Tay Sachs, Cystic Fibrosis, Sickle Cell Anemia and PKU Tay Sachs Disease (also called Hexosaminidase deficiency) Introduction 1. Tay Sachs is a rare condition named after 2 physicians, Tay and Sachs, who first described the disease in 1881 2. Tay Sachs is caused by a deficiency of an enzyme called hexosaminidase A (HEXA) HEXA gene 1. HEXA is encoded by the HEXA gene on chromosome 15 at locus 24.1 (15q24.1) 2. The HEXA gene stretches from base pairs 72,343,436 to 72,376,178. 3. The gene is 32,742 bp in length. 4. There are about 102 million base pairs on chromosome 15 5 chromosome 15 contains 1,428 genes (out of the 20,000 genes in the human genome) What does hexosaminidase (HEXA) do in cells? 1. hexosaminidase is a hydrolytic enzyme found in lysosomes 2. it breaks down glycolipids called gangliosides that are found in neurons a. there are 3 types of gangliosides: GM1, GM2, and GM3 b. gangliosides account for around 6% of the lipid content of plasma membranes of neurons; they are similar in structure to phospholipids 1
GM ganglioside Cause of Tay Sachs Disease 1. Tay Sachs is caused by a deficiency of the active form of hexosaminidase 2. if the enzyme doesn t function properly (due to mutated forms of the HEXA gene), then gangliosides buildup in neurons within the brain and spinal cord to toxic levels 3. the buildup of gangliosides (lipids) in neurons causes them to degenerate and die over time complications occur as brain neurons die 4. people with Tay Sachs disease have 100 to 1000 times more GM2 gangliosides inside brain neurons than a normal person 5. The death of neurons over a several year period as GM2 gangliosides build up causes the symptoms of Tay Sachs disease 4. Mutations in the HEXA gene on chromosome 15 cause the enzyme deficiency a. there are more than 120 mutations of the HEXA gene that cause Tay Sachs disease b. some mutations cripple the enzyme but it still partially functions giving rise to less severe forms of Tay Sachs c. other mutations lead to completely non-functional enzymes that lead to the severe forms of the disease that begin in infancy and lead to death by the age of 6. Tay Sachs is an Autosomal Recessive Condition 1. HexA gene a. A dominant HEXA allele codes for normal enzyme b. a recessive HEXA allele codes for nonfunctional enzyme. 1. This allele arose as a mutant allele. 2. It is recessive since it doesn t affect the phenotype in the heterozygote (Aa) 2. Since kids with Tay Sachs die young, they can t be parents. 3. Parents that have kids with Tay Sachs are Autosomal recessive carriers = Aa a. carrier parents show no symptoms b. Aa x Aa cross produces 3:1 phenotypic ratio where 1 in 4 kids have Tay Sachs disease c. Tay Sachs occurs when a person inherits two recessive alleles of the mutated gene (aa) from heterozygote carries who are normal (Aa) 2
Punnett Square: Aa x Aa offspring have 25% chance of inheriting the condition N alleles in unfertilized egg A a N alleles in sperm A AA Aa a Aa aa Zygote (aa) --> newborn with Tay Sachs 25% of kids born with Tay Sachs 75% of kids are normal, but 50% of kids are carriers Symptoms observed in babies with severe form of Tay Sachs 1. babies appear normal at birth, but deteriorate rapidly by 6 months and the baby is dead between the ages of 2 to 6 years 2. brain seizures: typically last 1-2 min, abnormal firing of brain neurons, muscle twitching, fall over or down, experience sensations (sounds, sights, tastes), loss of skeletal muscle control (weak muscles to paralysis) 3. vision and hearing loss (kids go blind and can t hear) 4. severe mental retardation 5. muscle paralysis kids cannot turn over, sit, or crawl 6. lose the ability to swallow and can t eat without feeding tubes 7. kids eventually die of suffocation since diaphragm stops working so can t inhale 8. less severe forms of Tay Sachs (late onset) allow one to live into adulthood but suffer from muscle weakness, loss of muscle coordination (ataxia), speech problems, and mental illness 3
