Chromosome Abnormalities & Cytogenetics - Notes PDF

Chromosome abnormalities & Cytogenetics Zsolt Fábián M.D., Ph.D., Dr. Habil. 1  Chromosome abnormalities & Cytogenetics Lesson 1 – Numerical chromosome abnormalities Zsolt Fábián M.D., Ph.D., Dr....

Chromosome abnormalities & Cytogenetics Zsolt Fábián M.D., Ph.D., Dr. Habil. 1  Chromosome abnormalities & Cytogenetics Lesson 1 – Numerical chromosome abnormalities Zsolt Fábián M.D., Ph.D., Dr. Habil. 2  Chromosome abnormalities & Cytogenetics Chromosomal abnormalities occur far more commonly than single‐gene defects, however they are usually prenatal lethal common cause of infertility and recurrent miscarriage  50% of first trimester miscarriages  multiple congenital anomalies in approx. 0.5% of newborns most are spontaneous, parents are normal accumulate in cancer cells consist of changes in chromosome number or structure  numerical abnormalities affect the total number of chromosomes  structural abnormalities do not affect the total number of chromosomes but alter the content of individual chromosome(s)  Chromosome abnormalities & Cytogenetics Numerical chromosome abnormalities affect the total number of chromosomes an individual carries per cell could result in Polyploidy (affects ~1% of conceptuses)  Changes in the number of complete sets of chromosomes  Affects all chromosomes equally  Examples:  Triploidy (3n), three full copies of all chromosomes  Tetraploidy (4n), four full copies of all chromosomes could result in Aneuploidy (affects ~1 in 300 newborns)  Changes in the number of one or more chromosomes  Chromosomes numbers are unequal  Examples:  Monosomy, 1 copy of a single chromosome (e.g. Turner syndrome)  Trisomy, 3 copies of a single chromosome (e.g. Down syndrome)  Tetrasomy, 4 copies of a single chromosome  Chromosome abnormalities & Cytogenetics Polyploidies Triploidy (A, B & C) ‐ incompatible with life Most commonly due to fertilization of an ovum by two spermatozoa Tetraploidy (D) ‐ incompatible with life Due to failure of first mitotic division after fertilization  Chromosome abnormalities & Cytogenetics Numerical chromosome abnormalities most numerical abnormalities of somatic chromosomes are incompatible with human life exceptions are trisomies of 13, 18 and 21 numerical abnormalities of the sex chromosomes are more likely to be viable  Chromosome abnormalities & Cytogenetics Aneuploidies Monosomy and Trisomy are caused by nondisjunctions during meiosis in one parent. Both monosomy and trisomy can be caused by non‐disjunctions in either M1 or M2. Look closely at the different outcomes for Trisomies caused by M1 vs M2 nondisjunctions. In trisomy there are 3 copies of a chromosome instead of the usual 2. Two of the copies come from the parent in whose meiosis the nondisjunction occurred. Remember the parent has 2 different copies of that chromosome (one paternal, one maternal). If the nondisjunction occurs during M1 in the parent, then the offspring will receive one copy of each of the parent’s two chromosomes (ie. one copy of the parent’s paternal and one copy of the parent’s maternal chromosome). If the nondisjunction occurs during M2 in the parent, then the offspring will receive two identical copies of one of the parent’s chromosomes (ie. two copies of the parent’s maternal or two copies of the parent’s paternal chromosome).  Chromosome abnormalities & Cytogenetics Nondisjunctions Nondisjunction can hypothetically occur in either the mother or father with any of the autosomes or the sex chromosomes in trisomies, the offspring inherits 2 copies of a chromosome from one parent, instead of the usual 1  if the two chromosomes from this parent are not identical (ie. one is the parent's maternal chromosome, the other is the paternal chromosome), then the nondisjunction occurred during Meiosis I  if the two chromosomes from this parent are identical (ie. 2 copies of the parents maternal chromosome, or 2 copies of the parents paternal chromosome), then the nondisjunction occurred during Meiosis II in monosomies, it is not possible to determine when the nondisjunction occurred. Both monosomy and trisomy can be caused by non‐disjunctions in either M1 or M2. Look closely at the different outcomes for Trisomies caused by M1 vs M2 nondisjunctions. In trisomy there are 3 copies of a chromosome instead of the usual 2. Two of the copies come from the parent in whose meiosis the nondisjunction occurred. Remember the parent has 2 different copies of that chromosome (one paternal, one maternal). If the nondisjunction occurs during M1 in the parent, then the offspring will receive one copy of each of the parent’s two chromosomes (ie. one copy of the parent’s paternal and one copy of the parent’s maternal chromosome). If the nondisjunction occurs during M2 in the parent, then the offspring will receive two identical copies of one of the parent’s chromosomes (ie. two copies of the parent’s maternal or two copies of the parent’s paternal chromosome).  