyping of variants can be useful in therapy, especially in personalized medicine, which involves prescribing drugs based on an individual's genotype

DNA Packaging

• DNA is packaged in the nucleus with positively charged histone proteins
• DNA + histones = chromatin
• Octamer of histones (2x H2A, H2B, H3, H4) forms a core with DNA around the outside, called a nucleosome
• Histone H1 links nucleosomes together
• Nucleosomes form a chromatin fiber, which loops out of a central nuclear matrix protein to create individual chromosomes

Human Karyotype

• Human cells have 46 chromosomes (23 from each parent), making them diploid (2n)
• There are 22 pairs of autosomes and 1 pair of sex chromosomes
• Staining chromosomes at Metaphase with Giemsa dye produces alternating light and dark bands, characteristic of each chromosome
• Normal female karyotype is 46 XX, and normal male karyotype is 46 XY
• Chromosome 1 is the largest, and chromosome 22 is the smallest

Organisation of Chromosomes

• After DNA replication, each chromosome consists of two identical chromatids linked together at the centromere
• The arms of the chromosomes are often different lengths, with a short (p) and long (q) arm
• The location of genes can be specified based on the G-banding pattern and the arm of the chromosome, e.g., alpha globin genes: 16p13.3

Mitosis

• Mitosis produces 2 cells with 46 chromosomes (diploid: 2n) that are genetically identical
• Involves one round of DNA replication and one round of chromosomal segregation
• DNA is replicated, and chromosomes are duplicated to create chromatids in prophase
• Chromosomes line up at the center of the spindle in metaphase
• Chromatids are separated to the poles of the cell in anaphase
• Nuclear membrane reforms, and the cell splits in two

Meiosis

• Meiosis produces 4 cells with 23 chromosomes (haploid: n) that are genetically variable
• Involves one round of DNA replication and two rounds of chromosomal segregation
• Steps of meiosis:
  1. Homologous pairs of chromosomes line up and DNA is exchanged by crossing over in prophase I
  2. Homologous pairs of chromosomes line up at the center of the spindle in metaphase I
  3. Chromosomes are separated to the poles of the cell in anaphase I
  4. Sister chromatids stay together; nuclear membrane reforms, and the cell splits in two
  5. Chromosomes line up at the center of the spindle in metaphase II
  6. Chromatids are separated to the poles of the cell in anaphase II
  7. Nuclear membrane reforms, and each cell splits in two – 4 cells in total (n)

X-Chromosome Dosage Compensation

• Genes from both maternal and paternal inherited chromosomes are usually active
• To prevent female cells from having double the amount of X-linked genes being active, there is a mechanism of X-chromosome dosage compensation
• Female cells are 46 XX, but only one of the X chromosomes is active
• This means that, with respect to X-linked genes, the level of expression is identical in male and female cells
• Early in embryology, one of the X chromosomes is inactivated randomly, and a gene on the X-chromosome is transcribed as Xist ncRNA, which triggers the transcriptional silencing of one of the X-chromosomes

Biological Sex Determination

• The presence of the Y chromosome determines the male sex
• Extremely rare mutations have identified the region of the Y chromosome responsible for male sex determination
• Males with 46 XX karyotype may have a translocation of part of the Y to the X chromosome
• Females with a 46 XY karyotype may have a deletion of part of the Y chromosome
• Identification of the SRY gene, which encodes a protein that acts as a testis-determining factor (TDF)

Abnormal Karyotypes

• A complete set of chromosomes is called euploidy, and an irregular number is called aneuploidy
• Aneuploidies are caused by the failure of homologous chromosomes to separate properly during meiosis, known as non-disjunction
• Responsible for 50% of spontaneous abortions, frequently caused by trisomies (3 copies of a chromosome)
• The only trisomies that lead to live births are:
  • Trisomy 13 (Patau syndrome): 47 XX +13 or 47 XY +13
  • Trisomy 18 (Edward's syndrome): 47 XX +18 or 47 XY +18
  • Trisomy 21 (Down syndrome): 47 XX +21 or 47 XY +21

Karyotype Determination

• Amniocentesis is carried out during weeks 15-20 of pregnancy to check for chromosomal abnormalities
• Removal of amniotic fluid, isolation of fetal cells, and quantification of chromosomes
• Risk of having a child with Down syndrome increases from 1:600 at maternal age of 33 to 1:50 at maternal age of 43
• 95% of aneuploid conceptions are due to non-disjunction during oogenesis

Monogenic Disorders

• A single gene variant (mutant) is linked to an inherited disorder
• Modes of inheritance:
  • Autosomal recessive
  • Autosomal dominant
  • X-linked (recessive)

Cystic Fibrosis

• Caused by a mutation of the Cystic fibrosis transmembrane conductance regulator (CFTR) gene
• Locus of the gene is on chromosome 7
• An individual inherits one version of the CFTR gene from each parent
• Sequence variation of a gene is called an allele
• Genome contains extensive gene polymorphism; some of these alleles are non-functional and are called mutant alleles or mutants
• For individuals to develop CF, they must inherit a mutant allele from both parents – it is an autosomal recessive disease

Autosomal Dominant

• Trait is frequent in pedigree
• Individuals are affected in each generation
• Trait affects males and females equally
• Trait is transmitted by either sex
• Parents of individual with phenotype also have the phenotype

Huntington's Disease

• Neurodegenerative disease; gradual cognitive decline
• Caused by mutations at the HD locus
• Normal HD gene has ~28 repeats of the CAG sequence in the gene
• Mutant HD gene has > 36 repeats of the CAG sequence
• This expansion of the polyglutamine region produces a version of the huntingtin protein that aggregates and is neurotoxic
• The inheritance of the disease is autosomal dominant

X-Linked Recessive Inheritance

• Trait is rare in pedigree
• Trait skips a generation
• Trait most frequently in males
• Fathers cannot transmit it to their sons
• Parents of affected individuals do not have the disease

Haemophilia A

• Haemophilia A is a blood clotting disorder and is due to a mutation in the factor VIII gene on the X chromosome
• It has X-linked recessive inheritance
• Female carriers of the mutation have 50% of the normal activity of factor VIII and this is sufficient to maintain normal blood clotting function

This quiz covers the basics of genomics, including DNA packaging and genotyping, as well as its application to personalized medicine and drug therapy.