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Unit 3: Genetic Disease
6 days
September 24:
• A molecular disease is when the primary disease causing agent is a mutation
• Occurs when an alteration in DNA causes gene changes in the amount or function of the gene products
• The pathophysiological process is not entirely understood for ANY disease
• Even sickle cell – which is pretty well documented – is not completely characterized– It was the first molecular disease identified
more than 50 years ago
Effects on Protein Function
• Loss of function
• Gain of function
• Acquisition of a novel property
• The expression of a gene at the wrong time (heterochronic expression) or in the wrong place (ectopic expression)
Loss of Function Mutations
• May result from alteration of the gene’s coding, regulatory, or other critical sequences
• Nucleotide substitutions, deletions, insertions, and/or rearrangements
• Include:– Thalassemia (caused by deletion of alpha
globin gene)– Turner Syndrome– Retinoblastoma (deletion in tumor supressor
gene)
• Can have a range if some residual function is retained
Gain of Function Mutations
• More is not necessarily better
• Enhancing the normal function of a gene
• Can be an increase in the abundance of the protein
• Can be an increase in the ability of a protein to perform its normal functions
Enhancing Normal Function
• Include:– Hb Kempsey (increases efficiency of oxygen
binding to Hb and reduces delivery to tissues)– Achondroplasia
Increasing Production
• Increasing synthesis of a protein in cells where it is already normally present
• Include:– Trisomy 21– Charcot Marie Tooth Disease (duplication of a
single gene)– Somatic mutations in cancer cells
• Add to tumor progression but not really to initiation
Novel Property Mutations
• Classic example is sickle cell disease
• Due to an amino acid substitution that has no effect on the ability of sickle hemoglobin to transport oxygen
• Sickle cell hemoglobin chains aggregate when they are deoxygenated and form polymeric fibers that deform the red blood cells
• Not observed with any other Hb mutation
• Rare because most mutations are neutral or detrimental
• Rarely does a mutation introduce a new property that is pathologically significant
Heterochronic or Ectopic Mutations
• Alter regulatory regions of genes
• Oncogenes– Usually promote cell proliferation
Biologically Normal Proteins
• Some mutations can disrupt the formation of biologically normal proteins
• Can disrupt transcription or translation
Hemoglobins
• Make great examples
• Varied mutations = varied disorders
• Called hemoglobinopathies
• Most common single gene diseases in humans
• Cause substantial morbidity
• WHO estimates that more than 5% of the world’s population are carriers
• One of the first protein structures to be decoded
• First disease related gene to be cloned
• Better understood than almost any other genetic disease
• Cast light on evolutionary process– Both at the molecular level– And the population level– Provide model of gene action during
development
September 25:
Structure and Function of Hemoglobin
• Oxygen carrier in all vertebrate red blood cells
• Four subunits– Two alpha chains– Two beta chains
• Each subunit composed of– Polypeptide chain– Globin– Heme (prosthetic group containing iron)
• In normal adult hemoglobin – Hb A – these chains are called α and β
• The four chains are folded together to form a tetramer
• Approximately 64,500 amu
• Abbreviated α2β2
• α chain has 141 amino acids
• β chain has 146 amino acids
• Resemble each other in amino acid sequence (primary structure)
• Also in three dimensional configuration (tertiary structure)
• Provide a lot of insight into how different mutations may cause disease
• Have ‘sensitive’ and ‘nonsensitive’ regions on the protein
Developmental Expression
• The change in the expression during development of the various globin genes
• Also called globin switching
• Classic example of the ordered regulation of developmental gene expression
• Genes in the α and β clusters are arranged in the same transcriptional orientation
• The genes in each cluster are situated in the same order in which they are expressed during development
• Embryonic globin synthesis occurs in the yolk sac
• From the third to eighth weeks of gestation
• At the fifth week the site of hematopoiesis begins to move to the fetal liver
• Hb F (α2γ2) is the predominant hemoglobin throughout fetal life – 70% of hemoglobin at birth– Less than 1% of total ‘adult’ hemoglobin
• β chains can be detected in early gestation
• Their significance only become significant near birth
• By 3 months old almost all hemoglobin is the ‘adult’ type (Hb A)
• Synthesis of the δ chain also continues after birth
• Hb A2 (α2δ2) never accounts for more than 2% of the adult hemoglobin
• Delta and gamma globin cannot compensate for defective beta globin
• Therapies are currently being investigated that would increase the production of these other two globins, and therefore Hb F and Hb A2
The Locus Control Region
• Expression of β globin gene is only partially controlled by the promoter and two enhancers in the immediate flanking DNA
• First discovered in patients who had zero β globin gene cluster expression, but intact genes
The Locus Control Region
• These informative patients were found to have major deletions upstream
• Removal of approximately 20 kb
• This region is called the Locus Control Region (LCR)
• Begins 6 kb upstream of the ε globin gene
• Significance of LCR is threefold:– Patients with deletions fail to express ANY
genes in the beta globin cluster– Components of the LCR are likely essential in
gene therapy for disorders of the beta globin cluster
– Knowledge of the molecular mechanisms that underlie globin switching may make it feasible to up-regulate the expression of other Hb’s
• There are 4 alpha globin and 2 beta globin genes per diploid genome
