Genetic Disorders. SCPA 501: General Pathology. Amornrat Naranuntarat Jensen

Genetic Disorders SCPA 501: General Pathology Amornrat Naranuntarat Jensen amornrat.nar@mahidol.ac.th

Human has 46 chromosomes (23 pairs) Sex chromosomes Human genome contains approximately 3 billion base pairs of DNA that contains about 20,000 protein-coding genes

Genetic disorder A disorder caused by an abnormality in an individual's genetic materials Abnormalities can range from a small mutation in a single gene to the addition or subtraction of an entire chromosome or set of chromosomes Genetic abnormalities maybe inherited, acquired during embryogenesis, or acquired later in life www.dirkschweitzer.net

Underlying causes of genetic disorders Mutations in both protein-coding genes and non-coding sequences

Genetic mutations Permanent changes in the DNA Mutations can interfere with protein synthesis at various levels Gene deletions and point mutations involving promoter sequences TFs Promoter Gene Normal gene transcription TFs Promoter X Gene Suppression of gene transcription

Effect of genetic mutations Mutations affecting introns or splice junctions TFs Promoter Gene RNA Exon1 Intron Exon2 Exon3 Abnormal mrna processing mrna Exon1 Exon2 Exon3

Effect of genetic mutations Introduction of a stop codon within an exon mrna Exon1 Exon2 Exon3 mrna Exon1 Exon2 Exon3 Translation Translation (Translational stop) Protein Truncated Protein Stop codon

Effect of genetic mutations Some point mutations in an exon mrna Exon1 Exon2 Exon3 Translation Abnormal Protein

Various types of mutations Point mutations - substitution of a single nucleotide base by a different base - missense mutation VS nonsense mutation

The genetic code for amino acids is a triplet code Image from bio.georgiasouthern.edu

Various types of mutations Point mutations - Substitution of a single nucleotide base by a different base - Missense mutation VS nonsense mutation Frameshift mutations - Insertion or deletion of one or two base pairs Trinucleotide-repeat mutations - Amplification of a sequence of three nucleotides

Point mutation: Premature chain termination in β-thalassemia Kumar et al., Basic Pathology, 7th edition, 2003

Frameshift mutation: Single-base deletion at ABO (glycosyltransferase) locus Kumar et al., Basic Pathology, 7th edition, 2003

Trinucleotide-repeat mutations: CAG repeats in neurological disorders

Underlying causes of genetic disorders Mutations in both protein-coding genes and non-coding sequences Amplification or deletion or translocation of protein-coding genes Polymorphisms Epigenetic alterations

Single nucleotide polymorphisms (SNPs) Brena R.M. et al., Nature Genetics, 2006

Underlying causes of genetic disorders Mutations in both protein-coding genes and non-coding sequences Amplification or deletion or translocation of protein-coding genes Polymorphisms Epigenetic alterations

Epigenetics Study of heritable changes in gene expression that occur without a change in DNA sequence [Wolffe and Matzke, Science, 1999] Images from Wikipedia and www.ccbusm.com

Evidence of epigenetic effect: Identical twins are not so identical Cancer No cancer www.dailymail.co.uk The differences seen in identical twins are not from the differences their DNA sequences but are due to an epigenetic effect

Major epigenetic mechanisms DNA methylation Histone modifications

DNA methylation An addition of a methyl group (CH 3 ) to cytosine base of DNA. DNMT Methylation 5-Methylcytosine DNMT = DNA methyltransferase DNA methylation usually occurs at the carbon-5 position of cytosine in CpG (cysteine-guanine dinucleotide) sites Image modified from: http://student.ccbcmd.edu

CpG sites CpG sites are regions of DNA where a cytosine nucleotide (C) occurs right next to a guanine nucleotide (G) in the linear sequence of DNA bases p = phosphate which links C and G together in DNA

DNA methylation represses transcription 1) Direct effects - Inhibit the binding of transcription factors to their recognition sites - Methyl group of 5-methycytosine extends into the major groove of DNA TF Transcription TF Transcription factors Methylation TF X X

DNA methylation represses transcription 2) Indirect effects - Involves proteins with high affinity for methylated CpGs such as methyl-cpgbinding proteins (MBPs) - Induce the recruitment of protein complexes involved in alterations of gene expression MBP No transcription X Methylation Protein that can repress transcription

DNA Hypomethylation VS DNA Hypermethylation Transcriptionally active Hypomethylation Transcriptionally silence Hypermethylation

Major epigenetic mechanisms DNA methylation Histone modifications

Histones pack DNA into nucleosomes Nucleosome DNA Histone octamer DNA

Histone modifications A reversible post-translational modification Histones have tails extending out of the nucleosome which can be covalently modified There are several types of histone modifications Methylation Acetylation Phosphorylation Ubiquitination Sumoylation Poly(ADP)-ribosylation Histone modifications alter the chromatin structure and/or aid in the recruitment of other proteins involved in gene transcription, DNA replication, and DNA repair

