Chromosome Abnormalities Flashcards
What are the difference categories of chromosomal abnormalities?
Numerical - gain or loss of whole chromosomes:
- Polyploidy
–Loss or Gain of whole chromosome set
–Triploidy, (3n) 69
–Tetraploidy, (4n) 92
- Aneuploidy
–Loss or Gain of chromosomes
–Autosomal trisomy, 47
–Sex chromosomes, 45, 47, 48, 49
Structural - partial chromosome gain /loss or rearrangement:
- Deletions
- Duplications
- Translocations
- Inversions
What are the features of a polyploidy karyotype?
Triploidy – 3n (69,XXY, XXX or XYY) - extra copy of every chro
• 1-3% of Conceptions
• 2 sperm cells fertilise a single egg
• Or have a diploid gamete.
• Embryos usually spontaneously abort.
• Triploids seldom survive to term, usually results in spontaneous abortion
What are the features of an aneuploidy karyotype?
One or more individual chromosomes extra or missing from a
euploid/balanced set (46)
• Monosomy - lack of a chromosome:
- Autosomal monosomy lethal in embryogenesis.
- Abort too early to be recognised as a conception
• Trisomy - three copies of a chromosome in a diploid cell
- Lead to early miscarriages.
- Only trisomy 13, 18, 21 survive to term (low density of genes) - not
many important genes in these
• Cancer cells often show extreme aneuploidy (multiple
chromosomal abnormalities)
What are the features of Down Syndrome?
• Autosomal aneuploidy
• Trisomy 21 – 47,XY,+21 Most common numerical abnormality in
newborns.
• Increased risk with increasing maternal age; 70% abort naturally
• Only autosomal trisomy compatible with survival to adulthood
• Cause is non-disjunction in 1st meiotic division (extra chromosome
from father in 15% of cases).
Phenotype:
- Dysmorphic face,
- Developmental delay
- Heart defects
What are the features of Klinefelters?
• Abnormal testicular development fail to produce normal levels of
testosterone - leads to breast growth (~40% )
• No spermatogenesis.
• Tall; long limbs
• Lower IQ (10-15 point reduction)
• Rare extreme forms of Kleinfelter’s syndrome occur with karyotype 48,
XXXY or 49, XXXXY. Individuals severely delayed.
What are the different mechanisms for chromosome abnormalities?
Failure of homologous chromosomes to separate in anaphase I
- So in the second stage of meiosis there is one cell with too many
any one with two few - continues throughout cycle.
Failure of sister chromatids to separate at meiosis II
- gives incorrect gametes. If these are fertilised with a normal gamete you get a trisomic or monosoic cell.
Nondisjunction in meiosis generates gametes with 22 or 24
chromosomes.
Fertilisation with a normal gamete gives a trisomic or monosomic zygote.
What are the different possible structural abnormalities?
Breakage occurs in at least 1 chromosome causing a structural abnormality
• Translocations – 2 different chromosomes break and rejoin incorrectly
• Inversions – 2 breaks in same chromosome; genetic material
inverted
- Insertions (Duplications, triplications etc) – Segment of chromosome inserted = extra genetic material.
- Deletions – Interstitial or terminal piece of chromosome missing
What are the features of Prader Willi Syndrome?
Microdeletion on the paternally derived chromosome 15q11
- 7 genes deleted – heterozygously expressed
Incidence 1/12000 – 1/15 000 live births
- Serious food seeking behaviour
- Obesity
- Learning difficulties
- Behavioural problems
- Normal karyotype
How do chromosome inversions occur?
- Rearrangement where segment of chromosome is reversed end to end.
- Occurs when a single chromosome undergoes breakage and rearrangement within itself
- Paracentric do not include the centromere (both breaks in one arm)
- Pericentric include the centromere (a break in each arm )
- Don’t usually cause abnormalities in carriers if rearrangement is balanced - e.g. inv 9, inv 2 are normal variants. But carriers have increased chance of producing offspring with unbalanced chromosome rearrangements.
What are the features of translocations?
Exchange of genetic material between non-homolgous chromosomes.
Reciprocal/Balanced translocation: Mutual exchange after two breaks; no loss or gain of DNA.
• Carried by 1/500 people
• Usually no clinical consequences.
• Carriers have increased risk of creating gametes with unbalanced chromosome translocations
• 3% of couples with recurrent miscarriages have translocation
• Reciprocal involves any chromosome
•Robertsonian involves acrocentric chromosomes (13, 14, 15, 21, 22)
What is the nature of robertsonian translocations?
Translocation between acrocentric chromosomes - short arms lost; long arms fuse at centromere. Gives a karyotype with only 45 chromosomes
• Results in 45 chromosomes - ( two chromosomes fused)
• Common with chr 13 and 14 - incidence 1 in 1300
• Carriers clinically normal. Risk of unbalanced gametes can lead to miscarriages or genetic disease
•Carriers of Robertsonian translocations involving chr 21 have higher
chance of having offspring with Down syndrome (5%)
When can apparently balanced abnormalities give a phenotype?
