Flipped lesson 3 - Haemoglobinopathies and mutation 23-24.pdf

Haemoglobinopathies and mutation MD210 – GGE – Genetics Flipped lesson 3 1 Essential Learning Outcomes By the end of this lesson you should be able to: • Describe codominant expression of genes with reference to the ABO blood antigen system • Describe the basic genetics of haemoglobin biosynthesi...

Haemoglobinopathies and mutation MD210 – GGE – Genetics Flipped lesson 3 1 Essential Learning Outcomes By the end of this lesson you should be able to: • Describe codominant expression of genes with reference to the ABO blood antigen system • Describe the basic genetics of haemoglobin biosynthesis, how it changes during development and how it is regulated • Describe the genetic mutation that leads to sickle cell anaemia and how it can be diagnosed clinically • Understand why mutations that disrupt the balanced biosynthesis of globin production lead to thalassaemias 2 Blood Group Antigens H Antigen H Locus on Chr 19: “Fucosyltransferase” Responsible for Synthesis of a Sequence of Monomers (Saccharides & Related) on RBC Surface Molecules ABO Group Determined by the ABO Locus Chr 9 Glycosyltransferase 3 Codominance • Two alleles of the same gene which code for proteins with different specific functions are co-expressed (both alleles are expressed completely) in (compound) heterozygote individuals (rare) • Whereas incomplete dominance is a blending of traits, in co-dominance an additional phenotype is produced E.g. the AB Blood group By InvictaHOG - Own work, Public Domain, https://commons.wikimedia.org/w/index.php?curid=1161572 4 ABO Blood Group Antigen 6 Genotypes: Homozygous AA – Group A BB- Group B OO- Group O Heterozygous AO – Group A BO- Group B AB- Group AB (co-dominant expression) 5 Haemoglobin - a tetramer 2 x α globins (141 aa) 2 x non-alpha globins (usually β globins -146 aa) Humans are Diploid = 23 pairs of chromosomes 2 copies (often different alleles) of each gene Hb genes have Biallelic Expression: Both Paternal and Maternal Alleles Are Expressed (αβ)2 (Haemoglobin A) Function – carry O2 6 7 Changes in Globin Synthesis In Embryonic Development https://www.youtube.com/watch?v=vhB0oNLYIqo 8 Naming Haemoglobin Hb F (α2γ2) Foetal Haemoglobin (HbFF) Hb A (α2βA2) Adult Haemoglobin (HbAA) Hb A2 (α2δ2) Haemoglobin A2 (HbA2A2) Hb S (α2 βS2) Haemoglobin S (HbSS) γ α α γ β α α β δ α α δ S α α S 10 Haemoglobin solubility • Free Hb is catabolised and excreted (Renal) • • So packaged in erythrocytes to keep it from being lost Concentration in RBCs: 320-350g/L of cytoplasm • Close to limit of solubility of Hb in physiological solution • • Globin chains (monomers) are not soluble Tetramer is highly soluble • Exceed solubility limit = • • • Polymerisation and Precipitation Distorted RBC shape and impaired function RBC lysis, release of Hb 11 Haemoglobin Biosynthesis Once transcription of globin genes activated lots of Hb is made Critical To Function More or less correct proportion of α and β chains Requires co-ordinated Gene Expression (Safety valve) Protease for Degradation of α chains can correct some excess of α globins But has finite capacity (it can be overloaded) Hb gene expression is coordinated by chromatin restructuring 12 Regulation of Hb synthesis – Locus Control Region Ch #11 Ch #16 13 • • • • Transcriptional Regulation cis-acting sequences Do Not Encode for Peptides (promoters/enhancers/silencers) • cis-acting = acting from the same molecule (intra-molecular action) act on the DNA strand on which they are encoded • • Inherited Abnormalities of Hb • 2 major types of problems, both due to mutation • > 1200 described mutations of the globin genes (> 1200 alleles) • • Haemoglobinopathies (normal quantities of globins that have abnormal sequences) • Causes globin chain polymerisation and misshapen RBCs • e.g. sickle cell disease • • Thalassaemias (normal globin chain sequences but the different chains are not in the correct proportions) • Causes not enough Hb (anaemia) and/or abnormal accumulation of globin subunits (toxic) Sickle Cell Anaemia Caused by mutation in β globin gene sequence Under Conditions of Low Oxygen Tension: Polymerisation of Hb Distortion of RBC Shape and Function Obstruction of Small Blood Vessels Intravascular Haemolysis