5.2 - Chromosomal Abnormalities

Question 1

Normal human karyotype

Answer 1

• karyotype - chromosome count of an individual
• contains 22 pairs of autosomal chromosomes and one pair of sex chromosomes (allosomes)
• females: two X chromosomes denoted by 46,XX
• males: have both an X and a Y chromosome denoted by 46,XY
• chromosomes karyotyped during metaphase - chromosomes condensed and comprises two chromatids

Question 2

How do you prepare a karyotype?

Answer 2

1. collect 5ml heparinised venous blood (peripheral) - can also use amniotic cells, CVS (chorionic villus sample)
2. isolate white cells
3. culture in presence of phytohaemagglutinin - stimulates T lymphocyte growth/differentiation
4. after 48h, add colchicine - causes mitotic arrest at metaphase
5. place in hypotonic saline
6. place on slide
7. fix and stain with Gisema
8. cut out individual chromosomes and arrange into karyotype

Question 3

What is DNA compaction?

Answer 3

• DNA does not exist as a simple double helix but is compacted around histones and further condensed into chromatin - what we see in karyotype
• proteins bound to the chromatin affect its regulation
• the 3D genome is important

Question 4

What is an ideogram?

Answer 4

• a way of showing a chromosome based on its G-banded architecture
• chromosomes have some common structural features
• Gisema staining leaves a recognisable pattern of bands (light and dark numbered bands)
• p-arm - shorter, top arm
• q-arm - longer, bottom arm

Question 5

How are bands formed and numbered?

Answer 5

• bands are caused by differentially staining - Gisema staining causes some to be darker than others as they take up more of the stain
• bands were originally identified with low level of resolution = only few bands were visible per chromosome e.g. 1, 2, 3
• improved technology = more bands visible which are named as sub-bands e.g. 11, 12, 21, 22, 23 etc
• further improvements = sub-sub-bands e.g. 11.1, 11.2 etc
• improved resolution helped identify smaller aberrations
• number of bands are measured in bphs (bands per haploid set)
• bands do not represent genes or families of genes
• there are regions of different compaction:
• dark (heterochromatin) = more compact, fewer genes
• light (euchromatin) = more open, more genes
• now this banding is often done during prophase not metaphase because chromosomes are less compact so get more detail from karyotype

Question 6

What is fluorescent staining?

Answer 6

• another staining method where chromosomes are stained fluorescent with a specific marker or individual portions of chromosomes are stained by individual markers

Question 6

Standard nomenclature

Answer 6

• p - short arm
• q - long arm
• pter - tip of short arm
• qter - tip of long arm
• cen - centromere
• del - deletion
• der - derivative chromosome (contains extra material)
• dup - duplication
• ins - insertion
• inv - inversion
• t - translocation
• +/- before chromosome number - gain/loss whole chromosome
• +/- after chromosome number - gain/loss part of chromosome

Question 7

What is aneuploidy?

Answer 7

• abnormal number of chromosomes - having a number that is not 46 or a multiple of it
• why is this an issue? the genome has developed such that having two copies of the chromosome is sufficient
• we can go about missing a single gene, but missing a chromosome means losing a large set of genes
• aneuploidy causes syndromes
• can have too many chromosomes (e.g. trisomy) or insufficient (e.g. monosomy)

Question 8

What is the purpose of meiosis?

Answer 8

• to achieve reduction from diploid (2n=46) to haploid (n=23)
• to ensure genetic variation in the gametes
• enables random assortment of homologues and recombination

Question 9

What is non-disjunction?

Answer 9

• non-disjuncture results in uneven number of chromosomes in daughter cells
• occurs when chromosomes are not split properly between daughter cells and can occur in meiosis I (all daughter cells affected) or meiosis II (half daughter cells affected)
• always results in either +1 or -1 chromosomes - when fertilised results in trisomy or monosomy

Question 10

Sex chromosome aneuploidy

Answer 10

• the most common form of aneuploidy
• affects 1/400 males and 1/650 females
  Why is sex chromosome imbalance tolerated whereas most other chromosome imbalances are not?
• due to X-inactivation of excess X chromosomes - in females, only one X chromosome is active (one is inactivated)
• also due to the low gene content of Y chromosome
  Why if inactivated does abnormal number X/Y have effect?
• both X and Y chromosomes have PAR (pseudo-autosomal region) which still produces genes and does not get inactivated - gives sex chromosomes aneuploidy phenotype

Question 11

Trisomy 21

Answer 11

• second most common form of aneuploidy
• trisomy of chromosome 21 (Down’s syndrome)
• most cases result from maternal non-disjunction
• risk of Down’s syndrome increases as age of mother increases, however 75% of cases born to mothers under 35 - this is because 90% of children are born to mothers of this age

Question 12

Why is there a maternal age effect on aneuploidy?

Answer 12

• due to the vulnerability of oogenesis
• primary oocytes are produced before birth then arrest in prophase I until puberty = finite number of primary oocytes and one primary oocyte –> one ovum
• secondary oocytes are then produced and arrest in metaphase II, and meiosis is only completed if the oocyte is fertilised
• the older the mother, the longer the oocyte has been paused in meiosis = increases chance of non-disjunction due to degradation of factors which hold homologous chromatids together - older you are, more disorganised chromatid arrangement

Question 13

Is there a paternal age effect on aneuploidy?

