Topic 11a: Cytogenic analysis of chromosomes Flashcards
trisomy
Chromosomal abnoramilities account for a large proportion of which conditions/diseases?
- spontaneous abortions,
- congenital anomalies,
- intellectual disabilities,
- infertility
what affects the risks associated with chromosomal imbalances or abnormalities?
- presence of haploinsufficient or triplosensitive genes in deleteted or duplicated segemnts
- the global/ cumulative effect of multiple genes on development
- generally, loss or gain of a while chromosome leads to substantial risk of disease (smaller changes can be ok)
what is the cause of the most common muations seen in humans?
aneuploidy (errors in chromosome segregation)
Name the three well-defined non-mosaic autosomal chromosome aneuploidy compatible with postnatal survival?
- Trisomy 21 (down’s syndrome)
- Trisomy 18 (Edward’s syndrome)
- trisomy 13 (patau syndrome)
why is it that we only see 3 compatible types of aneuploidies?
when involves chromosomes in gene-rich regions, it is incompatible with longterm survival and is frequently associated with pregancy loss
Of the conceptions that abort in the first trimester, what the most common cause?
chromosome anomalies (~50%), where the majority of these abnormalities are trisomies that are negatively selected for in the first trimester)
Triploidy is often the result of what?
noraml ovum fertilized by two sperms or by a diploid sperm
How will the majority of all trisomy 21 pregnancies conceived end?
`
spontaneous abortion
what is the most common genetic cause of moderate intellectual disability?
Down’s syndrome
What is the most frequent karyotype of Down’s syndrome?
47, +21 (95% of all patients)
what is non-dysjunction?
the process by which two homologous chromosomes or 2 sister chromatids migrate to the same pole as opposed to opposite poles during cell division
what are the 3 places that nondyjunction can occur?
- meiosis 1 (chromosome pairs do not seperate)
- meiosis 2 (chromatids do not seperate)
- mitosis (leads to mosaicism)
what is known to be correlated to increased chance of aneuploidy? why do people think we see this?
maternal age: which is linked to loss of cohesions between chromatids
Mechanism of origin of down’s syndrome
Meiotic nondisjunction (most common) but can also be caused by mitotic nondysjunction
when does the nondysjunction event leading to down’s syndrome typically occur?
maternal meiosis (M1) mostly (90%) and other 10% occurs in paternal meiosis
what cytogenic diagnostic techniques can be used to investigate trisomy 21 cases?
- quantitative fluorescent PCR
- chromosome banding
- FISH
- chromosomal microarray
what is QF-PCR?
During capillary electrophoresis, the amplified fluorescent microsatellites fragments of different lengths will migrate at different speed and the fluorescence is measured.
what are the advantages of QF-PCR?
- Fast turn-around-time for reporting (no need to culture)
- Automated
what are limitations of QF-PCR?
it is a targeted approach; allows detection og common trisomies and monosomies only
what is chromosome banding?
Will produce distinctive and reproducible patterns of transverse light and dark bands along the chromosomes
what are advantages of chromosome banding?
- View of all the chromosomes of one cell
- Can study multiple individual cells
- Can detect balanced and unbalanced rearrangements
what are limitations of chromosome banding?
- Limit of resolution around 5 -10 Mb
- Need an actively growing source of cells
What is FISH?
A DNA probe labeled with a fluorescent dye hybridize directly to the metaphase or interphase chromosome. A fluorescent signal identifies the probe on a specific region of the chromosome.
what are the advantages of FISH?
- Higher resolution as compared to G banding
- Can study non-dividing or fixed tissues
- Shorter turn-around time (TAT)
what are limitations of FISH?
Only going to see the region of the genome complementary to the probe
what is chromosomal microarray?
The patient DNA (green) and the control DNA (red) are mixed and allowed to hybridize to their complementary sequences on the array.
Red and green intensities are measured, indicating equivalent dosage between the two genomes (yellow) or a relative gain (green) or loss (red) in the patient sample
what are the advantages of chromosomal microarray?
Can identify copy number gains and losses across the genome and at a higher resolution compared to karyotyping.
what are limitations of chromosomal microarray?
balanced rearrangements cannot be detected
what could explain indiviuals that have down’s syndrome but also 46 chromosomes?
robertsonian translocation
what is a roberstonian translocation?
fusion of the entire long arms of two acrocentric chromosomes at the centromere
which chromosomes can be involved in robertsonian translocations?
all acrocentric chromosomes (i.e. those that have very short p arms that contain many repeating and non-coding DNA): 13, 14, 15, 21, 22
all have same material on p arm
Carriers of balanced Robertsonian translocation are at increased risk of what?
having an unbalanced conception, leading to down’s syndrome for example
what is a way the body tries to “rescue” a robertsonian translocation?
in order to avoid a trisomy, the body will “lose” one homologue which cna lead to uniparental disomy
what is the phenotypic effect of trisomy 21?