https://www.dnalc.org/resources/3d/17-sickle-cell.html 16 Sickle Cell – Genotypes CTGAGGAGA (HβA gene) – Beta haemoglobin CTGTGGAGA (HβS gene) – Sickle cell haemoglobin β α α β S α α S Codon for Glutamic Acid Becomes a Codon For Valine MIM ID #603903/ Gene map locus: 11p15.5 1 mutated allele = HbAS – Sickle cell trait 2 mutated alleles = HbSS – Sickle cell anaemia (no normal HbA) 17 Sickle Cell Diagnosis • Haemoglobin Electrophoresis • Based on differing charge of the different Hb tetramers and their differing migration patterns in an Electric Field 18 Thalassaemias The Most Common Genetic Disorders of Hb Inherited conditions characterised by defects in the balanced biosynthesis of normal haemoglobin globin chains RESULTS IN 1. Not enough Hb (anaemia) 2. Abnormal accumulation of Globin subunits 19 Thalassaemias β – thalassaemia Absent or reduced β globin https://www.youtube.com/watch?v=O9mPjGOxEsk α-thalassaemia Absent or reduced α globin https://www.youtube.com/watch?v=jHMfjYIf51w 20 β-Globin Gene: β-Thalassaemia Only one gene for β-globulin - HBB on Chr 11 But 2 copies expressed (one on each chromosome) Chrom #11 2 types of β-thalassaemia: Minor: (heterozygous mutation – one defective gene copy) Major: (homozygous/compound heterozygous mutations – both gene copies defective) Over 700 HBB variants described, problems at: Transcription (promoter) Processing of mRNA Translation of mature mRNA Post translation integrity of β globin 21 β-thalassaemia Minor (β-thal trait) • One copy of Ch11 affected • Heterozygous for defective β globulin expression • Either β0 (absent) or β+ (reduced) • Genotypes: Chrom #11 β/β0 • β/β0 • β/β+ • Clinically asymptomatic or mild symptoms (mild microcytic anaemia) Chrom #11 β/β+ • Small and hypochromic reds 23 β-thalassaemia Major • Both copies of Chr 11 affected • Homozygous for defective β globulin expression • Either β+ (reduced) or β0 (absent) • Genotypes: Chrom #11 • β0/β0 • β+/β+ • β0/β + (compound heterozygote) • Serious illness requiring lifelong transfusions β0/β + Chrom #11 24 α -Globin Genes: α -thalassaemia Two genes for α-globulins – HBα2, HBα1 on Chr 16 But all 4 copies expressed (two on each chromosome) Chr #16 4 types of α-thalassaemia: • Silent carrier: (one defective locus) • α-thalassaemia trait: (two defective loci) • HbH disease: (3 defective loci) • α-thalassaemia major aka HbBart: (4 defective loci) [Hydrops faetalis] 25 α-thalassaemias α+ thalassaemia (one defective locus) • a.k.a. α-thalassaemia minima or silent carrier • Clinically asymptomatic Chrom #16 α0-thalassamia ( 2 defective loci) • • α-thalassaemia minor/trait Often clinically asymptomatic but mild cryptic symptoms (mild hypochromic microcytosis, mild anaemia) Chrom #16 27 α-thalassaemia trait Asian/Mediterranean Populations Commonly deletion of both copies of gene from one Ch16 (cis deletion) with one normal Ch16 African Origin Typically one gene missing from each of the two copies of Ch16 (trans deletions) Chrom #16 Chrom #16 28 Severe α-thalassaemias • HbH disease (3 defective loci ) • Most common in Asian heritage – need one chromosome with no α gene Chrom #16 • Beta chain excess and production of HbH (4 x B-globins) β β • α-thalassaemia major β β • HbBart (4 defective loci ) [Hydrops faetalis] • Hb Bart – no α produced. Get production of γ4 tetramers (Hb Barts) no oxygen released to tissues and foetus dies – most serious form. • Most often gene deletions Chrom #16 γ γ γ γ Things to Remember 1. The AB blood group phenotype is an example of codominant expression 2. Hb is a mixed multimer (tetramer) and Hb genes have biallelic expression . The proportionate /balanced expression of genes for proteins that function as a mixed multimer is important 3. Hb genes on Chr 11 and Chr 16 are located in a 5’ to 3’ direction in the order in which they are expressed during embryonic/fetal life 4. The LCR controls the expression of beta globin genes in cis. It is located 1000s of base pairs upstream (5’) of the globin genes on Chr 11 5. Haemoglobin electrophoresis can be used to identify sickle cell anaemia (HbSS) and sickle cell trait (HbAS) and is based on the different charge and migration distance of Hb tetramers 6. Mutations remote from a structural gene may impact on gene expression and be associated with disease 31