Chromosome abnormalities & Cytogenetics Nondisjunctions after crossing over In reality the effect of recombination (crossing over) makes it very difficult or impossible to determine whether nondisjunction occurred in meiosis 1 or 2, at least without a detailed map of where all recombination events occurred (which we would virtually never have). So, we are left with trying to make the best guess we can based on the evidence we have available to us. My advice: If you're ever asked to determine in which meiosis a nondisjunction occurred it's best to ignore the effect of recombination.  Chromosome abnormalities & Cytogenetics Nondisjunctions Nondisjunction most often occurs in maternal M1  Chromosome abnormalities & Cytogenetics Lesson 2 – Structural chromosome abnormalities Zsolt Fábián M.D., Ph.D., Dr. Habil. 11  Chromosome abnormalities & Cytogenetics Structural chromosome abnormalities Change the internal structure of one or more chromosomes, but do not change the total number of chromosomes Balanced abnormalities  A chromosomal rearrangement of without any gains or losses of genetic material  Usually, no phenotypic effect Except in rare cases where the chromosomal breakpoint falls in an important gene.  e.g. Inversions, Translocations Unbalanced abnormalities  A chromosomal rearrangement that results in gain or loss of genetic material  Almost always have phenotypic effects  e.g. Deletions, Duplications Numerical abnormalities are always unbalanced. Structural abnormalities may or may not be unbalanced depending on the type. Individuals with balanced rearrangements are usually phenotypically normal. There are exceptions we will see in the future. e.g. The most common mutation causing Hemophilia A is an inversion that disrupts the F8 gene. e.g. Cases of Female Duchenne Muscular Dystrophy are caused by a translocation between the X chromosome and an autosome. Unbalanced abnormalities almost always result in phenotypes. Syndromes caused by chromosomal deletions tend to be more severe than those caused by duplications, suggesting a loss of genetic material is more disruptive than a gain. There are many examples of deletion syndromes we will see in the future: e.g. William’s Syndrome, DiGeorge Syndrome, Angelman Syndrome, Prader Willi Syndrome There are fewer examples of duplication syndromes. One we will see is WBSCR  Chromosome abnormalities & Cytogenetics Structural chromosome abnormalities Balanced Unbalanced Numerical Numerical abnormalities are always unbalanced. Structural abnormalities may or may not be unbalanced depending on the type. Individuals with balanced rearrangements are usually phenotypically normal. There are exceptions we will see in the future. e.g. The most common mutation causing Hemophilia A is an inversion that disrupts the F8 gene. e.g. Cases of Female Duchenne Muscular Dystrophy are caused by a translocation between the X chromosome and an autosome. Unbalanced abnormalities almost always result in phenotypes. Syndromes caused by chromosomal deletions tend to be more severe than those caused by duplications, suggesting a loss of genetic material is more disruptive than a gain. There are many examples of deletion syndromes we will see in the future: e.g. William’s Syndrome, DiGeorge Syndrome, Angelman Syndrome, Prader Willi Syndrome There are fewer examples of duplication syndromes. One we will see is WBSCR  Chromosome abnormalities & Cytogenetics Unequal crossing over Occurs when repeat sequences misalign during Prophase I of Meiosis I in a parent Correct alignment during Meiosis I Genes maternal Genes Repeats paternal Misalignment of repeats during Meiosis I Genes maternal Genes paternal Genes Genes Duplication