Example: 46,XY,t(1;8)(p23.1,q31.1) = 46 chromosomes, translocation between 1 and 8, p arm to q arm. ‘balanced’ translocation in a patient with learning delay, eye disorder, cleft lip/palate
• This is an unbalanced translocation - unequal exchange of
chromosome material resulting in loss or gain of gene(s) (partial
monosomy/trisomy).
• Breakpoint disrupts a gene or separates a gene from its
regulatory element (Could be no material missing but the break point
has disrupted a gene, so in effect you have lost the functionality of the
gene)
• Not detected by karyotyping - submicroscopic
What are the causes of structural abnormalities?
Chromosome breaks occur due to DNA damage (radiation; chemicals, viral infections) or as part of the mechanism of recombination
• Cells have enzyme systems that recognise and repair broken
chromosome ends; (joining two ends together, or capping a broken end
with a telomere).
• Cell cycle checkpoint mechanisms prevent cells with unrepaired
chromosome breaks from entering mitosis.
• If damage is not repaired cell commits suicide (apoptosis)
Structural abnormalities result from:
(i) misrepair of chromosome breaks. Chromosomes with a single
centromere can undergo mitosis, even if structurally abnormal
(ii) malfunction of the recombination system. Meiotic recombination
between mispaired chromosomes, especially in spermatogenesis
What are non-allelic homologous recombination events
between low-copy repeats (LCR-NAHR)?
-Homologous recombination between misaligned repeats
results in unequal cross-over at meiosis
- Get deletions, duplications, inversions
- Mechanism accounts for high incidence of de novo deletions
- Common mechanism in microdeletion microduplication (CNV) genetic disorders
How are chromosome abnormalities detected?
Conventional karyotyping:
resolution >5Mb. Detects gross chromosomal rearrangements.
FISH chromosome painting
Targeted FISH:
Detects smaller structural variants
Resolution >0.5 Mb but need to know where to look
- Eg deletions or substitutions. Targeted only.m
Comparative genome hybridisation Array-CGH:
Oligonucleotide arrays
Higher resolution > 10 kb (>2kb with SNP arrays)
- Very high resolution
Next generation sequencing:
- gold standard
- Detect down to 1 base pair
What kind of tissue samples are are used for chromosome testing?
Peripheral blood lymphocytes – treated with phytohemagglutinin to stimulate cell division.
Skin biopsy – also detect mosaics. Slow to grow.
Chorionic villi –pre-natal diagnosis (10-12 wk gestation). Cells dividing rapidly need no stimulation in culture.
Amniotic fluid - foetal cells (14-20 wk gestation). Slow to grow (~2 wks for analysis).
How is karyotyping performed?
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What are the advantages and disadvantages of Karyotyping?
Advantages:
•whole genome screen
• Can detect any gross abnormality:
- Numerical
- Deletions
- duplications
- inversions
- translocations
Disadvantages:
• Limited resolution > 4 Mb
• Slow, expensive, interpretation requires high degree of skill
What are the advantages and disadvantages of FISH?
Working in metaphase, using probes that fluoresce.
Need to know the underlying cause of the pathogenicity to know where to look - not a genome wide study
Advantages:
• High resolution, cheap, easy to interpret
• Can use a single probe or chromosome-specific cocktail (paint)
• Can use different coloured probes simultaneously
Disadvantages:
• Need to decide in advance which DNA sequence or chromosome to test
Multicolour spectral karyotype (SKY) Paint for each chromosome (mixtures of different paints)
How are microarray CHG performed?
Ubiub
2 channels: compare hybridization level to a common background reference
Loss or gain of chromosome material in patient DNA indicated by fluorescent spots on arrays showing abnormal signal intensity ratios
In the results every dot is a probe and this just shows one chromosome, any peaks formed by dots above the baseline is a duplication and below the 0 line is a deletion.
Resolution depends on the density of probes - high density can detect down to 1 or two kilobases
What are the advantages and disadvantages of array CHG?
Advantages:
• whole genome screen or any targeted chromosome
• resolution depends only on the choice of probes used
• oligonucleotide probes can give resolution ~ 2-10 kb
Disadvantages:
• Can’t detect balanced rearrangements (translocations, inversions)
• Gives no information on the chromosomal location of extra material
Array-CGH is displacing karyotyping as the first-line investigation in
genetic diagnosis
What are the features of 7q11.23 Duplication Syndrome?
• aCGH identifies a de novo duplication • FISH validation with 7q11.23 probe • 28 genes duplicated (over-expressed) • Duplication not as clinically severe as deletion condition Williams Syndrome
Clinical features:
- Deep set eyes;
- Long nose
- Speech
- dyspraxia
- Normal karyotype
How are copy number variants (CNVs) leading to new genetic discoveries?