1278+TATC Mutation of the HEXA gene 1. 1278+TATC is a Frameshift Mutation that leads to a Nonsense Point Mutation a. Frameshift mutation insertions or deletions that shift the reading frame for RNA polymerase. 1. the earlier in the sequence the insertion or deletion occurs, the more altered the protein 2. frameshift mutations typically alter the location of the first stop codon (UAA, UGA, UAG) resulting in an abnormally short polypeptide b. nonsense mutation is a change in nucleotide sequence of a gene that results in an early STOP codon. c. This results in the premature termination of transcription and a polypeptide that is shortened (truncated) 2. 1278+TATC is the most common mutation of the HEXA gene that causes the severe form of the disease (accounts for 80% of cases) 3. this is a 4 base pair insertion (+TATC) that comes after bp 1278 in exon 11 of the HEXA gene on chromosome 15 4. the TATC insertion alters the reading frame of the HEXA gene 5. this mutation leads to an early STOP codon (frameshift mutation that is known as a nonsense mutation since it results in a stop codon) 6. as a result of the STOP codon, the enzyme is only 429 aa s in length rather than the normal 529 aa s and non-functonal Treatment 1. there is no cure Tay Sachs disease 2. even with the best care the kids are dead by age 4 to 6 3. it is not possible to reverse or delay the progression of the disease Prevention 1. Blood tests can determine whether parents are carriers of the recessive allele (Aa) that causes Tay Sachs. Parents may decide not to have children 2. Prenatal Testing (CVS at 10-14 wks, amniocentesis at 15-18 wks) a. if both parents are carriers, then prenatal testing can determine whether the unborn child has inherited both recessive alleles (thus will be born with Tay Sachs). b. Couples may choose to abort 3. Preimplantation Genetic Diagnosis a. treat mother with drugs so that she superovulates a lot of eggs, then harvest 6 to 8 or more of the ovulated eggs from the oviduct and fertilize them with dad s sperm by in vitro techniques b. analyze some cells from the embryo before implantation for the presence of homozygous recessive alleles c. discard embryos with Tay Sachs alleles d. preimplantation diagnoses can be done for cystic fibrosis and sickle cell anemia and other genetic disorders 4
Cystic Fibrosis (CF) 1. autosomal recessive disorder that is lethal 2. What causes Cystic fibrosis (CF) a. CF is caused by mutations in the CFTR gene that lead to defective CFTR proteins in the plasma membranes of epithelial cells b. CFTR stands for cystic fibrosis transmembrane conductance regulator c the CFTR gene that is responsible for cystic fibrosis is on chromosome 7 1. the gene is on the long arm of chromosome 7 at position q31.2 2. the gene runs from base pair 116,907,253 to 117,095,955. It is 188,702 bp in length. 3. the gene has 27 exons and 26 introns c. the normal CFTR protein is 1,480 aa s in length CFTR gene on chromosome 7 at position 31.2 The CFTR protein is a channel protein that controls the flow of H2O and Cl- ions in and out of cells inside the lungs. When the CFTR protein is working correctly, as shown in Panel 1, ions freely flow in and out of the cells. However, when the CFTR protein is malfunctioning as in Panel 2, these ions cannot flow out of the cell due to a blocked channel. This causes cystic fibrosis, characterized by the buildup of thick mucus in the lungs. 3. What does CFTR protein do in the epithelial cells that line the bronchi of the lungs? a. CFTR protein is a membrane ion channel that regulates the flow of Na+, Cl- and water out of epithelial cells b. the channel first moves the negatively charged chloride ions out of epithelial cells into the lumen of the bronchi. 5
1. This is an active process that requires ATP hydrolysis to control the opening and closing of a gate associated with the channel (the gate regulates conductance or movement of ions through the channel). 2. The gate must be open in order for Cl- to move into or out of the cell. 3. Cl- move through the channel when the gate is open by diffusion which almost always favors movement out of the cell 4. CFTR is a membrane channel that acts as a Cl- transport pump c. Na+ then follow the Cl- by electrostatic attraction through the ENaC channel. 1. CFTR is necessary for the proper functioning of the ENaC channel. 