maternal Deletion paternal Unequal crossing over occurs during Prophase I of Meiosis I in either one of the parents. Recall chromosomes exist as bivalents during Prophase I. Bivalents consist of 2 homologs with 2 sister chromatids each. Any of the 4 DNA strands can undergo unequal crossing over (ie. between sister chromatids on the same homolog, or one sister chromatid on each of the two different homologs). During formation of the bivalent the homologs align due to homologous pairing which depends on sequence similarity. Since repeats have similar sequences it is possible they can misalign in the bivalent – as shown in the bottom half of the figure – Nb. The second red repeat on one strand is aligning with the first blue repeat on the other strand – this is a misalignment. Note when unequal crossing over occurs as shown here, both a chromosome with a deletion and a chromosome with a duplication will be generated. Thus, for every deletion syndrome there will be a corresponding duplication syndrome. Deletion syndromes are diagnosed far more often, likely because they result in more serious disease. Inversions and translocations can also occur by this mechanism however this isn’t shown here. Inversions occur due to misalignments of inverted repeats in the same chromatid arm (see hemophilia A later) Translocations occur due to misalignments of repeats on different chromosomes 14 Chromosome abnormalities & Cytogenetics Lesson 3 – Clinical cytogenetics Zsolt Fábián M.D., Ph.D., Dr. Habil. 15  Chromosome abnormalities & Cytogenetics Cytogenetics is the study of human chromosomes, their structure, inheritance and abnormalities acrocentric chromosomes in humans are chr. 13, 14, 15, 21 and 22 Until the 1970s chromosomes were identified by size and the position of the centromere as above Now, chromosomes are identified by staining techniques such as G‐banding The classification as meta, submeta, acro, telo‐ is still in use descriptively, but has been supplanted by staining for identification purposes. Telocentric chromosomes are not found in humans – the centromere is virtually at the p arm telomere  Chromosome abnormalities & Cytogenetics Karyotype Karyotype An individual’s complement of chromosomes Also describes a display of a persons chromosomes G‐banding Chromosomal proteins digested Stained with Giemsa Pattern of light and dark bands Light bands – Less condensed euchromatin Dark bands – More condensed heterochromatin G banding is the traditional way of staining chromosomal bands. There are several different staining methods that are beyond the scope of this lecture. More modern methods use fluorescent dyes. Q‐banding for example uses quinacrine, a fluorescent dye that stains DNA in a similar manner to Giemsa. We will discuss spectral karyotyping later when we look at Fluorescence In‐Situ Hybridization.  Chromosome abnormalities & Cytogenetics Karyotyping Peripheral blood is an ideal source of cells for cytogenetic analysis since it's easily obtainable. Other sources can be used as long as they contain rapidly dividing cells that can be cultured: Eg. Amniotic Fluid and Chorionic Villus (used in prenatal diagnosis – see Block 4 Genetic Counselling) Fibroblast cultures Bone marrow Solid tumors  Chromosome abnormalities & Cytogenetics Ideogram Schematic representation of the karyotype chromosomes are ordered from longest to smallest (1‐22) with X and Y separate p – short (petite) arm q – long arm  Chromosome abnormalities & Cytogenetics Karyotype The karyotype display at the bottom shows one X and one Y chromosome so is from a male. There are 2 copies of all autosomal chromosomes except for chromosome 18 which has three copies. The individual therefore has Edward’s Syndrome.  Chromosome abnormalities & Cytogenetics Nomenclature of chromosome bands regions have been assigned by a convention based on consistent morphological features of the chromosome. both regions and bands are numbered from 1 (nearest the centromere) to furthest 1. Identify arms  p – short (petite) arm  q – long arm 2. Identify region  1, 2, 3 etc. 3. Identify band  1, 2, 3 etc. 4. Identify subbands  1, 2, 3 etc. For example: chr. 1q22.3  reads as: one q two two point three (not 1 q twenty‐two)  means: Chromosome 1, q (long) arm, region 2, band 2, subband 3  Chromosome abnormalities & Cytogenetics Nomenclature of chromosome bands CFTR is the gene mutated in Cystic Fibrosis These chromosomal locations are determined by a committee of experts, you won’t ever be expected to identify the chromosomal region on a chromosome based on the banding pattern. But you should understand how to read the nomenclature. The Read and Donnai textbook describes the Region, Band and Sub‐band differently, as Major Band, Sub‐band and sub‐sub bands, but we will use Region, Band and Sub‐ band for our discussions.  