Normal genomes have a lot of genotypic variation anyway, explains why some of us are more predesposed to certain diseases, but also makes it difficult to determine which differences are disease causing and which are ‘normal variation’
• CNVs are submicroscopic structural variants
• Segment of DNA >50bp present at a variable copy number c.f.
reference genome
• CNVs are deletions, insertions, duplications.
• Abundant in the genome; 100% of populations
• Large CNVs (>1 Mb) less frequent (4% of Finnish study)
• Many overlap with disease genes & cause 8-15% of rare disease
• CNVs are not always associated with disease
• Underly genetic diversity and susceptibility to complex diseases,
(autism and cancer)
Pathological CNVs leading to recurrent and large events
• 17q21.31 Deletion Syndrome (Nat. Genet, 2006)
• 15q24.1 Deletion Syndrome (Hum. Mol. Genet, 2007)
• 17q12 Recurrent Deletion (Am. J. Hum. Genet, 2007)
• 15q13.3 Recurrent Deletion o(Nat. Genet., 2008)
What is the structural variation content in the human genome?
Non-redundant CNV coverage per chromosome.
• Abundant and vary in size (1kb-several Mb)
• 12% of human genome associated with CNVs
• Majority are relatively small <10 kb of sequence
Genomic alterations that involve segments of DNA > 50bp.
Database represents structural variation in healthy controls
But phenotypic documentation limited. Health is not static, status
of participant could change.
DGV does not exclude variations with late-onset phenotypes
Caution using it to predict health outcomes
What is the database of genomic variation?
Comprehensive summary of structural variation in normal human genome.
All variants > 50 bp and <3 Mb (10 Mb for inversions) are included.
Blue duplication, red deletion
Segmental duplication - when there’s a lot you see associated copy number variation due to instability.
DECIPHER V9.2 shows by comparison, CNVs linked to known diseases.
What are the uses of NGS over other genome analysis techniques?
Greater number of smaller-sized CNVs detected by NGS c.f. aCGH
- WGS - comprehensive assessment of CNV (and other SV) but costs limit widespread application. Only picking up the coding sequence - prone to misinterpretation
- WES detects CNVs to the level of an exon but lower confidence.
- Bottleneck – Expertise needed to interpret thousands of SV and translate to clinical environment
Most commonly used NGS technique now is Illumina machinery. Performs massively parallel sequencing to generate millions of short sequences (up to 250 bps)
What is the human genome mapping project?
Started in 1990, the U.S. Human Genome Project was a 13-year effort coordinated by the U.S. Department of Energy and the National Institutes of Health.
The project originally planned to last 15 years, but rapid technological advances accelerated completion date to 2003.
- The sequence is composed of the DNA of 10 to 20 anonymous
‘normal’ individuals across different racial and ethnic groups. - There are > 3 million locations where single-base DNA differences
(Single Nucleotide Polymorphisms) occur in humans. - This information is revolutionising the processes of finding
disease-associated sequences and tracing human history.
Information gained:
• Human genome is ~ 99.5% (not 99.9%) identical in all people
• < 2% of genome codes for genes (~ 25,000 genes)
• Functions are unknown for >50% of discovered genes (but
this is decreasing).
• > 98% of the genome does not code for genes – ‘junk DNA’
• Repetitive regions make the genome unstable with pathological consequences. Important disease mechanism.
• Genome architecture important for genome stability
What was learned from studying 1000 ‘normal’ human genomes?
- Sequencing of multiple human genomes indicates that every human has ~250 CNVs > 10kb
- Most are inherited
- 60% of the larger events >1 Mb are sporadic, not inherited
- 8% of “normal” humans have CNVs of at least 4Mb of DNA corresponding to 10-15 genes; compared to 25% of children with developmental delay - Clearly CNV architecture is a predeposing factor to conditions such as autism.
What did the ‘1000’ Genomes Project 2015 - updated catalogue of human genetic variants - show?
- Map of the genomic differences of 2,504 people
- from 26 populations across Africa, East Asia, South Asia, Europe and the Americas.
- Doubles number of known variant sites to 88 million in human genome. Leads to better understanding of the genetic size, diversity and history of the human population.
- Most SVs occur at low frequency (65% exhibit VAF < 0.2%)
- Rare SVs are typically specific to individual continental groups.
- African populations exhibited greatest genetic diversity. Other populations had some genetic variants found in African populations. Supports theory that Africa’s population is the oldest and from which other populations dispersed around the world.
- Naturally occurring homozygous gene knockouts occur in humans; >200 genes dispensable.
- Catalogue of SVs provides basis for understanding how inherited differences in DNA can contribute to disease risk.
- Useful for personal genomics and individualized medicine – identify a person’s medical predispositions