• This makes mutations in beta globin genes more likely to manifest physiologically
• One mutation affects 50% of the beta globin produced
• Alpha globin mutations only affect 25% of the chains
• Beta globin mutations have no prenatal consequences
• Gamma globin still prevalent at birth
• Alpha defect cause both prenatal and postnatal problems
September 30:
Hemoglobinopathies
• Hereditary disorders of hemoglobin
• Three main categories:– Structural variants (alter globin peptide but not
rate of synthesis)– Thalassemia (decreased synthesis of one or
more of the globin chains)– Hereditary persistence of fetal hemoglobin
(clinically benign, impair the perinatal switch from gamma to beta globin synthesis)
Structural Variants
• Most from point mutations in one of the globin structural genes
• More than 400 abnormal hemoglobins have been described
• More than half are clinically significant
Structural Variants
• Three main classes:– Variants that cause hemolytic anemia (sickle
cell)– Altered oxygen transport (formation of globin
incapable of reversible oxygenation)– Variants that cause thalassemia (reduced
abundance of the globin polypeptide)
Hemolytic Anemias
• Sickle cell hemoglobin (Hb S) was the first abnormal hemoglobin to be detected and is of great clinical importance
• Caused by single nucleotide substitution
• Changes the codon of the sixth amino acid of beta globin from glutamic acid to valine
• Homozygosity causes sickle cell disease
• Serious disorder
• Common in some parts of the world
• Characteristic geographic distribution
• Most common in equatorial Africa
• Less commonly occurs in Mediterranean area and India
• About 1 in 600 African Americans are born with this disease
• May be fatal in early childhood
• Longer survival is becoming more common
• Autosomal recessive
• Heterozygotes have sickle cell trait– Only have issues in situations with low oxygen
pressure (in vitro, airplanes)
Thalassemia
• As a group are the most common human single gene disorder
• Mutations reduce the synthesis or stability of the globin chains
• Comes from ‘sea’ thalassa – indicates the Mediterannean origin of the disease
October 2:
• Mutations in Different Classes of Proteins
Mucopolysaccharidoses
• Polysaccharide chains synthesized by connective tissues
• Normal constituents of many tissues
• Long disaccharide repeating units
Mucopolysaccharidoses
• Heterogeneous group of more than a dozen storage diseases
• Mucopolysaccharides accumulate in lysosomes
• Results from a deficiency in one of the enzymes required for degredation
Mucopolysaccharidoses
• 3 main disorders:– Hunter Syndrome – XR– Hurler Syndrome – AR– Scheie – AR
Hunter Syndrome
• Slow progression
• Corneal clouding
• Skeletal changes
• Hepatosplenomegaly
• Coarse facial features
Hurler Syndrome
• Same as Hunter:– Corneal clouding– Skeletal changes– Hepatosplenomegaly
• Death before 10 years of age
Scheie Syndrome
• Onset after 5 years of age
• Normal intelligence
• Normal life span
• Corneal clouding
• Valvular heart disease
• Genetic Complementation – the ability of a product from one mutant to correct the biochemical defect in another mutant
• Applies to Hunter and Hurler – affect different proteins
• Cells can take up the lysosomal protein that they lack from extracellular fluid
Transport Defects
• Cystic Fibrosis – – Most common fatal AR disorder of Caucasian
children– Lungs and pancreas affected– Males often infertile due to missing vas
deferens
Transport Defects
• The major symptoms of CF are caused by abnormal fluid and electrolyte transport across epithelial membranes
• Demonstrated through salty skin, because the sweat glands cannot reabsorb chloride ions
Structural Protein Disorders
• Duchenne Muscular Dystrophy:– Severe– Untreatable– Relatively common– Relentless clinical deterioration– X-linked
Duchenne Muscular Dystrophy
• Affected boys are normal for the first few years of life
• Develop muscle weakness between 3 and 5 years of age
• Confined to wheelchair by 12
• Usually do not survive past 20
Duchenne Muscular Dystrophy
• Die from respiratory failure
• Or cardiac failure
• 1 in 3,300 live male births
• 1 sperm with DMD mutation produced every 10 seconds
Duchenne Muscular Dystrophy
• Lethal in males
• 1/3 = new mutations
• 2/3 = carrier mother
• Typically women do not have overt symptoms
Duchenne Muscular Dystrophy
• The DMD gene is HUGE– 2,300 kb– 1.5% of the X chromosome
• Structurally complex– 79 exons– 7 tissue specific promoters– Differential splicing
Duchenne Muscular Dystrophy
• Most tissues express at least one structural isoform
• Most prevalent in the osseous tissue, muscle, and the brain– Obvious based on phenotypic symptoms
Duchenne Muscular Dystrophy
• Dystrophin is a structural protein which anchors a large protein complex at the cell membrane
• Carriers can be identified early
• Prenatal diagnosis accurate 95%
Structural Protein Disorders
• Osteogenesis Imperfecta– Collagen structure mutation
• Normal Type I Collagen is the major structural protein of bone and other fibrous tissues
Osteogenesis Imperfecta
• Special format of amino acids with Glycine at the axial position (it is the only one small enough to fit without disrupting the helical structure)
• Mutations that swap this AA are highly disruptive to the shape
Osteogenesis Imperfecta• Type I = mild, brittle bones, no deformity
– Defective production of type I collagen
• Type II = perinatal lethal (within 1 month), severe skeletal abnormalities– Structural defect in type I collagen
• Type III = progressive deforming, fractures, limited growth– Structural defect in type I collagen
• Type IV = mild to moderate bone deformity, short stature, fractures– Structural defect in type I collagen
Osteogenesis Imperfecta
• Typically autosomal dominant
• With Type I only half the number of normal molecules are made
• All Type II come from new mutations