Answer 13

• in spermatogenesis there isn’t an equivalent to oocyte mitotic arrest
• primary spermatocytes undergo around 23 mitotic divisions per year and potentially accumulate defects
• paternal age is not a risk factor for increased aneuploidy, but does affect a subset of single gene disorders caused by point mutations in FGFR2, FGFR3 and RET including Apert, Crouzon and Pfeiffer syndromes
• thought to be enhanced by ‘selfish spermatogonial selection’ resulting from advantage over neighbouring wild type cells
• paternal smoking is a risk factor for aneuploidy (causes 80% 45X, 46% 47XXY, 100% 47XYY, 8% 47+21)

Question 14

What is the effect of aneuploidy on pregnancy risk?

Answer 14

• aneuploidy causes 5% of still births and 50% of spontaneous abortions
• aneuploidy occurs in 5% of all clinically recognised pregnancies
• trisomy of all chromosomes has been detected prenatally
• most trisomies aren’t compatible with life except 21, 18, 13
• monosomy is very poorly tolerated
• estimated 50% of preimplantation embryos have some degree of aneuploidy which are then lost before implantation/before pregnancy is clinically recognised

Question 15

What is mosaicism?

Answer 15

• presence of two or more populations of cells with different genotypes
• e.g. females are often examples due to X inactivation where one X chromosome is randomly turned off early in development so tissues have different expressions of X chr encoded genes - but not TRUE mosaicism as each cell has same genotype
• mosaicism can arise from two mechanisms:
• non disjuncture during early development
• loss of extra chromosome in early development (if embryo starts of as aneuploid)
• they result in generally milder phenotype and some lethal aneuploidy is survivable if mosaic
• most common mosaic is 46,XX/45,X and 46,XY/45,X - where 1 set of sex chromosomes has been lost in development
• everyone is thought to be mosaic to a certain degree as loss of some genetic material occurs often

Question 16

How is chromosomal crossover a cause for structural chromosome abnormalities?

Answer 16

• occurs in prophase I and increases genetic diversity
• pairs of chromosomes align, chiasma form and crossover occurs - can occur at a single point where large terminal chromosome portions are swapped or in middle of chromosomal material in a double crossover
• 1 to 3 times per chromosome per meiosis
• however sometimes goes wrong

Question 17

What single chromosome abnormalities are there?

Answer 17

Deletion - loss of genes

• can be the result of unequal crossover, or breaks in chromosome during meiosis and loss of material
• can occur at chromosome ends or middle

Duplication -

• most often caused by unequal crossover where material from one chromosome is transferred to another
• 1 chr suffers from deletion and other has duplication insertion of genetic material

Inversions -

• occur in middle of chromosomes or around centromere
• breaks in chromosome, inversion, reinserted piece of DNA
• carriers often unaffected but can cause reproductive problems as children with deletions/insertions born
• paracentric inversion - does not occur in centromere
• pericentric inversion - occurs in centromere

Question 18

What are two chromosome abnormalities?

Answer 18

• genetic material transfer between non-homologous chromosome pairs
• if transfer is unidirectional, insertion occurs
• can also have mutual exchange of material - translocation
• if this is balanced, it does not affect carrier but may cause problems in offspring
• can cause partial trisomy or monosomy e.g. Cri-du-chat syndrome
• can occur in somatic cells

Question 19

Inheritance of chromosomal abnormalities

Answer 19

• many chromosomal abnormalities are de-novo (occur in individual gametes during oogenesis/spermatogenesis and parents not affected by condition)
• e.g. two chromosomes have balanced translocation - child can inherit both normal chromosomes (no issue), or both chromosomes with translocated bits (no issue, balanced translocation carrier themself) but if they inherit extra material from one –> unbalanced = problems

Question 20

Chromosomal deletions

Answer 20

• microscopic - detected easily by microscope e.g. Cri-du-chat syndrome 46,XY,del(5p) - means loss of whole short arm of chr 5
• microdeletion - seen in high resolution banding / require molecular genetic approaches to detect - despite name, large numbers of genes deleted (20+) e.g. Velocardiofacial/DiGeorge syndrome 22q11.2 del
• deletions cause a constellation of symptoms which are characteristic

Question 21

Williams syndrome

Answer 21

• 7q11.23 deletion
• long philtrum, short upturned nose, arched eyebrows, supravalvular aortic stenosis, friendly social personality with lack of social anxiety
• phenotypes caused by imbalance of genes - deletion from one chromosome - which are unrelated apart from their location
• deletion too small to detect using normal karyotyping - use targeted FISH (fluorescent in situ hybridisation)
• lack of elastin on affected chromosome

Question 22

7q11.23 duplication syndrome

Answer 22

• opposite of Williams syndrome
• delayed speech development, autistic behaviour that affects social interaction and communication, dilatation of aorta, flat eyebrows, broad nose and short philtrum
• duplications usually have a milder phenotype than corresponding deletions

Question 23

Robertsonian translocation

Answer 23

• classes of chromosomes - metacentric (short and long arms equal and centromeres in middle of chr), submetacentric (short arm shorter than long arm), acrocentric (short arm shortened down to residual stump)
• affects 1/1000 people
• occurs between acrocentric chromosomes
• can be homologous or non-homologous
• most commonly between 13 and 14, 14 and 15, 14 and 21
• most people show no effects - silent in carriers
• can cause problems in offspring
• loss of both short arms of 2 acrocentric chromosomes and they translocate to produce a derivative chromosome (long arms join to form this, short arms lost as a fragment)