2. ENaC epithelial sodium ion channel 3. the ENaC channel is a Na+/K+ ATPase channel d. the movement of Na+ and Cl- to the outside of the cell creates an osmotic gradient that water follows by osmosis 1. the NaCl creates a hypertonic environment outside the cell 2. water osmoses through aquaporins (water channels) in the membrane to the hypertonic NaCl solution outside the cell 4. the water moving out of the epithelial cells by osmosis thins out the mucus that covers the epithelial cells that line lung passageways a. this is normal so that the mucus doesn t get too thick (not hydrated enough) where it would move sluggishly and clog up the airways b. the mucus is part of the mucociliary elevator that cleanses the lungs of debris (bacteria, lint, dust particles). The cilia beat to move mucus and anything stuck to it (e.g., pollen, dust, bacteria) out of the lungs and into the throat where it is swallowed 5. Cystic Fibrosis a. Defective CFTR gene means defective CFTR proteins which means that the Cl- pumps in epithelial cells do not operate properly b. if the chloride transport pump is defective, a thick mucus forms on the outside of the epithelial cells clogging up the airways c. the mucus builds up in passageways of the lungs, pancreas, digestive tract, and other organs 6. Symptoms a. lung infections (bacteria live in the mucus that accumulates in the bonchi) b. difficulty breathing from clogged airways c. a decrease in the production of pancreatic juice that leads to digestive problems and malnutrition (mucus blocks pancreatic ducts that carry pancreatic juice to duodenum of small intestine) 7. if untreated, most kids die by the age of 5. if treated, they may survive to the age of 30 or beyond. 8. the most common point mutation of the CFTR gene a. mutation deletes 3 base pairs within the gene. b. As a result, phenylalanine is not included at the 508th position of the resultant CFTR protein. c. The CFTR protein made by the mutant allele does not fold properly without phe at position 508 doesn t function properly as a Cl- transporter 6
d. The normal protein has 1,480 amino acids, whereas the mutant form has only 1,479. 7
Sickle-Cell Anemia 1. autosomal recessive disorder that affects one out of every 400 African-Americans. It is estimated that one out of every African-American is a heterozygote for the sickle cell allele; the sickle cell gene is on chromosome 11 2. Sickle cell disease is caused by a single amino acid substitution in the hemoglobin protein of red blood cells. a. The protein portion of hemoglobin consists of four polypeptides linked together. There are 2 polypeptides called alpha subunits and two that are called beta subunits. b. In sickle cell disease, the amino acid valine is substituted for the normal glutamic acid at position 6 on the 146 amino acid long beta chains (i.e., val replaces glu) 3. When the oxygen level within affected individual s red cells is low (during exercise or at high altitudes) then the sickle-cell hemoglobin molecules clump so as to deform the cell into a sickle shape (as compared to their normal biconcave shape) 4. Sickled cells my stick together and clog up blood vessels. This blockage deprives the cells downstream of oygen and nutrients. This leads to a variety of symptoms to include pain, brain damage, paralysis, and muscle weakness 5. A simple blood test can determine if a person is a carrier of the defective allele. There is a test to detect the presence of the allele in newborns by analyzing the hemoglobin within red cells. 8
PKU (Phenylketonuria) 1,. Autosomal recessive condition on chromosome 12 2. people with the disorder do not make a functional copy of the enzyme phenylalanine hydroxylase. As a result they don t convert the amino acid phenylalanine to tyrosine 3. Phenylalanine is a common amino acid that comes into the body with the proteins that we eat. 4. Phenylalanine accumulates in the bodies of those with the disorder 5. PKU babies look normal at birth, but if untreated they become irreversibly mentally retarded within a year of birth. 6. The most common mutation that causes PKU is the substitution of tryptophan for the normal amino acid arginine at position 408 of the 451 amino acid long enzyme (phenylalanine hydroxylase) 7. Newborns are screened for PKU at birth by a blood test for abnormally high levels of phenylalanine. If they have PKU, they can receive treatment in the form of a special diet for the rest of their life that prevents most of those from becoming mentally retarded. 9