The existence of a trisomic state results in the production of 50 percent more of the products of genes present on the chromosome, and these gene dosage effects are, in turn, responsible for the abnormalities of development.
what kind of genetic material is contained in chromosome 21?
repeats (36%), open chromatin (3%), regulatory (12.4%), exons (3.6%)
what are some prenatal ultrasound features of down’s syndrome and when can they be detected?
at 17-18 weeks; abnormal facial features (nose), cardiac anomalies, duodenal atresia, generalized edema
what are some dysmorphic features of down’s syndrome that can be seen at birth?
- hyponia
- intellectual attinment
- brachycephaly
- protruding tongue
what is constitutional mosaicism?
Defined as at least 2 cell lines with different chromosomal complement in a fetoplacental unit derived from a single zygote
what determines the percentage of cells and location with each karyotype in mosaisim?
timing of event and cell selection.
how does down’s syndrome usually present in individuals with mosaisism?
- generally milder features
- but can be quite varied (basically normal to full expression)
this depends on the % cells with trisomy
what are some tissue specific influences in mosaic trisomy 21?
- The level of trisomy in buccal mucosa was significantly correlated with IQ
- Congenital heart defects were significantly correlated with lymphocytes
why is it imporatnt to study individuals with partial trisomy?
they can show what region of chromosome 21 is likely to be responsible for specific components of the Down syndrome phenotype and what regions can be triplicated without causing a phenotype
what region is common in all Down’s syndrome cases?
distal 21q22.13, now known as the down syndrome critical region
which congenital disease is often seen in down’s syndrome patients?
congenital heart disease
what gene was found to eb common between down’s syndrome and congenital heart disease?
DSCAM gene: Down syndrome cell adhesion molecule (present in 21q22.3)
what did animal studies show involving overexpression of genes DSCAM
and COL6A2?
affected the morphology of the heart and resulted in septal defects, supporting involvment of dosage gain in congential heart defects
both genes present in down’s syndrome and CHD
how does down’s syndrome relate to alzheimer’s disease?
Almost all patients with Down syndrome develop neuropathology that is similar to that of Alzheimer’s disease
where is APP gene located?
21q21.3
what was shown when looking at a woman with DS with partial trisomy but without AD?
no triplicate APP gene
are chromosome abnormailites seen in “unselected” newborns, and if so, which are seen?
there are some, but very low incidence, and if there are some, mostly balanced transloactions or X and Y aneuploidies are seen.
what is the first-line test in contexts where we see a newborn (or child) with abnormal development?
Chromosomal microarray (CMA)
we see CVNs in 15-20%
what are copy number variants (CNVs) and which genes are often affected?
are variations in the number of copies of larger segments of the genome, and generally involves genes that are implicated in traits that have a gene dosage effect.
what are the main clinical indications for CMA testing?
- hypotonia
- growth abnormality
- neurologic impairment (intellectual delay, seizures)
- dysmorphic facial features
- cardiovasuclar malformations
- other congenital abnormaities
what are the two major classes of CNVs?
- recurrent
- non-recurrent/idopathic
which CNV class is associated with a clinically defined syndrome?
recurrent
why do we say that some CNVs are recurrent?
- there are certain centromeric and telomeric breakpoints that cluster among different patients (i.e. non-related patients)
- hinting at a predisposition of these breakpoints leading to the same rearrangments
recurrent breakpoints ofte map to which regions? how can this mechanistically explain the origin of recurrent CNVs?
Regions with nearby low copy repeats (LCR), leading to abberent or uneuqal crossing over.
there are two products that occurs in this meiotic non-allelic homolgous recombination event: an allele with a duplication and one with a deletion
recurrent syndromes are also called what?
contigous gene syndromes or genomic syndromes
recurrent syndromes are caused by what?
small deletions or duplications involving few genes at closely adjacent loci
microdeletion or microduplications
are clinical phenotypes similar in patients with the same recurrent syndrome? why?
no, it is quite variable due to variable expressivity and penetrance (even if the natire of the geneic abnormaility is pretty much the same)
What type of syndrome is DiGeorge Syndrome and what genetic defect is seen?
is its a recurrent microdeletion syndrome, where individuals ahve a microdeletion in 22q11.2
what features and symptoms are associated with DiGeorge syndrome?
- bulbous nose
- immunodeficiency
- developental delays
- heart defects
what are the main genetic causes of Prder-Willi syndrome and angelman syndrome?
microdeletion and uniparental disomy
what features/symptoms are associted with Prader-Willi syndrome?
Hypogonadism
Hypotonis
Hyperphagia
the 3 Hs
what features/symptoms are associated with Angelman syndrome?
- ataxia
- severe learning difficulty
- inapproproate laughter
what are the two common breakpoints associted with PWS/AS?
in the 15q11.2q13.3 region:
BP1 to BP3 OR BP2 to BP3
BP1 right next to centromere, BP2 a little further, BP 3 is next to the telomere
how is it that the same deletion cause causes 2 diseases (PWS vs AS)?