Acetylation of Histones The best studied type of histone modifications Lysine + + + + + + + + + + + + + + + + + + Histones have long tails with positive charges Acetylation alter the electrostatic charge of Acetylation Decetylation the histone decreasing the affinity between histones and DNA

Acetylation status of histones correlates with transcriptional activity + + + + + + + + HATs + HDACs Transcriptional repression Transcriptional activation HATs: Histone Acetyl Transferases HDACs: Histone Deacetylases Image from www.ucsf.edu

Interplay between DNA methylation and histone modification MBP HDAC No transcription X HDAC Methylation Protein represses transcription Histone Deacetylase MBPs can recruit HDAC resulting in histone deacetylation downstream of the methylated CpG sequence

Summary: Epigenetic Mechanisms DNA methylation Hypermethylation --> GENE OFF Hypomethylation --> GENE ON Histone modification Acetylation of histone --> GENE ON Deacetylation of histone --> GENE OFF

Genetic disorders are relatively common It has been estimated that at least 7.6 million children are born with severe genetic or congenital malformations each year The incidence of genetic disorders in infants is 25-60 per 1000! Source: WHO report, 2005

Classification of human genetic disorders 1. Mendelian disorder/ Single-gene disorders/ Monogenic disorders 2. Chromosomal disorders 3. Multifactorial/ multigenic disorders

Mendelian disorders Disorders related to mutations in single genes Mendelian disorders are relatively rare but affecting millions of people worldwide Often have simple and predicable Picture from Topnews.in inheritance

Transmission patterns of single-gene disorders Mutations involving single genes typically follow mendelian patterns of inheritance: 1) Autosomal dominant 2) Autosomal recessive 3) X-linked www.scq.ubc.ca

Transmission pattern of autosomal dominant disorders

Examples of autosomal dominant disorders Autosomal Dominant Disorders System Disorder Nervous Huntington disease Neurofibromatosis Myotonic dystrophy Tuberous sclerosis Urinary Polycystic kidney disease Gastrointestinal Familial polyposis coli Hematopoietic Hereditary spherocytosis von Willebrand disease Skeletal Marfan syndrome [*] Ehlers-Danlos syndrome (some variants) [*] Osteogenesis imperfecta Achondroplasia Metabolic Familial hypercholesterolemia [*] Acute intermittent porphyria Kumar et al., Basic Pathology, 7th edition, 2003

Huntington s disease Neurodegenerative disorder that cells in the basal ganglia, the part of the brain that controls movement, emotion, and cognition (thinking ability) is destroyed Caused by a mutation in the gene encoding for huntingtin. Patient has usually high number of CAG repeated in the coding region of the gene Huntingtin has been shown to be essential for development, however its function is yet unclear http://learn.genetics.utah.edu/ http://edoc.hu- berlin.de/dissertationen/zabel-claus- 2003-01-17/HTML/N100BA.html

Familial hypercholesterolemia The most common mendelian disorders (prevalence: 1 in 500) Caused by a mutation in LDLR gene encoded for LDL receptor protein LDL receptor functions in removing LDL from the blood circulation Patient with familial hypercholesterolemia has abnormally high level of LDL in the blood Kumar et al., Basic Pathology, 8th edition, 2007

The LDL receptor pathway Kumar et al., Basic Pathology, 8th edition, 2007

LDL receptor mutations are found at various points of the LDL receptor pathway Kumar et al., Basic Pathology, 8th edition, 2007

Transmission patterns of single-gene disorders 1) Autosomal dominant 2) Autosomal recessive 3) X-linked

Autosomal recessive disorders The largest group of mendelian disorders The diseases occur only when both alleles of the genes are mutants Disease onset is usually early in life

Transmission pattern of autosomal recessive disorders

Examples of autosomal recessive disorders Autosomal Recessive Disorders System Metabolic Hematopoietic Endocrine Skeletal Nervous Disorder Cystic fibrosis Phenylketonuria Galactosemia Homocystinuria Lysosomal storage diseases 1-Antitrypsin deficiency Wilson disease Hemochromatosis Glycogen storage diseases Sickle cell anemia Thalassemias Congenital adrenal hyperplasia Ehlers-Danlos syndrome (some variants) Alkaptonuria Neurogenic muscular atrophies Kumar et al., Basic Pathology, 8th edition, 2007

Cystic fibrosis Disease that affects the respiratory and digestive systems Mutation of CFTR gene encoding chloride channel essential for production of sweat, mucus, and digestive components More than 1,000 different mutations in the CFTR gene have been identified Patients have salty tasting skin, poor growth, excess mucus production, frequent chest infections and shortness of breath