Chromosome abnormalities & Cytogenetics Nomenclature of chromosome bands some terms are descriptive of chromosomal locations or features e.g. p, q, cen, ter others describe structural abnormalities e.g. del, dup, inv, t, /, + or – ish – in situ hybridization  is a cytogenetic technique used to label specific genes or loci i – iso – two copies of the same chromosome arm attached to the centromere normal karyotypes are 46,XX or 46,XY  Chromosome abnormalities & Cytogenetics Examples of structural abnormalities 46,XY,del(22)(q21) a male with 46 chromosomes and – a deletion on chromosome 22 – with a breakpoint at band q21 46,XX,inv(7)(p11;q22) a female with 46 chromosomes and – an inversion on chromosome 7 – with breakpoints at bands p11 and q22 47,XX,+21 a female with – an extra copy of chromosome 21, trisomy 21 or Down syndrome 46,XX,t(1;6)(p23;q21) a female with 46 chromosomes and – a translocation between chromosomes 1 and 6 – with breakpoints at band p23 on the short arm of chromosome 1 and at band q21 on the long arm of chromosome 6 45,X/46,XX a mosaic female with some 45,X cells (Turner Syndrome) and some 46,XX cells  Chromosome abnormalities & Cytogenetics Summary 1. Numerical chromosome abnormalities involve alterations in the normal number of chromosomes (46). 2. Polyploidy involves changes in full sets of chromosomes. Triploidy = 3 full copies, 69 chromosomes. Incompatible with life. 3. Aneuploidy involves changes in numbers of 1 or more chromosomes unequally. Trisomy and monosomy, involving a single chromosome are most common, but changes in multiple chromosomes also occur. 4. Aneuploidy is caused by nondisjunctions in a parental meiosis. 5. In a trisomy, if the 2 chromosomes coming from the same parent are identical, the nondisjunction occurred in Meiosis II. If they are different it occurred in Meiosis I. 6. Structural chromosome abnormalities do not change the total number of chromosomes but alter the internal structure of one or more chromosomes. 7. Balanced rearrangements have no gain or loss of genetic material and usually have no phenotypic effect, albeit with rare exceptions. Inversions and translocations are balanced rearrangements. 8. Unbalanced rearrangements involve a gain or loss of genetic material and almost always have phenotypic effects. Deletions and duplications are unbalanced rearrangements. 9. Unequal crossing over is the mechanism that causes most structural abnormalities. It involves misalignment of repeat sequences during a parental meiosis.  Chromosome abnormalities & Cytogenetics Summary II 10. A karyotype is an individual’s complement of chromosomes and also a display of those chromosomes. 11. Chromosomes 13, 14, 15, 21 and 22 are acrocentric and can take part in Robertsonian translocations. 12. Chromosomal band nomenclature is a standard method used to describe chromosomal locations. It will be useful to describe locations of abnormalities. 13. Karyotype nomenclature is a standard method used to describe normal karyotypes and numerical and structural chromosome abnormalities.  Chromosome abnormalities & Cytogenetics Learning objectives 1. Contrast numerical and structural chromosomal abnormalities and describe the mutational mechanisms that result in these abnormalities. 2. Define Polyploidy and Aneuploidy and summarize how these effect human viability. 3. Determine in which parental meiosis a non‐disjunction occurred to cause a trisomy. 4. Describe the major types of structural chromosomal rearrangements. 5. Determine if a chromosomal rearrangement is balanced or unbalanced and predict the phenotypic effect. 6. Describe a karyotype and identify gross abnormalities using a karyotype display. 7. Interpret chromosomal band nomenclature, and karyotype nomenclature to recognize chromosomal abnormalities and use these to predict the resulting clinical condition.