A role for genomic imprinting in producing the distinct phenotypes: if deletion in maternal – AS, if deletion in paternal – PWS
why is it that we see more cases of maternal uniparental disomy, leading to PWS, as compared to paternal uniparental disomy leading to AS?
higher rates of trisomy in females (i.e. more frequent nondysjunction)
what makes a CNV non-recurrent?
- when the deletions or duplications may be overlapping but have highly variable breakpoints
- these are “random” breakpoints that may or may not lead to a defined syndrome
often de novo
how do non-recurrent CNV arise?
from non-homologous end joining after a double strand break or recombination between copies of repetitive repeats
case study:
enlargement of cerebral ventrical + CNS malfunction on fetal ultrasound.
amniosentesis: CMA shows small deletion on ch.5q11.2
what does this show and what follow up tests could we do?
This deletion is overlaps with other deletions in CHARGE syndrome, where variable breakpoints are seen but all have an overlaping region – non-recurrent CNV
Follow-up tests should include looking at the parent’s chromosome vis FISH, mapping/probing for that segement
not g-banding since deletion too small
what are some unique chromosomal abnormalities that can lead to disease?
unbalanced segregation
of parental balanced chromosome abnormalities
what risks are associated with parents that are carriers of reciprocal translocation?
Carriers produce gametes in which many of the possible combinations occur:
* generally, carriers have a 40-50% chance of having normal or balanced gametes
* of the abnormal, many different posibilites can occur
what are differences and similarites between X and Y chromosomes?
X-chromosomes: larger (150 Mb) and contains 1000 genes
Y-chromosomes: smaller (23 Mb) and contains fewer genes (including SRY gene located at the top or p arm below the PAR1)
both: common regions called pseudautosomal regions (PAR1 top, and PAR2 bottom) to allow homologous recombination between the two
there is obligatory recombination between which region in X and Y chromosome?
PAR1
what is the function of the sex chromosomes?
to determine biological sex and provide proper sexual development and differentiation
when does the firts genetic step in sex determination occur?
at fertilization, where sex of the zygote is determined by the fertilizing sperm
what is the SRY gene and what is its role?
on the Y becomes active early in development and causes the bipotential gonad to start development into testes
what allowed us to understand the role of the X and Y chromosomes in sex determination and differentiation?
Genetic studies of patient with sex-reversal syndromes or patients that are either genetically female with XY and male with XX.
what is the Lyon hypothesis?
proposed that dosage compensation in mammals occurs by the inactivation of one X chromosome in females
how does X-inactivation in the context of genetics occur?
X inactivation starts at the X inactivation center (XIC) which contains XIST and TSIX: Inactive X expresses XIST, which leads to inactivation. Active X expresses TSIX, which represses XIST.
what is required for x chromosome inactivation?
the presence of at least 2 copies of the XIC on different chromosomes
when does x inactivation occur and is it reversible?
- 3 days after fertilization and is completed by the first week
- is permanent and clonally propagated BUT is reactivated in the oogonia (i.e. cells that undergo meiosis in females)
when does non-random or skewed x inactivation occur?
post-inactivation selection: where there is the elimination of functionally unbalanced chromosomeal cell lines or cells with activation of an x bearing an x-linked mutation
this skewing may be either complete or partial, leading to mosaicism.
this will depend on selective pressure and biological impact
what is Klinfelter syndrome and its genetic cause?
features (generally mild): sparse hair, tall stature, small testes, slight deficit in verbal IQ, infertility
genetic:47, XXY genotype where there is an non-dyjunction event in either paternal M1 or maternal M1/2
what is turner syndrome and common genetic causes?
phenotypic features: heart defects, short stature, ovarian failure, IQ slightly reduced
genotype: generally 45,X, but other common ones are: 46, Xi, 46, Xr or some other deletion or defects in one of the X
Given that one X chromosome is inactivated in normal females, what explains at the genetic level the somatic features observed in Turner syndrome individuals?
some genes (~15%) escape x-inactivation and remain active in both chromosomes
what do we have to look for particularly in Turner’s syndrome (i.e. what is a potential risk factor)?
look for family history of x-linked conditions since there cannot be skewed inactivation due to presence of only 1 “healthy” x-chromosome
what is an isodicentric Y and how is sexual phenotype determiend in this cases?
when the y chromsome has 2 centromeres at either end (does not contain the distal portion of the q arm), which is the most common structural anomaly of Y
The sexual phenotype of patients depends on the position of the breakpoint and the proportion of each cell line in the gonadal tissue
what is the effect of the presence of a dicentric chromosome in cell division?
leads to mitotic instability, where the two centromeres of the same chromosome can bind to opposite microtubules, leading to anaphase lag
what is the DAX1 gene and its role?
DAX1 acts as a dominant-negative regulatory protein that inhibits the transcriptional activity of other nuclear receptors, including the androgen receptor.
Duplications in DAX1 in XY individuals undergo sex reversal and develop as females