Sickle cell anemia A disorder that affects red blood cells Caused by a missense mutation of a single nucleotide in a gene encoding for beta subunit of hemoglobin Hemoglobin molecules do not form properly, causing red blood cells to be rigid and have a concave shape http://learn.genetics.utah.edu

Transmission patterns of single-gene disorders 1) Autosomal dominant 2) Autosomal recessive 3) X-linked

X-linked disorders There are relatively small number of x-linked diseases All sex-linked disorders are x-linked Almost all x-linked disorders are recessive Female carrying one mutant x allele usually does not carry a disease due to the presence of another normal allele Affected male dose not pass x-linked trait to his son http://genome.wellcome.ac.uk/doc_wtd020851.html

Examples of X-linked disorders X-Linked Recessive Disorders System Disease Musculoskeletal Duchenne muscular dystrophy Blood Immune Metabolic Nervous Hemophilia A and B Chronic granulomatous disease Glucose-6-phosphate dehydrogenase deficiency Agammaglobulinemia Wiskott-Aldrich syndrome Diabetes insipidus Lesch-Nyhan syndrome Fragile-X syndrome Kumar et al., Basic Pathology, 8th edition, 2007

Hemophilia Disorder that affects ability to control blood clotting Patients with hemophilia have a prolonged bleeding following the injury Caused by mutations in F8 and F9 genes encoding for coagulation factor Mutations in the F8 or F9 gene lead to the production of an abnormal version of coagulation factor VIII or coagulation factor IX, or reduce the amount of one of these proteins www.ynooban.co.cc

Human genetic disorders 1. Mendelian disorders/ Single-gene disorders 2. Chromosomal disorders 3. Complex multigenic disorders

Chromosomal Disorders Approximately 1 of 200 newborns has some form of chromosomal abnormalities

How to read human chromosome Karyotype: an organized profile of a person's chromosomes Karyotype is done by staining the chromosome with a Geimsa dye (G-banding) after the cells have been arrested in metaphase or prometaphase http://learn.genetics.utah.edu/content/begin/traits /scientists/ http://www.nature.com/scitable/topicpage/karyotyp ing-for-chromosomal-abnormalities-298

Cytogenetic terminology Karyotypes are usually described using a shorthand system of notations in a following order: 1) Total number of chromosomes 2) Sex chromosome complement 3) Description of abnormalities in ascending numerical order Can you explain this karyotype? 47,XY,+21 Short arm of a chromosome is designated p Long arm is referred to as q

Cytogenetic terminology: An example Where is the location of Xp21.2?

Chromosomal abnormality 1. Alterations in chromosome number 2. Alterations in chromosome structure

Alterations in chromosome number Nondisjunction occurs when homologues fail to separate during meiosis Trisomy cell has one extra chromosome Monosomy cell has one missing chromosome

Chromosomal disorders: Alterations in chromosome number Examples: Down syndrome (trisomy 21) Klinefelter syndrome (47, XXY) Turner syndrome (Monosomy X)

Down syndrome/ Trisomy 21 Result of an extra copy of chromosome 21 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 Affects 1 in 800 children - the most common chromosomal disorder! 19 20 21 22 23 Patients have characteristic facial features, short stature, heart defects, and short lifespan Maternal age has a strong influence on the incidence of Down syndrome

Klinefelter syndrome (47, XXY) http://64.143.176.9/library/healthguide/enus/support/topic.asp?hwid=zm2596 A disorder that affects only male Male with Klinefelter syndrome have an extra chromosome X At puberty, patients often develop more breast tissue than normal, have a less muscular body, and grow very little facial or body hair

Turner syndrome/ Monosomy X The only viable monosomy in humans!! Found in 1 of every 2,000 to 2,500 newborns Patients have Short stature, do not mature sexually during puberty, and sterile 98% of fetuses die before birth

Alterations in chromosome structure Usually result from chromosomal breakage followed by loss or rearrangement of the chromosome

Types of chromosomal rearrangements Kumar et al., Basic Pathology, 8th edition, 2007

Diseases caused by chromosomal translocations http://en.wikipedia.org

Cri-du-Chat Syndrome Partial deletion of a small arm of chromosome 5 - one of the deleted genes is telomerase reverse transcriptase Severe mental retardation Patients with Cri-du-chat syndrome usually cry like a cat due to abnormal larynx development, have a small head, and unusual facial features

Human genetic disorders 1. Mendelian disorders/ Single-gene disorders 2. Chromosomal disorders 3. Complex multigenic disorders

Complex multigenic disorders/ Multifactorial disorders Result from interactions between mutations in multiple genes and environmental factors (alcohol, pollutants, food, etc.) Environmental influences can increase or decrease the risk of the disease Often cluster in families BUT do not have a clear mendelian pattern of inheritance Low tendency to be inherited compared to single gene disorders

Example of complex multigenic disorders Asthma Diabetes Epilepsy Hypertension Coronary artery disease